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RADIATION BIOLOGY
VOLUME I— PART II

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RADIATION BIOLOGY

VOLUME I: HIGH ENERGY RADIATION

Edited by
ALEXANDER HOLLAENDER

Director of Biology Division
Oak Ridge National Laboratory

With the cooperation of

Austin M. Brues Berwind P. Kaufmann

Hermann J. Muller Lauriston S. Taylor

Prepared under the Auspices of the Committee

on Radiation Biology, Division of Biology and Agriculture

National Research Council

National Academy of Sciences

Washington, D.C.

Part II
Chapters 9 to 18

New York Toronto London
McGRAW-HILL BOOK COMPANY, INC.

1954

RADIATION BIOLOGY
VOLUME I— PART II

... • *♦.,

' • Cfopyrigtjgt*, J$54, by the McGraw-Hill Book Company, Inc. Printed in the

United^ Spates of America. All rights reserved. This book, or parts
'tf'thereo*;' may not be reproduced in any form without permission of the

publishers.

Library of Congress Catalog Card Number: 53-6042

THE MAPLE PRESS COMPANY, YORK, PA.

CHAPTER 9

Chromosome Aberrations Induced in Animal Cells
by Ionizing Radiations1

Berwind P. Kaufmann

Department of Genetics, Carnegie Institution of Washington, Cold Spring

Harbor, New York

Introduction. Nature of the induced rearrangements: Methods of diagnosis — Types of
induced chromosomal aberrations. The process of structural rearrangement: The breakage
process. Differences in sensitivity to ionizing radiations: Relative sensitivity of different
organisms — Effect of ploidy — Relative sensitivity of chromosomes in different types of
cells of the same species — Changes in sensitivity of chromosomes in cells of the same type.
Chemical and cytochemical studies. References.

1. INTRODUCTION

The effects on living cells of higher plants and animals of exposure to
ionizing radiations are evidenced by various alterations in the constitu-
tion and behavior of cellular materials. Among the most readily detect-
able reactions are those that modify the form and pattern of association
of chromosomes and the course of their separation in the cycle of mitosis.
Cytologic examination of irradiated cells has disclosed a variety of aber-
rant types, in which the chromosomes were either adherent or clumped,
fragmented or reconstituted, excessive or deficient in number. Many of
these abnormalities were observed in the pioneer studies of the biologic
effects of radiation carried on during the early years of the century (for
example, Bergonie and Tribondeau, 1904, 1906, on rat testes; Krause
and Ziegler, 1906, on various mammalian tissues; Perthes, 1904, P. Hert-
wig, 1911, Payne, 1913, and Holthusen, 1921, on eggs of Ascaris; Koer-
nicke, 1905, and Gager, 1908, on somatic and meiotic cells of plants;
Mohr, 1919, on testes of grasshoppers; Amato, 1911, and Grasnick, 1918,
on cells of amphibia). However, no consistent interpretation of the
nature and sequence of origin of the various types of aberrations was
developed. This was partly because most of the observations were made

1 This manuscript was prepared prior to Apr. 1, 1951. Although a few changes
have been made since then, this essentially represents the literature available at that
time.

627

68910

628 RADIATION BIOLOGY

on sectioned material, in which induced breaks were not easily diagnosed.
Moreover, adequate consideration was not given to the effect on the
induced reaction of the length of time elapsing between irradiation and
fixation of the cell.

Recognizing the need for a more extensive evaluation of these factors,
Alberti and Politzer (1923, 1924) examined entire cells of the corneal
epithelium of salamander larvae, fixed at varying periods of time after
the animals had been exposed to X-ray treatments of different intensities.
The observed types of cellular disturbance were interpreted as revealing
a reaction system that involves (1) a period of "primary effect," begin-
ning shortly after irradiation, characterized by a decline in the frequency
of mitoses and the appearance, especially after treatment with high doses,
of pycnotic nuclei with adherent chromosomes, (2) a subsequent period
of mitotic inactivity, and (3) a period of "secondary effects," character-
ized by abnormal mitoses with fragmented or reconstituted chromosomes,
whose frequency is dependent on the duration of the treatment.

It is now recognized that during the period of "primary" or "physio-
logical" effect the materials of the chromosomes are altered, and the
mitotic mechanism governing the normal orderly distribution of the
chromatids into daughter nuclei is inhibited. In cells that have been
exposed to moderate doses of radiation, the chromosomes in late prophase,
metaphase, or anaphase stages may continue the course of division, but
the mitotic progress of cells in earlier prophases is arrested, and at times
their chromosomes evince regressive changes that suggest a return to
interphasic conditions (see, for example, Carlson, 1940). Because the
onset of new mitoses is inhibited, there occurs shortly after irradiation
a period of mitotic inactivity, in which practically all cells appear to be
in resting stages. If the treatment has not been too intense, the mitotic
rhythm may subsequently be reestablished, whereupon the condensed
chromosomes may reveal various types of induced abnormalities. At
times the delay effected by radiation may be followed by precocious
differentiation of the treated cells (e.g., primary spermatocytes into giant
spermatids, Creighton and Evans, 1941).

When the treatment is more drastic, the chromonematic threads of
early mitotic stages may reveal nodal thickening along their lengths, and
the condensed chromosomes may adhere or clump to form irregular
aggregates of chromatin material, presumably as a consequence of changes
in viscosity of component proteins and nucleic acids. Such alteration
disrupts the normal pattern of chromosome division, at times producing
bizarre mitotic figures (see, for example, Helwig, 1933, White, 1937,
Carlson, 1938a, Carothers, 1940, Creighton and Evans, 1941, and Bishop,
1942, on grasshopper cells; Bauer and LeCalvez, 1944, on eggs of Ascaris;
Tansley, Gray, and Spear, 1948, Duryee, 1949, and Rugh, 1950, on
amphibian cells; Welander et al., 1948, on cells of embryos and larvae of

CHROMOSOME ABERRATIONS IN ANIMALS 629

salmon; Lasnitzki, 1943b, 1948, on avian tissue cultures; Pfuhl and
Kiintz, 1939, on connective tissue cells of rabbits; Roller, 1947, on
normal and malignant cells of man). A few selected illustrations are
presented in Fig. 9-1. Alberti and Politzer realized that pycnotic chro-
mosome masses represent one aspect of cellular necrosis. When alter-
ation of the chromosomes is so complete that individual members of the
set cannot be recognized, the aberrations are not serviceable for quanti-
tative studies of breakage, but are useful, as will be shown presently,
for chemical and cytochemical analysis of the changes effected in the
nucleic acids and proteins that represent the major organic constituents
of chromosomes.

Of greater use in quantitative studies are those "secondary" or
genetic" effects induced by moderate doses of radiation, in which

<<

Fig. 9-1. Abnormal mitoses, with adherent chromosomes resulting from exposure of
cells to ionizing radiations, (a) (b) From grasshopper neuroblasts (Carlson, 1941b);
(c) (d) from malignant cells of man (Roller, 1947).

damage is sufficiently localized to break the chromosomes without impair-
ing permanently the synthetic and reparative processes essential to
mitotic and other vital cellular activities. Under these conditions the
chromosomes may establish new associations by union of their broken
ends. The resulting rearrangements can be detected by examination of
the treated cells or their descendants, or by genetic analysis of individuals
carrying the aberrations. Production of such viable chromosomal
exchanges by X-ray treatment was first reported by Muller (1928a, b)
and Muller and Altenburg (1928, 1930), who designed excellent methods
for detection and preservation of the induced rearrangements. These
furnished a wealth of experimental material, whose analysis during the
second quarter of this century has greatly furthered understanding of
the mechanisms of heredity and evolution. Determination of the fre-
quency of these gross chromosomal aberrations under various experi-
mental conditions has also furnished basic data for a preliminary analysis
of the processes involved in chromosome fragmentation and reconstitu-
tion. The significance of such data in supplying information about the
mode of action of ionizing radiations and the possible control of deleteri-
ous effects has become increasingly apparent in recent years.

630 RADIATION BIOLOGY

Many aspects of such studies have been examined in extensive detail
in a series of reviews, texts, and symposia since the publication of Dug-
gar's "Biological Effects of Radiation" in 1936 [e.g., Timofeeff-Res-
sovsky, 1937; Timofeeff-Ressovsky and Zimmer, 1939; Bauer, 1939c;
Delbriick, 1940; Muller, 1940, 1950b; Fano and Demerec, 1944; Catche-
side, 1945, 1946, 1948; Lavedan, 1945; Gray, 1946; Spear, 1946; Lea,
1946; Giese, 1947; Buzatti-Traverso and Cavalli, 1948; Fano, Caspari,
and Demerec, 1950; Sparrow, 1951; and the numerous contributions to
the 1941 Cold Spring Harbor Symposium on Genes and Chromosomes,
the 1946 London Conference on Certain Aspects of the Action of Radia-
tions on Living Cells (published in 1947), the 1948 Brookhaven Confer-
ence on Biological Applications of Nuclear Physics, the FIAT Review of
German Science from 1939 to 1946 (published in 1948), and the 1948 Oak
Ridge Symposium on Radiation Genetics (published in 1950)].

The present review will deal with chromosomal aberrations induced in
animal cells by ionizing radiations. Separation of animal from plant
materials, even for the sake of description, imposes arbitrary limitations
that are not always advantageous; accordingly, pertinent botanical
literature will be cited when it seems desirable. Designation of chromo-
some aberrations and gene mutations as sharply delimited classes is also
to a large extent arbitrary. Small chromosomal aberrations cannot
always be distinguished from the so-called "point mutations." More-
over, radiation studies have clearly shown that the reaction system
responsible for the characteristic phenotypic expression of a gene may
be profoundly altered by a realignment of parts of chromosomes in the
process of structural rearrangement. It is thus apparent that no sharp
line can be drawn between gene mutations and chromosome aberrations;
but this article will be limited to a consideration of the types of gross
chromosomal alterations that can be detected either by direct cytological
examination or by breeding tests. Even with attention focused on this
restricted segment of a large body of information, it will not be possible to
review all the available experimental evidence, and the discussion will be
concerned primarily with the types of induced aberrations, the methods
used in their diagnosis, and their significance with respect to evaluation
of the processes of chromosome breakage and recombination. The
effects of radiations in retarding mitosis and in modifying the normal
distribution of chromosomes on the spindle are considered in Chap. 11
by Carlson.

2. NATURE OF THE INDUCED REARRANGEMENTS

Various types of cells from many different species of plants and animals
have been irradiated in order to obtain information about the process of
structural rearrangement, but the majority of available data have been

CHROMOSOME ABERRATIONS IN ANIMALS

631

obtained in studies of the effects of treating microspores, or pollen grains,
of plants of the genus Tradescantia, and spermatozoa, or male gametes, of
flies of the genus Drosophila. Analysis has been facilitated by the rela-
tively small number of chromosomes in these two types; in the species of
Tradescantia most commonly used the haploid number is six, and in
D. melanogaster, the most widely studied species of Drosophila, the hap-

DROSOPHILA

MAGO

VIABLE
TYPES

IMAGO

LARVA

PUPA

VIABLE RE-
ARRANGEMENTS

IRRADIATION

DOMINANT
LETHALS

ALL TYPES OF
CHROMOSOMAL
ABERRATIONS

/

ANTHER

NTERPHASE METAPHASE

MICROSPORE

REVEAL DOMINANT
"VISIBLES" TRANSMIT
RECESSIVE MUTATIONS

PISTIL

MATURE
PLANT

TRADESCANTIA
Fig. 9-2. Diagram of development stages in Drosophila and Tradescantia, indicating
sources of materials for cytogenetic studies described in text. (From Kaufmann,
1948a.)

loid number is four (either X, 2, 3, 4 or Y, 2, 3, 4 — the letters indicating
the sex chromosomes, the numerals the autosomes). The techniques
employed in studying these organisms illustrate the methods commonly
used to detect chromosomal aberrations (Fig. 9-2).

2-1. METHODS OF DIAGNOSIS

In studies on Tradescantia the irradiated cells are themselves examined
after an interval of time sufficient to permit the treated chromosomes to
reach the condensed stages, when determination of the number of frag-
ments or chromosome exchanges is feasible. The earlier studies, on both
plant and animal cells, employed this method in basic form; but the

632 RADIATION BIOLOGY

development of smear techniques, especially for spreading and staining
the microspores of plants (Taylor, 1924; Kaufmann, 1927), presented the
opportunity — used advantageously in the work on Tradescantia — of
irradiating large numbers of cells in a known stage of microsporogenesis
and obtaining, by subsequent inspection of the stained metaphases and
anaphases, extensive and comprehensive data on the frequency of induc-
tion of fragments and various types of chromosomal rearrangements.
Diagrams illustrating the kinds of aberrations detected in such studies of
Tradescantia have been presented by Catcheside (1945, 1946, 1948);
Catcheside, Lea, and Thoday (1946a); and Lea (1946).

Comparable although less extensive studies have also been made by
direct examination of irradiated animal cells (for example, the studies of
Carlson, 1938a, 1941b, on neuroblast chromosomes of the grasshopper,
Chortophaga) . The effects of the treatments are detectable as breaks or
lesions along the chromosomes, or as new associations of the breakage
ends (represented diagrammatically in Figs. 9-3 and 9-4). An interpreta-
tion of the mode of origin of the types of breaks designated in these
diagrams as "chromosome," "chromatid," and "isochromatid" will be
presented subsequently.

From these illustrations it is apparent that cytologic examination of
condensed chromosomes at metaphase or anaphase will reveal all types
of induced aberrations, including lethal as well as viable aberrations.
Differentiation of the two classes is desirable in some types of analysis.
It is also desirable to know more about the precise location of breaks and
the complexity of individual rearrangements than can be inferred from
observations of mitotic chromosomes. Studies on Drosophila have been
especially useful in supplying such information.

In the method commonly used for detection of induced chromosomal
rearrangements in Drosophila, gametes are irradiated by treatment of
males, which are then mated with untreated virgin females. Experi-
ments with D. melanog aster by Muller and Settles (1927) and Demerec
and Kaufmann (1941) have indicated that doses of X rays approaching
the limit of tolerance of the adult fly (ca. 5000-10,000 r) do not usually
inactivate the spermatozoa, which fertilize the eggs and participate in
zygote formation. Some of the fertilized eggs fail to hatch, death of the
embryos being attributable in many cases to loss or duplication of sec-
tions of chromosomes in early cleavage mitoses (Sonnenblick, 1940).
The abortive embryos constitute a class of so-called "dominant lethals,"
whose frequency can be determined by counting the number of eggs laid
and the number from which larvae do not emerge.

Larvae hatching from eggs fertilized by irradiated spermatozoa may or
may not carry detectable chromosomal rearrangements. The relative
frequencies of the two classes resulting from any given treatment can be
determined by either cytological or genetical techniques. The cytologic

CHROMOSOME ABERRATIONS IN ANIMALS

633

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RADIATION BIOLOGY

CHROMOSOME ABERRATIONS IN ANIMALS 635

approach involves inspection of the salivary-gland chromosomes of the
first generation (Fi) larval progeny of the irradiated fathers. Since
analysis of a given rearrangement by this method is restricted to the
chromosomes of a single individual, aberrations of special interest cannot
be perpetuated for genetic analysis and subsequent experimental use.
On the other hand, the large size and precise pattern of banding of the
salivary-gland chromosomes (as shown in Figs. 9-5, 9-6) offer unparalleled
opportunities for determining the complexity of a rearrangement and the
positions of the breaks involved in its production — as was first demon-
strated by Painter in 1933. Methods of preparing for cytological exam-
ination the aceto-carmine or aceto-orcein smears from which these photo-
graphs were made are briefly outlined in the "Drosophila Guide," by
Demerec and Kaufmann (1950).

One limitation of the salivary-gland-chromosome method of diagnosis
is the difficulty of detecting rearrangements that are restricted to proximal
heterochromatic regions; these parts of the chromosome have poorly
defined bands, and aggregate to form a so-called " chromocenter " (illus-
trated in Fig. 9-5a). Exchanges involving breaks in proximal hetero-
chromatic regions can sometimes be detected in neuroblast cells of the
larva by the pattern of somatic pairing of the chromosomes. Thus the
cross-shaped configuration shown in the small inset of Fig. 9-5a (cf. Fig.
9-7b), results from the side-by-side association of unaltered second and
third chromosomes, maternal in origin, with second and third chromo-
somes of the paternal set that had exchanged parts (reciprocal transloca-
tion) as a result of irradiation of the spermatozoon. Identification of
intrachromosomal exchanges in neuroblast cells is difficult, but is some-
times possible because of the presence of constrictions, including those
associated with the formation of the nucleoli, that are visible in late
prophase stages (Fig. 9-7a, c, d, e). From these considerations it is
apparent that a comprehensive quantitative study of induced rearrange-
ments should include analysis of both salivary-gland and neuroblast
chromosomes from the same larva. The labor involved in cytological
examination of the neuroblasts is so considerable, however, that they are
rarely utilized for this purpose. Before the advantages of salivary-gland
chromosomes were recognized, cytological studies of genetically detected
rearrangements were made exclusively on chromosomes of neuroblast
cells of larvae or gonial cells of adults (see, for example, Dobzhansky,
1936; Stern, 1931).

Genetic techniques for determination of induced chromosomal ex-
changes were described by Muller and Altenburg (1930) and Dobzhansky
(1929, 1930). Both intra- and interchromosomal rearrangements had
previously been known in Drosophila. Sturtevant (1926) had shown that
a reduction of crossing over in the third chromosome of D. melanogaster
was due to inversion (rotation through 180° as a consequence of breakage

Fig. 9-5. Salivary-gland chromosomes of Drosophila melanogaster. Rearrangements
detected in larval progeny of irradiated fathers, (a) Unaltered complement; chromo-
somes aggregated at their proximal heterochromatic regions to form the chromocenter.
(Insert shows chromosomes of neuroblast cell, not from same individual but photo-
graphed to same scale as salivary-gland chromosomes.) (6) Transposition of section
of right limb of third chromosome, (c) Inversion in X chromosome; inversion loop
surrounds nucleolus, (d) Intercalary duplication (reverse repeat) in left limb of
third chromosome. (Photographs of preparations by the author.)

636

Fig. 9-6. Salivary-gland chromosomes of Drosophila melanogaster. Rearrangements
detected in larval progeny of irradiated fathers, (a) Duplication of terminal section
of left limb of second chromosome, (b) Reciprocal translocation between the Y
chromosome and the right limb of the third chromosome, (c) Reciprocal transloca-
tion between the left limb of the second chromosome and the right limb of the third
chromosome, (d) An inversion-translocation complex involving breaks in 2L, 2R,
and 3L. (Photographs of preparations by the author.)

637

638 RADIATION BIOLOGY

and reinsertion) of a section of that chromosome. This inversion, and
others analyzed by Sturtevant (1931), were found in nature or arose
spontaneously in laboratory cultures. Determination of crossover values
is laborious, and this has acted as a deterrent to study of induced inver-
sions by means of genetic techniques. Spontaneously arising transloca-
tions had also been detected in Drosophila by genetic analysis (Bridges,
1923; Bridges and Morgan, 1923; Stern, 1926, 1929). The method of

Y/2R

(6)

3/4

Id) (e)

Fig. 9-7. Chromosome exchanges as revealed in late prophase or metaphase stages
in neuroblast cells of larvae of Drosophila melanogaster . (a) Reciprocal translocation
between the right limb of the second chromosome and the Y chromosome (2R/Y).
(6) An exchange between the second and third chromosomes; characteristic cross-
shaped configuration results from somatic pairing, (c) A reciprocal translocation
between the second and fourth chromosomes, (d) An exchange between the third
and fourth chromosomes, (e) An exchange between the second and fourth chromo-
somes; the fourth chromosome carrying the translocated tip of the second is repre-
sented in duplicate, in addition to the normal fourth chromosome. Nucleoli stippled.
(Original drawings by the author.)

diagnosis of the induced types is based on the finding that translocations
produce linkages between genes located in different chromosomes that
would normally segregate independently.

In the genetic, as in the cytologic, method for detection of transloca-
tions, males are usually irradiated and mated with untreated virgin
females. If the males are wild type, they may be crossed with females
whose chromosomes carry marking mutant genes. In a typical experi-
ment of this type, outlined in Fig. 9-8, irradiated males having wild-type

CHROMOSOME ABERRATIONS IN ANIMALS

NONIRRADlATED IRRADIATED

639

X

»

Pm;H Pm.Sb Cy.Sb Cy, H

Unrelated wild -type $

+ ; +
Translocation

No trans-
location

))

))

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)>

+ / + : +/ +

Cy/+ ;H/ +

Eggs

Sperms

Cy/+; H/ +

(Crossed with unrelated
+ /+;+/+? gives
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+;+ progeny)

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+ ; +

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))

))

Cy/+; H/+

Cy/+ ; +/+

+ /+: H/ +

+ /+- ; +/ +

Balanced Unbalanced Balanced

and Duplication and Deficiency and

Viable Inviable Viable

Fig. 9-8. Diagram of the genetic method for detecting reciprocal translocations
between the second and third chromosomes in Drosophila melanogaster. Second
chromosomes shown at left in outline, third chromosomes at right in solid color.
The chromosomes with dominant marking genes carry inverted sections which are
not indicated in the diagram. Only crosses involving Curly and Hairless (Cy;H)
are detailed, but similar results are obtainable with Plum and Hairless {Pm;H),
Plum and Stubble {Pm;S,b) and Curly and Stubble (Cy;Sb).

640 RADIATION BIOLOGY

second and third chromosomes (represented by the symbols +; + ) are
mated with females whose second chromosomes carry the dominant
markers Curly (Cy) and Plum (Pm), and the third chromosomes the
dominants Hairless (H) and Stubble (Sb). The heterozygous Fi flies,
which are of four types with respect to the dominant marking genes —
namely, Cy;H, Cy;Sb, Pm;H, and Pm;Sb — are mated individually with
unrelated wild-type flies of the opposite sex. Figure 9-8 represents a
cross between heterozygous Fi males and wild-type females. The pres-
ence or absence of translocations between the second and third chromo-
somes is determined by examination of the F2 cultures. If no transloca-
tion has been induced by irradiation, each of the second and third
chromosomes of the Fi males, whether paternal or maternal in origin,
will carry a normal complement of genes. Independent assortment at
meiosis will yield four types of spermatozoa, with respect to the mutants
under consideration, which will produce, by fertilization of eggs bearing
the wild-type chromosomes, four kinds of F2 progeny in approximately
equal numbers (for example, from the cross Cy ;H d" by + ; + 9 Cy;H,
Cy;-\-, -\~',H, and + ;+ males and females). If, on the other hand, an
induced translocation is present, only the maternally derived second and
third chromosomes will carry an unaltered complement of genes, since
those of paternal origin will have exchanged parts with each other.
Independent assortment at meiosis will yield four types of spermatozoa,
but two of them will carry some genes in duplicate and be deficient for
others (bottom row of Fig. 9-8). Eggs fertilized by such spermatozoa
will not as a rule give rise to viable progeny, although a duplication or
deficiency zygote may occasionally survive to produce an individual
possessing special somatic characteristics. Eggs fertilized by the other
two types of spermatozoa (those carrying the second and third chromo-
somes with the dominant markers, and those carrying the two chromo-
somes that have exchanged parts), both of which transmit a complete
set of genes, will produce viable progeny. The occurrence, in the cross
illustrated, of only two classes of F2 progeny — namely, Cy;H and + ;H —
will thus serve as an index to the induction of a reciprocal translocation.

Translocations may be detected in a similar manner by irradiating
males whose chromosomes carry dominant marking genes, and mating
them with nonirradiated wild-type females. This procedure is described
in detail by Dobzhansky (1936).

Another technique for the detection of translocations involving a par-
ticular chromosome is based on phenotypic modification accompanying
change in position of a specific gene. Thus the cubitus interruptus (ci)
position effect in D. melanogaster, which alters the normal pattern of wing
venation, is caused by a translocation involving the fourth chromosome,
whereby the dominance of the wild-type allele of cubitus interruptus is
weakened (Dubinin and Sidorow, 1934). Using this criterion of assay,

CHROMOSOME ABERRATIONS IN ANIMALS

641

Eberhardt (1939) irradiated flies carrying a normal fourth chromosome
and determined the frequency of translocations from the proportion of
progeny showing interruptions in the cubital vein.

In a more elaborate experiment designed to disclose exchanges among
all four chromosomes of an irradiated spermatozoon, Patterson, Stone,
Bedichek, and Suche (1934) mated irradiated wild-type males with
females having attached-X chromosomes homozygous for the mutant
gene yellow (yy), the second chromosome homozygous for brown (bw),
the third for ebony (e), and the fourth for eyeless (ey). The hetero-
zygous F! males were backcrossed individually to yy; bw, e; ey females.

Fig. 9-9. Diagram illustrating the genetic technique for determining points of ex-
change between chromosomes in Drosophila melanogaster. A translocation-carrying
female heterozygous for a series of genes (left) is crossed to a male free from the
translocation and homozygous for the same genes (right). (From Dobzhansky, in
Duggar's "Biological Effects of Radiation," 1936.)

Since these males carried X chromosomes received from their irradiated
fathers, many different types of reassociation could be detected by
examination of F2 cultures, e.g., X;2, X;3, X;4, 2;3, 2;4, 3;4, X;2;3,
X ;2 ;4, X ;3 ;4, 2 ;3 ;4, and X ;2 ;3 ;4. Some duplication and deficiency types
were also viable and could be detected by criteria that will be indicated
presently. Translocations involving the Y chromosome were not
detected in these experiments because the females that came from eggs
fertilized by Y-bearing spermatozoa were not tested.

When translocations have been diagnosed by such genetic methods,
they can usually be perpetuated in cultures, and the positions of the
breaks involved in the rearrangement can subsequently be determined by
either of twro procedures. The most informative and least laborious is
examination of salivary glands of individuals carrying the translocation
in heterozygous condition. The alternative method, which was the first

642 RADIATION BIOLOGY

to be used, requires determination of crossover values between the
chromosomes involved in a translocation and their homologues. As indi-
cated previously, the apparent linkage between genes located in different
chromosomes in translocations is attributable to the inviability of the
recombination classes carrying duplications and deficiencies. This
apparent linkage can be utilized for determining genetically the loci at
which the chromosomes were broken and reunited. The method was
outlined by Dobzhansky (1936) in the review from which Fig. 9-9 is
reproduced. This represents a translocation between the second (black)
and third (stippled) chromosomes, both of which had been broken near
the middle. Females heterozygous for this translocation, which carried
in their normal homologues the series of genes indicated, were mated
with normal males that carried the same series of marking genes. Cross-
ing over took place in the female between the chromosomes involved in
the translocation and their normal homologues. The strongest linkage
was observed in this case between the genes cu and c, and between st and
pr, indicating that breakage and recombination had occurred between
these loci. The validity of this method of diagnosis has been confirmed
in several instances by parallel cytologic studies.

2-2. TYPES OF INDUCED CHROMOSOMAL ABERRATIONS

Any deviation from the standard pattern in number of chromosomes
or arrangement of their component units may be regarded as an aberra-
tion (Dobzhansky, 1936). Although changes conforming to these speci-
fications regularly occur under natural conditions, the frequency of their
occurrence can be increased enormously by ionizing radiations. The
types of alterations that have been induced in this manner include
duplications and deficiencies affecting individual chromosomes and sets
of chromosomes, and inter- and intrachromosomal rearrangements.

2-2a. Haploids and Polyploids. Diploid individuals normally have
every chromosome represented in duplicate in cells that have not under-
gone meiosis. In such individuals occasional cells, or groups of cells,
may have three, four, or more sets of chromosomes, as a result of failure
of chromatids to separate after their multiplication by normal mitotic
processes or by endomitosis. Such polyploid cells can sometimes be
induced in a diploid organism by irradiation. Makino (1939) detected
tetraploid spermatocytes in testes of adults of Podisma mikado (Acrididae)
developing from irradiated nymphs. The observations, which were
restricted to three individuals, were not in themselves conclusive, since
polyploid cells occur in nature in the testes of some Orthoptera (Mickey,
1942; Ray Chaudhuri and Bose, 1948). However, Creighton and Evans
(1941) reported the formation in Chorthippus of giant spermatids by
direct transformation of primary spermatocytes whose normal course of

CHROMOSOME ABERRATIONS IN ANIMALS 643

development was inhibited by X-ray treatment; and White (1935a, b)
and Carlson (1941b) reported the induction by X rays in Orthoptera of
tetraploid spermatogonial cells containing diplochromosomes (tetraploid
with respect to chromatids but only double with respect to centromeres).
It is suggested that they may have originated in prophase cells that
reverted in phase at the time of treatment, and whose chromosomes
underwent a second doubling as they progressed toward metaphase the
second time.

Individuals of a species in which both sexes are normally diploid may
occasionally have one, three, four, or more sets of chromosomes. For
example, Fankhauser (1945) noted the occurrence of haploid, triploid,
tetraploid, and pentaploid individuals among normal embryos raised
from eggs of diploid salamanders. Triploid females of D. melanogaster
arise spontaneously in normal diploid lines (Morgan, Bridges, and
Sturtevant, 1925). Deviations in the number of sets of chromosomes
may also be induced by ionizing radiations. Thus, in the classic experi-
ments of G. Hertwig (1911, 1927), P. Hertwig (1916, 1924), and Dalcq
and Simon (1932), exposure to radiations inactivated the nuclei of
amphibian eggs, which upon insemination with untreated spermatozoa
produced haploid embryos (androgenesis). In other experiments (cf.
Hertwig, 1911. 1913; G. Hertwig, 1927; and Dalcq and Simon, 1932),
treatment of spermatozoa with doses of radiation that did not impair their
motility or ability to penetrate the egg altered the subsequent behavior of
their chromosomes so that haploid embryos developed with only egg
chromosomes (gynogenesis) . In more recent experiments by Rugh
(1939), treatment of spermatozoa of Rana pipiens with doses of X rays
ranging from 15-10,000 r caused progressive decrease in the frequency of
viable embryos; at 10,000 r only 1.6 per cent hatched. With further
increase in dosage, however, the number of viable embryos increased, so
that at 50,000 r about 90 per cent hatched. These embryos were pre-
sumably gynogenetic haploids (see also Rugh and Exner, 1940).

The method of inactivating a gamete nucleus by irradiation has been
used by A. R. Whiting (1946) to secure androgenetic males in the wasp
Habrobracon juglandis. Diploid males, which are nearly always sterile,
can be produced experimentally in this species, although the fertile males
are normally gynogenetic and haploid. Whiting irradiated females of an
inbred wild-type stock, whose egg chromosomes were in the metaphase
stage of the first meiotic division, with doses of X rays (up to 42,000 r) ;
they were then mated with untreated males carrying recessive marking
genes. Among the surviving progeny were fertile males showing the
characters of the recessive mutants (haploid and paternal in origin), in
addition to the expected wild-type males (haploid and maternal) and
females (diploid and biparental). Cytologic examination (A. R. Whiting
1948) indicated that the treatment retards and distorts the egg pronucleus

644 RADIATION BIOLOGY

to such a degree that the sperm pronucleus cleaves and develops into a
normal, fertile, haploid male with paternal traits only.

2-2b. Monosomic and Polysomic Types. If, in an otherwise diploid
organism, three chromosomes of one type are present, the individual is
called "trisomic." If one of the chromosomes is represented only once,
the individual is called "monosomic." Irregularities in the mitotic
mechanism account for such deviations from the normal diploid condi-
tion. Nondisjunction of chromatids in somatic mitosis, or of homologues
in meiosis, may result in the passage of both to one pole of the spindle.
Bridges (1916) found that eggs of normal, untreated females of D.
melanogaster may occasionally carry two X chromosomes or none, rather
than a single X. If such females are mated with males carrying sex-
linked dominant markers, the progeny arising from these exceptional
eggs can readily be detected. They will be either recessive females
(XXY, derived from XX eggs fertilized by a Y-transmitting sperma-
tozoon) or dominant males (XO, derived from no-X eggs fertilized by an
X-transmitting sperm), in contrast with the normal progeny, which are
dominant females and recessive males. Although the exceptional eggs
originate for the most part as a result of nondisjunction of the X, they
apparently can also arise from losses of that chromosome, since XX eggs
are less frequent (1 :2500) than no-X eggs (1 :600).

The frequency of nondisjunction and chromosome elimination can be
increased by treatment of females with X rays, as was shown in the
studies of Mavor (1922, 1924a) and Anderson (1924, 1925a, b, 1931) on
D. melanogaster, and Demerec and Farrow (1930a, b) on D. virilis. Loss
of an X chromosome in a cleavage division of a normal female may give
rise to a gynandromorph composed of XX and XO tissues, and such
individuals have been obtained in X-ray experiments (Mavor, 1924b;
Patterson, 1930; Bonnier, Liming, and Perje, 1949). Losses of X and Y
chromosomes following irradiation of spermatozoa, which result in death
of the embryos, will be considered in the discussion of dominant lethals.

Individuals that are monosomic for the fourth chromosome arise
spontaneously in Drosophila, and may also be induced experimentally.
Dobzhansky (1930) observed mosaics among the progeny of treated flies,
which were haplo-fourth in part of the body. The mosaic individuals
described by Mohr (1932) presumably belonged to this category, as did
some of the Minutes described by Muller (1928a, 1930) and others.
Flies that are trisomic for the fourth chromosome are viable (cf. Fig.
9-7e), although individuals tetrasomic for this chromosome are not.

Nondisjunction or loss of the second or third chromosomes as a result of
irradiation of gametes would also lead to the formation of zygotes that
were either monosomic or trisomic for these longer autosomes; but indi-
viduals of these types have not been detected by genetic or cytologic
methods of analysis, and presumably are eliminated in embryonic stages.

CHROMOSOME ABERRATIONS IN ANIMALS 645

2-2c. Deficiencies and Duplications. A segment, rather than an entire
chromosome, may be eliminated from the normal complex or added to it
as a result of irradiation damage. The loss of a section of a chromosome
is designated as a "deficiency," and the repetition of a section as a
"duplication." The length of the section removed or added and the
genetic properties of the chromatin involved determine the extent of
genie unbalance and the capacity of the cells receiving the altered
chromosomal complement to survive. Deficiencies and duplications con-
fined to heterochromatic ("inert") regions, such as exist in the proximal
third of the X chromosome and throughout the Y chromosome of D.
melanog •aster, have less effect in modifying normal developmental proc-
esses than alterations of comparable length within the euchromatic
("genetically active") portions of the chromosomes. (As indicated
previously, the loss of the entire Y leads to the production of an XO male,
which is viable though sterile; and the addition of a Y, as in an XXY
female, does not appreciably reduce viability or fertility.)

Deficiencies confined to euchromatic regions have, in general, more
deleterious effects than duplications of comparable length. Studies on
Drosophila have indicated that deficiencies are usually lethal when
homozygous or hemizygous (as, for example, when a deficient X chromo-
some is present in duplicate in an XX female zygote, or as a single X in an
XY male zygote), although some short terminal deficiencies are not
(Demerec and Hoover, 1936). Many of the so-called "lethal mutations"
of Drosophila are attributable to minute deletions (Slizynski, 1938). A
deficiency may at times be detected by the absence of specific marking
genes. Thus, a deficiency induced in the 1A5-8 region of the X chromo-
some of D. melanogaster by irradiation of wild-type males, which were
mated with y sc females, was detected by the appearance of female
progeny with yellow body color (Sutton, 1943). A deficiency for band
3C7 of the X chromosome acts as a dominant (Notch), producing flies
with serrated or notched wings (Demerec, in Demerec and Kaufmann,
1937). Such correlations of phenotypic effects with chromosomal aber-
rations have been useful in determining the loci of specific genes on the
chromosomes, as is illustrated in Fig. 9-10.

Deficiencies may be either terminal or intercalary in position. Ter-
minal deficiencies are frequently detected in irradiated cells as fragments
separated from the portion of the chromosome having the centromere or
spindle-attachment region (Figs. 9-3 and 9-4). A terminal fragment
detached from an ordinary, or monocentric, chromosome lacks the
centromere essential for its normal transportation to the spindle pole at
anaphase. Consequently, it is usually not included in either of the
daughter nuclei (however, see Carlson, 1938b). Fragments detached
from chromosomes having compound or diffuse centromeres — such as
those of Ascaris, the coccids, or the bearberry aphid — provide an excep-

646

RADIATION BIOLOGY

tion to the general rule, since they are transported to the spindle poles
together with the main body of the chromosomes from which they have
been detached (White, 1936; Bauer and LeCalvez, 1944; Hughes-
Schrader and Ris, 1941; Ris, 1942). Exclusion of detached fragments
from the daughter nuclei upsets genie balance ; it has been suggested that
the reduction in fertility that follows irradiation of gametes in various
species of animals is due in large measure to fragment production. This
topic has been reviewed by Lea (1947b).

0.8

1.5 1.7 3.0

Fig. 9-10. Salivary-gland chromosome map of the prune-echinus region of the X
chromosome of Drosophila melanogaster, indicating the extent of 14 deficiencies that
have been studied. (From Demerec, in Demerec and Kaufmann, 1937.)

Terminal deficiencies are probably produced by irradiation of sperma-
tozoa of Drosophila, but they presumably have dominant lethal effects
(Pontecorvo, 1941, 1942; Pontecorvo and Muller, 1941; Muller, 1941;
Fano, 1941; Demerec and Fano, 1944; Lea and Catcheside, 1945; Catche-
side and Lea, 1945a). Loss of a fragment from one of the longer auto-
somes in cleavage mitosis will upset genie balance and cause abortion of
the embryo. The mechanism probably involves establishment of a
breakage-fusion-bridge cycle in the centric portion of the chromosome
(similar to that described by McClintock, 1939, for maize) as a conse-

CHROMOSOME ABERRATIONS IN ANIMALS 647

quence of end-to- end union of sister chromatids at the site of the break,
formation of a chromatin bridge between the two separating centromeres
at anaphase, and subsequent breakage of the extended strand at an
indeterminate position. The irregular cleavage mitoses lead to abnormal
embryonic development, and occasionally cellular multiplication may
occur without differentiation (Sonnenblick and Henshaw, 1941).

If the breakage-fusion-bridge cycle occurred in either the fourth
chromosome or a sex chromosome, it might not be expected to prove
lethal in all cases, since haplo-fourth and XO individuals are viable.
Muller (1940) and Pontecorvo (1941) found, however, that the number of
individuals surviving loss of a sex chromosome was much lower than
expected. It thus appears that the actual mechanism of loss may
involve mechanical difficulties that upset the course of mitosis and modify
subsequent developmental processes, even though the absence of the
chromosome from the complex would not in itself cause death of the
embryo (cf. Fano and Demerec, 1944; Catcheside and Lea, 1945b;
Catcheside, 1948). Viable losses, which occur with a frequency of about
1 per cent at 4000 r according to Pontecorvo (1942), presumably occur
when the break is produced close to the centromere, so that a short
bridge is formed at anaphase. Formation of daughter nuclei could
presumably then proceed without interference from the chromosome in
the process of elimination (see also Catcheside, 1948).

For the reasons indicated, most of the detectable deficiencies induced
by irradiation of spermatozoa of Drosophila are intercalary rather than
terminal. Some exceptions have been reported, however (Demerec and
Hoover, 1936; Sutton, 1940; Catcheside and Lea, 1945a), and it has also
been noted that terminal inversions occur in nature (e.g., by Kaufmann,
1936 and Kikkawa, 1938, on D. ananassae). These observations present
an objection to Muller's suggestion (1941) that the induced losses are
always intercalary rather than terminal, the deleted section approaching
the tip but not including the terminal band or bands. Muller's assump-
tion (1940) that the chromosomes of Drosophila contain specialized
terminal chromomeres or telomeres essential to their survival has also
been contested, on the basis of an analysis of breaks induced in ring-X
chromosomes (Catcheside and Lea, 1945b).

The extent of an induced intercalary deficiency may be determined
roughly by a method designed by Painter and Muller (1929), which
measures the suppressing action of genes in the remaining or centric
portion of the chromosome. Irradiated males are mated with females
having attached-X chromosomes carrying recessive marking genes.
When a spermatozoon carrying an X from which a section has been
deleted as a result of radiation injury fertilizes an egg with attached-X
chromosomes, the individual created has X-chromosome material in
excess of that of a normal diploid female. The wild-type genes in the

648 RADIATION BIOLOGY

duplicated section will suppress the expression of recessives in the
attached X's. Dobzhansky (1936) has summarized data from one of his
experiments in which the females were homozygous for y, wa, ec, and /.
The results are shown in Table 9-1.

Table 9-1. Progeny Obtained in a Cross between Irradiated Wild-type Males
of D. melanoyaster and Attached-X Females Homozygous for the Sex-linked

Recessives y, wa, ec, and /
(Dobzhansky, 1936)

y wa ec / 9 (normal offspring) 2185 wa ec f 9 (duplication for y) 8

Wild-type c? (normal offspring) . . . 1879 ec f 9 (duplication for y, wa) 9

Superfemales 4 /9 (duplication for y, wa, ec) 6

Wild-type 9 (from detachment of Wild-type 9 (duplication for y, iva,

attached X's) 3 ec, /) 1

y ioa ec f o" (from detachment of y wa ec 9 (duplication for /) 1

attached X's) 1

A more accurate measure of the extent of an induced duplication or
deficiency may be obtained by cytological analysis. The procedure has
generally involved examination of the salivary-gland chromosomes of the
Fi larval progeny of irradiated fathers, although a duplication may occa-
sionally be recognizable in neuroblast prophases (Fig. 9-7e). In the
salivary-gland-chromosome studies many of the longer duplications have
been detected as mosaics, being present in some, but not all, cells of the
gland. Since individuals have rarely been detected in which all the cells
of the gland carried extensive duplications, it seems probable that aber-
rations of these types interfere with normal embryonic development,
although they are not necessarily lethal in salivary-gland cells.

The nature and extent of several different duplications is represented in
diagram form in Fig. 9-11. These aberrations are readily recognized
because a portion of the chromosome is present in triplicate, two strands
having been contributed by the irradiated father, and one by the
untreated mother (as is shown by the photographs reproduced as Figs.
9-od and 9-6a). Figure 9-1 lg represents a duplication covering prac-
tically the entire length of the right limb of the third chromosome ; it was
observed in some cells of a mosaic gland. The rearrangement dia-
gramed in Fig. 9-1 la was present in some cells of the salivary gland as a
duplication of the region 5F to 15F of the X chromosome; other cells of
the same gland were deficient for this region. This aberration indicates
that a duplication and a deficiency may arise as complementary types as a
result of exchange between sister chromatids and their separation into
daughter nuclei (Kaufmann, in Demerec, Kaufmann, and Sutton, 1939;
diagram in Fig. 9-4, column 3, of this chapter).

The origin of intercalary duplications by sister-chromatid exchange
reveals the method whereby replicated sections have been built into the
chromosomes of various species of Diptera in the course of phylogeny

{a) xg

3Lg

if)

(6)

2Rg;

x n >

.,. y>:- '"":\"[ii////m//////m//////////m/////////imimmm

3 o::... '.''.'T'-'

3RS

(M

a:-"":"»

(cj

2R

2l ' li^^^^^^* ^^^^ ^^^^M>

'biii/iii/tiititiiitti, :::.:;::: » 3R <5

,)... :..::..• .... -7— - ■. - ■;. /;///////////)

Y

s

(d) (A)

2R

3L

s

2Rg

3R CMmm^^mmmmmm^^ 3R tmwMwmmm»mmmM//w/mMMm

<j^— .;:.. .;:;;.;",■ ■ ~> r^

(<?) (2)

Fig. 9-11. Chromosome and chromatid breaks in Drosophila melanogaster. Dia-
grams showing positions of breaks in nine rearrangements having duplicated sections
in one of the chromosomes represented. The chromosomes (2L, 2R, 3L, 3R, X, 4)
are diagramed in the two-strand stage; centromeres are shown at left. Approximate
positions of breaks are indicated by gaps, the chromosome type occurring at the
same level in both strands, the chromatid type represented in only one strand. Por-
tions of the chromosomes that were identified in the salivary-gland nuclei are repre-
sented in solid color; portions not recovered, in cross hatching. The rearrangement
indicated in (a) appeared as a mosaic, some nuclei of the salivary gland showing a
duplication of the reversed-repeat type, other nuclei showing a deficiency resulting
from recombination of the stippled parts of the chromosome. When chromosomes
other than those showing the duplicated sections were involved in a complex rear-
rangement, the positions of breaks in these chromosomes are also indicated. Thus,
in 0') duplicated sections of 2L were identified, but the rearrangement also involved
breaks in 2R, 3R, and the Y chromosome. The rearrangement diagramed in (d)
had 14 breaks, although only 12 are shown because the positions of the breaks in one
of two reversed repeats could not be identified. The rearrangement represented in
(g), which is a duplication for the entire length of 3R, was found in some cells of a
salivary gland but was lacking in others.

649

650 RADIATION BIOLOGY

(Fig. 9-5d; see also Kaufmann and Bate, 1938). The existence of such
repeats can readily be disclosed by examination of patterns of banding of
salivary-gland chromosomes (e.g., as reported by Bridges, 1935, Offerman,
1936, and Kaufmann, 1939a, in D. melanogaster; by Metz and Lawrence,
1938, and Metz, 1947, in Sciara ocellaris and S. reynoldsii) .

Muller (1950a) has reported experiments designed to determine
whether small deficiencies induced by irradiation arise as a result of such
exchange between two different breaks in sister chromatids or as a con-
sequence of two adjacent breaks in the same strand with consequent
deletion of the intermediate piece. In these studies the effects induced in
ring-X chromosomes were compared with those induced in the normal X.
On theoretical grounds it would be assumed that ring chromosomes would
yield a lower frequency of recoverable deficiencies because of the produc-
tion of dicentric double chromosomes resulting from an exchange between
sister strands (cf. Bauer, 1939a, 1942; Muller and Pontecorvo, 1942;
Pontecorvo, 1941). However, the ring chromosomes gave just as high a
yield of deficiencies as the non-rings for a given dose of irradiation. It is
assumed, therefore, that very few minute deficiencies arise by the method
of "unequal crossing over."

The genetic effects of duplication have been studied extensively in
recent years. Burdette (1940) reported that homozygous duplications
decrease fertility and viability in D. melanogaster as compared with the
corresponding heterozygous duplications, and that these in turn are
usually less viable than normal. The series of rearrangements studied by
Burdette and by Patterson, Brown, and Stone (1940) indicated that the
effect of a duplication is not necessarily proportional to its length, so that
duplications of similar lengths in different regions may have dissimilar
effects on phenotype, fertility, and viability. Specific phenotypical
effects of duplications of short regions of the chromosomes of Drosophila
have been reviewed by Lewis (1950).

2-2d. Intrachromosomal and Interchromosomal Rearrangements. For
descriptive purposes it will be assumed that in these aberrations, as con-
trasted with those previously mentioned, there is no gain or loss as com-
pared with the normal condition, but merely a realignment of parts of one
or more chromosomes.

The viable types of intrachromosomal rearrangement include inver-
sions and transpositions. In the former, a section between two breaks is
displaced and reinserted in reverse order, as if revolved through 180°
(Fig. 9-5c). Thus, in a chromosome in which the normal sequence of
genes is represented by the letters ab cdef ghij, the arrangement may be
altered by inversion to ab fedc ghij. Two or more inversions may occur in
the same chromosome, and occasionally they are observed in tandem,
with one point of breakage common to the two inverted sections (Hoover,
1937).

CHROMOSOME ABERRATIONS IN ANIMALS 651

The term transposition is applied to a rearrangement in which a seg-
ment is transferred from one position to another within the chromosome.
Three breaks are involved. If the unaltered chromosome is represented
by the letters ab cdefg hi j, transposition may produce such types as
ab hi cdefg j, or ab ih cdefg j, in which the loci of the transposed segment
are either in their normal sequence or inverted. Pairing between a
chromosome with a transposed section and its normal homologue pro-
duces the type of configuration indicated in Fig. 9-5b. The rearrange-
ments shown in Fig. 9-5b and c are intrabrachial, or paracentric; that is,
they are restricted to one limb of a chromosome. An interbrachial, or
pericentric, inversion, which extends across the centromere and encom-
passes both limbs of a chromosome, furnishes a pattern of pairing with
its normal homologue in salivary-gland cells of D. melanogaster similar to
that of a reciprocal translocation, because the characteristic inversion
loop is distorted by the chromocentral attraction of the heterochromatic
regions adjoining the centromere.

A reciprocal translocation, involving a mutual exchange between two
different chromosomes, is clearly denned in salivary-gland cells by the
cross-shaped configuration established by pairing of the rearranged
chromosomes and their normal homologues. The translocation illus-
trated in Fig. 9-6c is between the left limb of the second chromosome and
the right limb of the third. If we assign the letters abcde fghij to the
normal sequence of parts in 2L and the letters klmnopq rst to the normal
sequence in 3R, we may indicate the new sequences produced by trans-
location as abcde rst and klmnopq fghij, the inequality in length being
due to the location of one break near the tip of 3R and the other near the
middle of 2L. A reciprocal translocation between the right limb of the
third chromosome and the Y chromosome is illustrated in Fig. 9-6b.
Since the Y chromosome is represented in the salivary-gland cells of male
larvae of D. melanogaster by only a few bands that form part of the
chromocenter, the position of breakage in the Y is not determinable in
these cells, but can at times be determined by inspection of neuroblast
chromosomes, as is shown in Fig. 9-7a.

An inversion-translocation complex is illustrated in Fig. 9-6d. In the
production of this rearrangement, the distal part of the left limb of the
second chromosome (2L) was transferred to the base of 2R, the distal
part of 3L to the base of 2L, and the distal part of 2R to the base of 3L.
Pairing between these rearranged chromosomes and their normal homo-
logues produced the pattern of radiating arms shown in the photograph.
Descriptive details of other rearrangements have been published by
Kaufmann (1939b).

Several independent two- or three-break rearrangements may be pro-
duced in the same irradiated spermatozoon. A large number of breaks
within a single spermatozoon may also combine to form one or more

652 RADIATION BIOLOGY

complex cyclic rearrangements. In the most complex rearrangement so
far reported (induced in a spermatozoon of D. melano g aster) , at least 32
breaks were involved, and the broken ends had reassociated to form sev-
eral independent exchanges (Kaufmann, 1943). Because of difficulties of
diagnosing breaks in the proximal heterochromatic regions of the chromo-
somes, it was not possible to determine exactty all the details of recombina-
tion, but on the simplest possible explanation there were seven independ-
ent exchanges: four with two breaks, one with four breaks, and two with
ten breaks each. In one of the latter a cyclic rearrangement occurred
among the ends of chromosomes broken at the ten points indicated by the
map numbers 87B/51E/81/49F/70C/33E/42C/56B/64C/76A. (Using
this system of notation a simple reciprocal translocation would be repre-
sented by two breaks, such as 3C/24D.)

Chromosomal aberrations in Drosophila, even though they reveal no
perceptible loss of chromosomal material when examined in salivary-gland
preparations, are for the most part inviable when homozygous (Schultz,
1936, has reviewed the supporting evidence in considerable detail).
This suggests that the production of a rearrangement alters the genetic
constitution of the chromosomes involved. Analysis of radiation-
induced lethal mutations has shown that a considerable fraction of them
are associated with chromosomal rearrangements. The frequency varies
with the dose (Oliver, 1932; Demerec, 1937; Herskowitz, 1946); it is
about 35 per cent at 3000 r. (The association of a high proportion of
chemically induced lethals with chromosomal aberrations has also been
established by Slizynska and Slizynski, 1947.) Demerec (1937) tested a
series of 26 X-ray-induced lethals associated with chromosomal rearrange-
ments, and found that in 24 the locus of the lethal coincided with one of
the breakage points. This correlation suggests either that a submicro-
scopic change (in the nature of a gene mutation or inactivation) has
occurred adjacent to a break, as a consequence of the passage of an ion-
izing particle through the chromosome at that position, or that functional
modification has followed displacement of a gene from its normal rela-
tions with adjacent regions of the chromosome (position effect). Evi-
dence in favor of the former of these alternatives has been obtained in
Drosophila by analysis of the dose-frequency relations determined experi-
mentally in studies of lethal mutations and chromosomal aberrations
(Lea and Catcheside, 1945; Lea, 1946; Herskowitz, 1946), and by analysis
of the modifying effect of near-infrared radiation on the frequency of
X-ray-induced lethals and chromosomal rearrangements (Kaufmann and
Gay, 1947). When spermatozoa of D. melanogaster were exposed to near-
infrared radiation before treatment with X rays, they showed an increase
in frequency of chromosomal rearrangements, as compared with controls
that received only the X-ray treatment; but there was no increase in the
frequency of sex-linked recessive lethals, although about one-third of them

CHROMOSOME ABERRATIONS IN ANIMALS 653

were associated with gross chromosomal aberrations. An increase would
be expected if the lethals were dependent for their expression on the
production of rearrangements involving two independent breaks (Fano,
in Kaufmann et al., 1947). This evidence, and that afforded by the
dose-frequency analyses of Lea, Catcheside, and Herskowitz, indicate
that the lethals associated with induced chromosomal rearrangements in
Drosophila do not represent a special class caused by a position effect.
Muller (1950a) has objected to this interpretation, with the suggestion
that near-infrared pretreatment may actually increase the frequency
of position-effect lethals associated with gross chromosomal exchanges,
but that such increase is counterbalanced by a decrease in the frequency
of lethals associated with small rearrangements. This explanation
involves the assumption, for which there is no experimental evidence,
that small rearrangements, unlike the gross ones, are not increased in fre-
quency by pretreatment of spermatozoa with near-infrared radiation.

Another aspect of the phenomenon of chromosomal exchange involves
the breakage and recombination of homologous chromosomes, designated
by the term "crossing over." In females of Drosophila, crossing over
occurs regularly in the course of meiosis, presumably during prophase in
the oocyte. In males crossing over occurs spontaneously with very low
frequency, but can be induced by X rays, as shown by Friesen (1933,
1934), Patterson and Suche (1934), and others. The frequency of cross-
ing over in the female may also be increased by irradiation (see, for
example, Whittinghill, 1938). Naturally occurring and induced crossing
over have many similarities; exchange takes place at identical loci while
the chromosomes are in the four-strand stage, and the recombinants are
usually not lethal when homozygous (references in Shapiro, 1945).
Many cases of mosaic formation in X-rayed larvae are assumed to result
from crossing over that is induced in somatic tissues (Stern, 1936). In
these cells the presence of chiasmalike configurations, which may afford
the structural basis conducive to crossing over, has been demonstrated by
Kaufmann (1934). Similar configurations have been observed by
Cooper (1949) in spermatogonial cells.

The question thus arises whether irradiation promotes crossing over by
facilitating a method of exchange comparable to that normally operating
in the course of meiosis in the female, or whether the breakage-recombina-
tion process resembles that involved in exchange between nonhomologous
regions. A related problem concerns the stage of gametogenesis during
which induced crossing over occurs. Solution of these problems has
generally been assumed to require analysis of the progeny of individual
irradiated flies so as to detect the possible occurrence of "clusters" of
crossover types. The production of such clusters has been interpreted as
indicating that crossing over is induced in gonial stages (e.g., Whitting-
hill, 1938, 1950; Hinton and Whittinghill, 1950). Some parallel lines of

654 RADIATION BIOLOGY

evidence conform with this interpretation. It has been reported that the
exceptional flies which result from interchange between X and Y chromo-
somes in attached-X females of D. melanogaster (Kaufmann, 1933) fre-
quently occur in clusters, suggesting that they originate in a gonial cell
(Neuhaus, 1936; Cooper, 1946). Whittinghill (1947) has observed that
the presence of a Curly inversion in the second chromosome of males of
D. melanogaster does not influence the induced frequency of crossing over
in the third chromosome, although it is known (e.g., Schultz and Redfield,
1932) that the presence of an inversion may affect the frequency of
recombination when crossing over occurs in the course of meiosis in the
female.

On the other hand, Parker (1948) suggests that induced crossing over
in the male may result from breakage in the spermatocyte of both
homologous chromosomes by the action of a single ionizing particle — a
phenomenon that will be discussed subsequently. This hypothesis
assumes that synapsis of homologues occurs within a restricted region of
the chromosomes being tested, since it was found that crossing over is
limited to a short portion of the chromosome at any time. The produc-
tion of clusters is attributed to similarity of all cells in a cyst with respect
to their crossing-over potentialities. Dose-frequency relations as deter-
mined by Parker for the 500- to 2000-r range were in accord with the
assumption that a single ionizing particle is concerned; there was an
increase in crossing over with increase in dosage, but no increase in the
proportion of double crossovers. Shapiro (1945), however, observed a
much higher frequency of crossing over at 3000 than at 1500 r (0.69 as
compared with 0.23 per cent in one experiment), but only a slight addi-
tional increase (to 0.79 per cent) when the dose was increased to 4500 r.
He concluded that crossing over may be the result of breakage of chromo-
somes by ionization, especially in late spermatogonia or in spermatocytes,
but that selective elimination of germ cells may account for the declining
proportion of detectable recombinants with increasing dosage. A further
line of evidence that the frequency of crossing over is a function of the
distribution of ionizations is afforded by the observation of Lefevre (1948)
that neutrons are relatively more effective per ionization than are y
rays in inducing somatic crossing over in Drosophila. It thus seems
probable that irradiation-stimulated crossing over involves the produc-
tion of chromosomal breaks as a consequence of ionization; but the
period of gametogenesis during which this occurs is not clearly defined.

3. THE PROCESS OF STRUCTURAL REARRANGEMENT

From the foregoing discussion it is evident that the breakage and
recombination phases must both be considered in an analysis of the
process of induced structural rearrangement. Whether breakage pre-

CHROMOSOME ABERRATIONS IN ANIMALS 655

cedes recombination, or whether both phases are part of a single process
and occur simultaneously, could not be decided with certainty from the
data obtained in the earlier genetic experiments; but more recent evi-
dence, both genetical and cytological, is in accordance with the first of
these alternatives (reviewed in Bauer, 1939c; Muller, 1940). After the
breaks — or potential breaks — have been induced, the establishment of
new associations of ends is influenced by movement of the chromosomes.
In dividing cells such movement takes place as mitosis progresses, with
the result that breakage and recombination occur in close succession.
In the sperm head of Drosophila, the chromosomes are quiescent, so that
the types of rearrangement that depend on establishment of associations
among breakage ends at different loci are not realized until after the
spermatozoon has penetrated the egg in fertilization, when chromosome
movement incident to the formation of the male pronucleus presumably
provides opportunities for recombination (Muller, 1940; Kaufmann,
1941a; Dempster, 1941a; Makhijani, 1945). The delay between break-
age and recombination, which can be controlled by withholding the
treated males from copulation, permits experimental determination of
the time and method of combination of breakage ends in the production
of detectable aberrations.

3-1. THE BREAKAGE PROCESS

3-la. Induction of Breaks. Physical data indicate that ionizations are
localized along the paths of ionizing particles throughout the cells of an
irradiated tissue. The sequence of events that transpires between
ionization and the production of chromosomal lesions remains largely
conjectural (see Zirkle, 1949), although an understanding of possible
intermediate steps is slowly emerging. It is now generally agreed that
ionization produces chemical changes in the irradiated tissue (see, for
example, the discussion of Allsopp, 1948; Allsopp and Catcheside, 1948).
Available evidence suggests that chemical changes resulting from ioniza-
tion of materials within the chromosome itself may initiate the series of
reactions that lead to its breakage. Whether the chemical reactions
originate for the most part in molecules of constituent nucleic acids and
proteins, or are mediated through associated aqueous solutions, remains
to be determined. For this reason the process of induced chromosome
breakage cannot at present be described as exclusively direct or indirect
according to the definition formulated by Bacq (1951), which restricts the
direct effects to changes initiated by ionizations occurring in or on the
surface of organic molecules. Studies conducted in the past few years
showing the dependence of the yield of chromosomal aberrations on the
physiological conditions of the cell or organism speak strongly in favor of
the indirect reaction. Thus Baker and Sgourakis (1950) have shown
with Drosophila that the frequency of dominant and recessive lethals is

656 RADIATION BIOLOGY

40-70 per cent lower if the flies are irradiated in an atmosphere of nitrogen
than when they are irradiated in an atmosphere of oxygen. (Similar and
more extensive results have been obtained in studies of plant chromo-
somes; e.g., by Thoday and Read, 1947, 1949, and by Giles and Riley,
1949.)

One of the criteria of the indirect effect is the protection afforded by
dissolved substances, other than those being tested, present in the aque-
ous medium. Further evidence, therefore, of the indirect nature of the
reaction system is afforded by the findings that various substances, such
as cysteine, glutathione, thiourea, ethyl alcohol, azides, and cyanides,
when used in proper concentrations, afford protection against the effects
of ionizing radiations. Pioneering work along these lines with respect to
biologically active materials has been done on isolated enzymes (e.g., the
studies of Dale, 1940, 1942; Dale, Davies, and Meredith, 1949; and
McDonald, reported in Kaufmann et al., 1950), and has recently been
extended to whole-body irradiation (reviewed in Bacq, 1951). From the
results obtained in irradiating entire animals, which indicated that the
median lethal dose is much higher when protective agents are used, it can
only be inferred that chromosome breaks are involved, although the
dosage in most of these experiments was of the order of magnitude that is
known to produce chromosomal rearrangements.

Whatever the precise course of events in the production of any specific
break may be, it appears probable that molecules near a high concentra-
tion of ionizations may undergo chemical change although not themselves
ionized, and that a large number of chemical activations per chromosome
is required before any observable breaks occur (Fano, Caspari, and
Demerec, 1950).

If an indirect reaction system can produce chromosome breaks, it
may conceivably originate in some part of the cell other than the chromo-
some (a remote effect). Duryee (1939, 1949) has reported that the
chromosomes of isolated oocyte nuclei of amphibia are much less sensitive
to radiation damage than chromosomes in the intact cell. Chromosomal
injuries are also produced by exposure of isolated nonirradiated nuclei to
irradiated cytoplasmic brei. Although these findings may suggest "that
nuclear damage results mainly from chemical change in the cytoplasm,"
as Duryee has proposed, the intense X-ray doses used and the surgical
procedures employed in isolating nuclei involve a reaction system differ-
ent from that which leads to the formation of viable chromosomal
rearrangements in the intact cell. It has been shown, however, that the
types of viable aberrations induced by ionizing radiations can be pro-
duced by treating cells with chemical mutagens, which do not reach the
nucleus directly but diffuse into it, either in their original form or as
degradation products. For example, chromosome breaks have been
induced in Drosophila by mustard gas and nitrogen mustard (Auerbach

CHROMOSOME ABERRATIONS IN ANIMALS 657

and Robson, 1946; Auerbach, 1949, 1950; Kaufmann, Gay, and Roth-
berg, 1949), and in chick fibroblasts by urethane (Paterson and Thomp-
son, 1949).

Such results have raised the question whether breaks in chromosomes
are ordinarily produced by a series of reactions originating in remote
parts of the cell. A. R. Whiting (1949, 1950) examined this question in
considerable detail. Her experiments on Habrobracon confirm and extend
the earlier findings on various species, including D. melanogaster (reviewed
in Schultz, 1936). Cytoplasmic injury produced in eggs by very intense
doses of radiation inhibits the development of all embryos. After lower
doses, however, which still may be twenty times greater than that
required to induce dominant lethals in all the egg chromosomes, andro-
genetic males may develop (from nonirradiated chromosomes in X-rayed
cytoplasm). No mutations are induced in these males, which are
normal in morphology and fertility and produce fully viable progeny.
If irradiated cytoplasm exerts any mutagenic action, it must be effected
within a shorter period than that tested in these experiments. Auerbach
and Robson (1947) also concluded that there was no delayed effect on
untreated chromosomes of cytoplasm treated with mustard gas. Other
studies indicate that the lethal effects of radiation, such as killing of
Drosophila eggs, depend primarily on nuclear damage (e.g., Langendorff
and Sommermeyer, 1940).

It appears improbable, therefore, that ionizing radiations or chemical
mutagens effect genetically significant types of chromosome breaks
through the accumulation of toxic substances in the cytoplasm. The
fact that chemicals can induce rearrangements similar to those induced
by radiations may merely indicate that breaks are produced with com-
parable potentialities regardless of the agent used in their production; it
does not, as some authors have assumed, dispose of the possibility that
the sequence of events leading to breakage can originate within the
chromosome. Similar end results, as represented by a series of breaks or
rearrangements, may conceivably be mediated through a variety of
channels. A more specific definition of the radiation-induced reaction
system therefore depends on more precise information concerning the
effects of ionizations on the genetically active materials of the chromo-
some and on other cellular materials. This being the case, the question
whether an effect is localized or remote is reduced to a choice of prob-
abilities between the effectiveness of a sequence of events originating in
the chromosome and that of one initiated elsewhere in the cell (Kauf-
mann, 1948a).

The target theory, in its more general terms, is formulated in accord-
ance with the first of these alternatives. Primary production of breaks
occurs with a frequency that is proportional to dose over a wide range of
wave lengths, including X rays and y rays. The effect is independent of

658 RADIATION BIOLOGY

the duration of a given dose or the number of treatments into which it is
divided, and is not appreciably modified by temperature within the
narrow range that can be utilized while maintaining viability (and, in
some cases, fertility) of the test organisms (summary of evidence in
Catcheside, 1945). These experimental results add further weight to
the argument that chromosome breakage is not primarily attributable to
a long chain of chemical reactions operating over considerable distances
within the cell. But, as Fano, Caspari, and Demerec (1950) have
pointed out, experiments on time distribution could only show interaction
between separate events whose activity, even though not permanent,
lasts much longer than that of a chemical activation. Phenomena
exhibiting such properties are not among the usual physical effects of
radiation on matter and must therefore be characteristic of biological
materials. These authors have also pointed out that the direct propor-
tionality between dosage and genetic effect would hold for indirect as
well as direct action of radiation, provided that the efficiency of the
indirect mechanism was not sensitive to changes in temperature and
other factors.

The more rigid concept of the target theory, that a single ionization
within a sensitive zone suffices to induce a break, was an outgrowth of
earlier studies of the conditions governing the induction of gene mutations
(summary of experimental data in Timofeeff-Ressovsky, 1937; and
critical evaluation in Muller, 1940, 1950a; Fano, 1942; Fano, Caspari,
and Demerec, 1950). This view has been tempered in recent years by
development of the interpretation that chromosome breakage is refer-
able to a cluster of ionizations produced by the passage of a single ion-
izing particle. Such clustering takes place at the tail end of the electron
track, or at places where the main track branches out into other short
tracks; it may occur along the entire length of the track of a recoil proton
(Lea and Catcheside, 1942; Fano, 1943a). Relative efficiencies of differ-
ent wave lengths and types of radiation have been explained on this
basis (Giles, 1940; Lea and Catcheside, 1942; Lea, 1946, 1947a; Gray,
1946; Catcheside, 1948). For example, soft X rays of wave length
4 A produce more breaks per roentgen than harder X rays, because the
spacing of ionizations is such that the whole track corresponds to the
"tail" end of tracks of hard X rays. If the spacing is too close there is a
loss of efficiency owing to oversaturation. Thus, wave length 8 A is less
effective per roentgen than other wave lengths. Additional evidence in
support of the concept that the large concentrations of energy resulting
from clusters of ionizations are required to induce chromosome breakage
has been obtained from comparative studies of X rays and neutrons
(summarized in Fano, Caspari, and Demerec, 1950). Thus the concept
that a single ionization serves to induce a chromosome break no longer
seems tenable. "When the identification of a hit as a chemical activa-

CHROMOSOME ABERRATIONS IN ANIMALS

659

tion is abandoned, the concept itself of a hit acquires a macroscopic sig-
nificance, while the possibility of establishing a direct atomic physical
link between the hit and its biologic consequence becomes more remote."
(Fano, Caspari, and Demerec, 1950.)

3-lb. The Sphere of Action of an Ionizing Particle. Development of
the interpretation that a single ionizing particle, rather than a single ion-
ization, produces a chromosome break has modified the original concept
that methods using ionizing radiations are adequate to determine the
number of component strands of a chromosome at the time of treatment.
On the assumption that a single ionization would suffice to induce a
break, it was inferred that irradiation of the divided, or double, chromo-
some would produce breaks independently in the two chromatids (chro-
matid breaks), whereas irradiation of the undivided, or single, chromo-
some would produce breaks that would be transmitted in the subsequent
division equally to the daughter chromatids (chromosome breaks) . The
experimental results indicated in a general way that prior to a certain
stage in mitosis the chromosome reacts as a unit to the ionizing radiations
(as shown in Fig. 9-3, column 1) and that subsequently each of the two
chromatids is affected independently (Fig. 9-4, column 1). As an
example, Carlson (1941b) presented the data shown in Table 9-2, which

Table 9-2. Percentage Neuroblasts of the Grasshopper, Chortophaga, Reveal-
ing Chromosome Aberrations at Stated Time Intervals after Treatment with

125 r of X Rays

(Carlson, 1941b)

Hours after X irradiation

Chromatid translocations

Chromatid + chromosome translocations.

Chromosome translocations

Chromosome fragments

96

72

48

36

24

0

0

1

3

12

0

0

0

0

4

13

25

26

15

8

17

34

41

43

57

12

12
0
0

38

indicate the frequency of interchanges involving chromosome and
chromatid breaks in neuroblast cells of the grasshopper Chortophaga
viridifasciata. In cells irradiated 72 to 96 hours before they reached meta-
phase, at which stage they were examined cytologically, only chromosome
breaks were produced; and in those irradiated 12 hours before examina-
tion only chromatid breaks appeared ; but in the intervening period both
types of breaks were produced. In some cases chromosome and chro-
matid breaks were detected in the same chromosome. The underlying
structural basis for such response has been examined in detail by several
investigators (reviewed by Kaufmann, 1948b). There is convincing
evidence from studies on Tradescantia, in which cytological examination
sometimes permits determination of the number of strands in the chromo-
somes at the stage of mitosis during which they are irradiated, that two
chromatids may be severed conjointly by a single ionizing particle to

660 RADIATION BIOLOGY

produce a so-called " isochromatid break" (Sax, 1941; Swanson, 1943;
Catcheside, Lea, and Thoday, 1946b; Catcheside, 1948; Bishop, 1950).
In the grasshopper, Chortophaga, isochromatid breaks were observed by
Creighton (1941) in cells that presumably were irradiated in late prophase
stages. She suggested that in these cases breakage was probably attrib-
utable to an ion cluster rather than to a single ionization. The present
author (unpublished data) has confirmed by direct observation on living
neuroblasts of Chortophaga the fact that isochromatid breaks can be
produced by irradiation at late prophase stages when the chromosomes are
visibly separated into chromatids (Fig. 9-3, column 2). Such evidence
provides no support for the contention of Darlington and Koller (1947)
that isochromatid breaks are "fictitious." Accepting the validity of
their occurrence — and there are many other reasons for doing so, as
Catcheside (1948) has indicated — it follows, as Lea (1946) has empha-
sized, that evidence obtained from X-ray studies that a chromosome is
unsplit is far from conclusive.

Some of the rearrangements detected in Drosophila by salivary-gland-,
chromosome analysis have involved both chromosome and chromatid
breaks in the same chromosome. A typical rearrangement (Fig. 9-1 lc)
included two chromosome breaks, one proximal and the other distal, and
in the region between them two chromatid breaks, one in each of the
two sister strands. On the assumption that chromatid and chromosome
breaks reveal whether a chromosome has or has not divided, the chromo-
somes of the spermatozoon at the time of irradiation in this case would
appear to have been in the process of division. The condensed, inactive
condition of the chromosomes of the sperm head, however, makes division
at this stage highly improbable. On the assumption that the chromo-
some was single at the time of treatment, potential breaks were induced
at the four loci indicated, and were duplicated subsequently in each sister
chromatid. Restitution must then have occurred in each of the two
strands at that locus at which a break is detectable only in the sister
strand. Under such conditions a higher proportion of duplications might
be expected than the analysis of a large group of induced alterations has
indicated, for the reason that broken ends in both sister strands would be
available for recombination. On the assumption that the chromosome
was longitudinally double in the mature spermatozoon, either chromatid
or isochromatid breaks might be produced, as revealed in the described
rearrangement. An unequivocal decision in favor of one or another of
these alternatives is complicated by the fact that the chromosomes have
not been identified in the spermatozoon, so that no cytological evidence
is available concerning the time of division into chromatids.

Despite this lack of direct evidence, individual rearrangements have
been observed that can best be interpreted on the assumption that the
chromosomes of the spermatozoon are longitudinally double (Kaufmann,

CHROMOSOME ABERRATIONS IN ANIMALS 661

1941b, 1948b). One of these, reported by Demerec and Sutton (1940)
involved a change in the Notch region of the X chromosome of D.
melanogaster. Cytogenetic analysis showed that a deficiency in one
chromatid extended from 3C8 to 3E5, inclusive, whereas in the other the
deficiency extended from 3C7 to 3E5. Thus the two breaks to the right
occurred at the same point, the two to the left at different points although
very close together. It seems probable that the breaks in 3C7 and 3C8
were caused by the passage of a single ionizing particle but that the spread
of the effect was greater along one chromatid than along the other.

Such observations on chromosome rearrangements, and a comparable
series on gene mutations in Drosophila, are difficult to reconcile with the
theory that the chromosomes of the spermatozoon are unsplit, as some
investigators have assumed (e.g., Muller, 1940, 1941). Treatment of
Drosophila spermatozoa with chemical agents, such as nitrogen mustard,
produces types and proportions of lethals and chromosomal rearrange-
ments that can be explained most readily on the assumption that to a
large extent they originate as mosaics (Kaufmann, Gay, and Rothberg,
1949). Perhaps the specific action of this type of agent will provide the
precise information that X-ray treatment does not provide concerning
the number of experimentally separable strands that comprise the
chromosomes of the spermatozoon of Drosophila.

In some materials lesions or constrictions are seen at anaphase in
chromosomes that have been irradiated at an earlier stage, and these have
been interpreted as half-chromatid breaks (e.g., in Carlson, 1938a, on
Chortophaga). Half-chromatid breaks have also been observed in
Tradescantia (Swanson, 1943, 1947), especially after treatment of the
cells with ultraviolet radiation. Since the sphere of action of an ultra-
violet quantum rarely encompasses more than one strand of a longi-
tudinally split chromosome, the half-chromatid breaks suggest that
under certain conditions radiations may act selectively on the com-
ponent units of a multiple-strand chromosome. A striking demonstra-
tion of such action has been reported by Slyzinski (1950), confirming an
earlier observation of Marshak (1936) that X-ray treatment of Dro-
sophila embryos induces structural rearrangements affecting only a few
of the chromonemata of the chromosomes that will form the giant struc-
tures seen in the salivary-gland cells. In these bodies the rearrangements
are discernible as deficiencies, inversions, and translocations, involving
in some cases less than one-sixteenth of the diameter of the chromosome
(Fig. 9-12).

3-1 c. The Distribution of Breaks. Cytological examination of salivary-
gland chromosomes permits precise determination of the location of
radiation-induced breaks participating in viable recombinations. Exam-
ination of many hundreds of rearrangements has shown that the breaks
occur in the intervals between bands, suggesting that the integrity of the

662

RADIATION BIOLOGY

cbromomeric regions is maintained. Moreover, breaks rarely occur
between the halves of a "doublet" (exceptions are discussed by Metz,
1947), although in some cases the doublets unquestionably represent
duplicated regions rather than a single gene locus. These observations
suggest that breakage does not represent an indiscriminate severance of
the chromosome but an uncoupling of specific linkages.

The breaks that have been located in Drosophila are distributed among
the chromosomes of the set in proportion to their respective lengths
(Bauer, Demerec, and Kaufmann, 1938; Bauer, 1939b; Heifer, 1940;
Roller and Ahmed, 1942). Summarized data from the first two of these

100

Fig. 9-12. Examples of partial structural changes in salivary-gland chromosomes of
Drosophila melanogaster. (A) Deficiency. (B) Inversion. (C) Translocation.
(D) Deficiency. (E) Ring formation. (From Slizynski, 1950.)

studies, based on the analysis of larval F! female progeny of irradiated
fathers, are presented in Table 9-3. Expected values were computed on

Table 9-3. Distribution of Induced Breaks among the Limbs of the Chromo-
somes of D. melanogaster. Expected Frequencies Based on Metaphase Chromo-
some Length
(Bauer, 1939b)

X

2L

2R

3L

3R

4

Totals

Observed

Expected

325
299.8

268
279 . 3

X2

298
325 . 7
= 5.2 A

303
296 . 9
' = 5 P

394
386.2
= 0.5 -

18
18.0

0.3

1606
1605.9

the basis of length of the chromosome limbs in mitotic cells. Although
there is no direct evidence that the chromosomes of the spermatozoon are
structurally comparable to those of a mitotic cell, they give an intense
Feulgen-positive coloration similar to that of metaphase and anaphase
chromosomes, and also seem to be coiled in a similar manner [as inferred

CHROMOSOME ABERRATIONS IN ANIMALS 663

from analysis of patterns of recombination, which suggest that the X
chromosome of the spermatozoon is coiled in a series of 13 or 14 gyres
(Kaufmann, 1946a)].

In a parallel study of the salivary-gland chromosomes of male larvae,
it was found that breakage occurs in the Y chromosome with a frequency
that is approximately equal to that in the limbs of the autosomes (Kauf-
mann and Demerec, 1937). Only 262 breaks were diagnosed in these
experiments, as compared with the 1606 indicated in Table 9-3, and there
was much greater variability in the distribution of breaks among the
chromosomes. Fewer breaks were detected in the Y chromosome than
would be expected on the basis of length in mitotic cells, but this was in
part due to inability to recognize in salivary-gland chromosomes the types
of rearrangements that are restricted to heterochromatin. Since the Y
chromosome is essentially heterochromatic, and is represented in the
salivary-gland nucleus by only a few discs, practically all inversions and
intercalary deficiencies escape detection. If the estimated number of
breaks involved in such rearrangements is added to the observed number,
the total for the Y chromosome approaches that expected on the basis of
mitotic chromosome length. These findings do not support the assump-
tion of Lea (1946) and Catcheside (1948) that greater sensitivity of the
X chromosome, as compared with the Y, accounts for the fact that in
some studies (e.g., Bauer, 1942; Catcheside and Lea, 1945a) the propor-
tion of female flies hatching from eggs fertilized by irradiated spermatozoa
was found to be lower than the proportion of male flies. Demerec and
Fano (1944), on the contrary, found only a slight effect of irradiation
on sex ratio, and questioned its significance. They suggested that loss,
as a result of a breakage-fusion-bridge cycle, may occur in either Y or
X chromosomes with approximately equal frequency, and that the num-
bers of dominant lethals contributed by these two chromosomes are
roughly comparable.

The close correspondence between break frequency in the Y chromosome,
which is largely heterochromatic, and the autosomes, which are largely
euchromatic, suggests that breaks are distributed at random and in propor-
tion to length of the chromosome at the time of irradiation, regardless of
its euchromatic or heterochromatic properties. On the other hand, recent
studies, by genetic methods, of the frequencies of translocations induced
by X rays in D. virilis suggest that the Y chromosome participates in
exchange only about half as frequently as the various autosomes, although
it is of similar length (Baker, 1949). To what extent this result depends
on primary break production and to what extent on subsequent behavior
of breakage ends remains to be determined. Muller and his associates
also had decided, primarily on the basis of genetic analysis of X-chromo-
some inversions in D. melanog aster, that breaks in this chromosome are
not distributed at random. They suggested that breaks are produced at

664 RADIATION BIOLOGY

only a few points between blocks that are virtually unbreakable (Muller
and Gershenson, 1935; Muller, Raff el, Gershenson, and Prokofyeva-
Belgovskaya, 1937). On this assumption the close correlation between
break frequency and mitotic chromosome length revealed in the cyto-
logical studies would be attributable to mere coincidence (Muller, 1945).

In order to evaluate these two alternatives more adequately, an exten-
sive study was made of the distribution of breaks along the X chromo-
some of D. melanogaster (Kaufmann, 1939a, 1946a). The proximal third
of this chromosome is heterochromatic in mitotic cells of the stock
studied, being represented in salivary-gland chromosomes by only a few
discs, whereas the distal two-thirds is primarily euchromatic and con-
stitutes the major portion of the salivary-gland chromosome. The loca-
tions of about 1400 breaks were determined with reference to the lettered
subdivisions of Bridges' salivary-gland map of this chromosome. About
one-fourth of these were in the proximal heterochromatin, but rearrange-
ments confined to this region were not detected. When the estimated
number of breaks involved in such rearrangements was added to the
observed number, the proportion in the proximal region was increased to
about one-third of the total for the entire chromosome. This value cor-
responds closely to that expected on the basis of length in late prophase
stages of mitosis. A similar close correlation between mitotic chromo-
some length and break frequency in the proximal heterochromatic regions
of the second and third chromosomes was presented by Bauer (1939b).
The frequency of breaks induced in the second and third chromosomes by
treatment of spermatozoa with the chemical mutagen, nitrogen mustard,
also showed close correlation with mitotic chromosome length (Kauf-
mann, Gay, and Rothberg, 1949). Approximately equal numbers of
breaks occurred in each of the four arms, and the proportion in the
proximal heterochromatic regions was similar to that obtained in X-ray
experiments (e.g., 17.6 and 13.6 per cent in the second and third chromo-
somes, respectively, after nitrogen mustard treatment of males, as com-
pared with 18.3 and 12.5 after X-ray treatment).

In the light of these findings it seems highly probable that break dis-
tribution is a function of chromosome length at the time of irradiation.
Breakage disrupts the fundamental structural units that give the chromo-
some its linear continuity; and there is no evidence that the chromatids
and their subsidiary strands have different basic patterns of organization
in heterochromatin and in euchromatin, although there may be con-
siderable differences in the types and proportions of constituent nucleic
acids and proteins. The discrepancy in expression of euchromatin and
heterochromatin as the mitotic-type chromosome becomes transformed
into the salivary-gland type of structure is a problem of differentiation
(see Krivshenko, 1950), and does not necessarily reflect any dissimilarity
in structure at the time of irradiation. All these considerations indicate

CHROMOSOME ABERRATIONS IN ANIMALS 665

that more convincing evidence is needed if the close correlation between
break frequency and mitotic chromosome length is to be regarded as
merely coincidental, especially since cytogenetic studies have shown that
the presumably indivisible blocks in the proximal region of the X may be
broken in a number of different places by X-ray treatment (Kaufmann,
1944). The general conclusion drawn from these various lines of inves-
tigation is that breaks induced in the spermatozoa of Drosophila are dis-
tributed at random among the chromosomes and along their lengths, and
represent a random sample of the number originally induced by the
ionizing radiations.

In the study of the distribution of breaks along the X chromosome of
D. melanogaster it was found that certain intercalary subdivisions, such
as 11 A, 12D, and 12E (Table 9-9; see also Kaufmann, 1944, 1946a), have
high coefficients of breakage (determined by comparing the number
observed with that expected on the basis of length represented in the
salivary-gland chromosome). These regions thereby simulate the
behavior of the proximal heterochromatin, and it is suggested, in the
light of the foregoing considerations, that they also contain hetero-
chromatin. The probability that other intercalary heterochromatic
regions are scattered along the chromosome is indicated by the essen-
tially normal frequency distribution curve obtained by plotting the
coefficients of breakage.

Breaks detected in salivary-gland chromosomes of Sciara after irradia-
tion of oocytes are not distributed at random among chromosomes
(Bozeman and Metz, 1949). The rearrangements, however, are almost
exclusively intrachromosomal (Crouse, 1950), suggesting a recombination
pattern different from that of breaks induced in chromosomes of sperma-
tozoa of Drosophila. The absence, in the chromosomes of most organ-
isms, of the clear pattern of linear differentiation that characterizes the
salivary-gland structure makes precise determination of break position
much more difficult. There are, however, several reports of nonrandom
break distribution. In the ( Jrthoptera some members of the chromosome
set appear to be more susceptible to fragmentation than others (White,
1935b; Bishop, 1942), and some regions of the chromosomes also appear
to be especially fragile (Helwig, 1933, 1938), although they are not
necessarily characterized by any special morphological features (Bishop,
1942). If it is assumed that breaks are distributed at random among and
along these chromosomes, the observed departures from randomness are
to be attributed to the subsequent behavior of the breakage ends with
respect to restitution or recombination, or to the difficulty of detecting all
breaks in later stages of the same mitotic cycle in which they are pro-
duced. These aspects of the problem will be discussed later.

3-ld. Dose-Frequency Relations. Breaks that are detected cyto-
logically represent only a part of those induced by ionizing radiations.

666 RADIATION BIOLOGY

Some of the breaks primarily produced are eliminated by restitution, or
restoration of the original sequence of parts, so that the sites of the
original lesions are not recognizable in the repaired chromosome (see, for
example, Carlson, 1941a; Faberge, 1940; Sax, 1940). Calculations by
Lea and Catcheside (1945), based on a limited number of observations,
indicate that about a third of the breaks induced in the X chromosomes
of spermatozoa of Drosophila by a 3000-r X-ray treatment undergo
restitution. It has been suggested that all recessive lethal mutations not
associated with cytologically detectable chromosomal aberrations repre-
sent restitutional breaks (Lea and Catcheside, 1945 ; Lea, 1946) ; but
Fano (1947) and Muller (1950a) have noted the difficulty of reconciling
this theory with the total data available for the production of recessive
and dominant lethals at different dosage levels.

Breaks that do not undergo restitution are detectable, either because
they remain "open" or "unhealed," or because the breakage ends join
inter se or participate in structural rearrangement. Several lines of
experimental evidence indicate that the detectable breaks are scattered
at random within and among cells, and therefore presumably represent
a random sample of those originally induced.

Single breaks, such as those that separate fragments from the centro-
meric portions of the chromosomes, are produced with frequencies pro-
portional to the dose, at least when the dose is sufficiently low that the
mean number of breaks per cell is less than one (Fano, 1941). The dose-
frequency relation is expressed by the formula N/N0 = kD, in which N
is the number of breaks occurring in iVo cells, D the dose, and k a propor-
tionality constant. When the frequency N/N0 approaches 100 per cent,
the deviations from linearity that occur can be attributed to the produc-
tion of more than one break per cell. The linear relation of breaks to
dose suggests that breakage is due to a single "hit," use of this term being
subject to the various qualifications that have been outlined by Muller,
Fano, and others (summarized in Fano, Caspari, and Demerec, 1950).

The experimental results are well illustrated by Carlson's study (1941a)
of fragment production in neuroblast cells of the grasshopper, Chortophaga
viridifasciata, by X-ray treatment. The distribution of the fragments
among nuclei was found to conform to a Poisson distribution. The
number of breaks was proportional to the amount of the dose within
the range tested, which extended from 7.8 to 125 r (Fig. 9-13a). Appar-
ently there is no threshold below which breaks are not produced; it thus
appears to be an all-or-none effect.

In Drosophila the increase in frequency of dominant lethals induced by
X rays, as determined from the proportions of eggs that fail to hatch, is
essentially linear at lower dosage levels, indicating that the single-break
type of aberration is primarily involved (Demerec and Fano, in Demerec,
Kaufmann, Sutton, and Hinton, 1940; Demerec and Fano, 1944; Catche-

CHROMOSOME ABERRATIONS IN ANIMALS

667

side and Lea, 1945a). At higher dosage levels the frequency increases
more rapidly than the first power of the dose. This deviation from
linearity has been attributed to the dominant-lethal effects of deficiencies
and asymmetrical exchanges involving two or more breaks, which are
produced with higher frequencies at higher doses (Fano and Demerec,

<

Id

CD

u_
O

LlI

o

LlI
CL

0 10 20 30 40 50 60 70 80 90 100 110 120 130

DOSAGE, r

500 1000

1500 2000 2500 3000
DOSAGE, r

3500 4000 4500 5000

— ■ ADULT WASPS FAILING TO EMERGE

DROSOPHILA EGGS OF OREGON R STOCK FAILING TO

HATCH (SONNENBLICK)

DROSOPHILA EGGS OF OREGON R STOCK FAILING TO

HATCH (DEMEREC AND FANO)

ADULT DROSOPHILA OF OREGON R STOCK FAILING

TO EMERGE (DEMEREC AND FANO)

Fig. 9-13. (a) Relation between X-ray dose and the production of single breaks in
neuroblasts of Chortophaga. (From Carlson, 1941a.) (b) Dose-frequency relations
for dominant lethals in Drosophila and Habrobracon. (Data of Demerec and Fano,
1944; Sonnenblick, 1940; Heidenthal, 1945.)

1941; Demerec and Fano, 1944; Lea and Catcheside, 1945; cf. also the
study of Bauer and Lerche, 1943, on the midge, Phryne fenestralis).
Another line of evidence that the frequency of the single-break type of
dominant lethal in Drosophila is directly proportional to dose has been
obtained by Bauer (1942), in a study of the distortion of the sex ratio
resulting from X-ray treatment of spermatozoa carrying a ring-X chromo-
some. In studies of Habrobracon (Heidenthal, 1945; cf. A. R. Whiting,

668 RADIATION BIOLOGY

1945b, P. W. Whiting, 1938a, b), it was found that the proportion of
adult wasps failing to emerge among the offspring of irradiated haploid
males gave a dose-frequency curve closely paralleling that for dominant
lethals in Drosophila. The results are illustrated in Fig. 9-13b.

Dose-frequency relations determined in studies of induced crossing
over have been discussed previously. Related studies concern the fre-
quency of separation of attached-X chromosomes in irradiated cells.

Separation in nonirradiated XXY females involves crossing over with
the Y chromosome, so that the detached X carries either the short or the
long arm of the Y (Kaufmann, 1933). Mainx (1940) found that the
frequency of exceptional flies resulting from separation of attached-X
chromosomes after irradiation of unfertilized females increased approxi-
mately linearly with increased dose in the 1000- to 3000-r range. There
was a marked increase over values expected on the basis of linear propor-
tionality, however, when the dose was increased to 4000 and 4500 r. It
is assumed that at the higher dose a considerable proportion of the
exceptional progeny was derived from eggs in which the X chromosomes
were broken by processes other than the process involved in crossing
over with the Y.

The frequency of production of small deficiencies, detected by genetic
methods, has been reported in some cases as proportional to dose (e.g.,
Muller, 1940, 1941) and in others as approaching a "two-hit" curve
(e.g., Panschin, Panschina, and Peyrou, 1946). Rick (1940) also
reported dose-frequency relations for small spherical fragments induced
by irradiation of Tradescantia microspores which suggest that many of
the rearrangements are produced by a "two-hit " process. In Drosophila
the small deficiencies detected by genetic methods constitute a graded
series, some of which are discernible in salivary-gland chromosomes and
some of which are not (see Slizynski, 1938). From an analysis of the
frequency of deficiencies in the Notch region of the X chromosome of D.
melanogaster, Demerec and Fano (1941) concluded that the longer defi-
ciencies involve two breaks and depend for their production on the pass-
age of two separate ionizing particles. Deficiencies of sections shorter
than 15 bands on the salivary-gland chromosome (or ca. 60 m/x on the
sperm chromosome) presumably involve single events, and therefore
increase in number in proportion to dose. Breakage in these cases may
be visualized as a process whereby the effect of the radiation spreads
along the chromosome for a short distance and produces two uncouplings
rather than one.

Another type of two-break rearrangement that is apparently produced
with a frequency proportional to the dose involves breaks, often in
separate chromosomes, induced by irradiation with neutrons (e.g., the
studies of Giles, 1940, and Thoday, 1942, on Tradescantia). The results
suggest that both strands involved in the exchange are disrupted simul-

CHROMOSOME ABERRATIONS IN ANIMALS

669

taneously by the densely ionizing track of a single recoil proton. Com-
parison with X rays indicated that neutrons produce a higher frequency
of chromosomal aberrations for the same dose. Studies on Drosophila
have not afforded such convincing data as those obtained from the
Tradescantia experiments. The frequency of viable types of aberrations
induced in D. melanogaster spermatozoa was determined by salivary-
gland-chromosome examination at two levels of neutron dosage (Demerec,
Kaufmann, and Sutton, 1942). The frequency at the lower dose did not
differ appreciably from that induced by an equivalent dose of X rays;
but the higher dose of neutrons was less efficient than X rays, as deter-
mined by the frequency of rearrangements and by the total number of
breaks. The types of rearrangements induced by neutrons and X rays
seemed to differ only with respect to the proportion of intercalary defi-
ciencies, which was greater in the neutron than in the X-ray series
(Kaufmann, 1941b). Genetic tests for translocations made by Catsch,
Peter, and Welt (1944) also indicated that neutrons are less effective
than X rays (Table 9-4), whereas similar tests by Dempster (1941b) sug-

Table 9-4. Frequencies of Translocations between the Second and Third
Chromosomes of D. melanogaster Induced by X Rays and Fast Neutrons

(Catsch, Peter, and Welt, 1944)

Frequency of translocations

Dose

No. of sperms
tested

Per cent
translocations

X rays, 1000 r

Neutrons, 1050 r-cq

X rays, 2000 r

Neutrons, 2160 r-eq

X rays, 4000 r

Neutrons, 4480 r-eq

6,016
3,287

5,775
3 , 159

10,223
2,665

1.33 ±0.15
1.07 ±0.18

3.79 ± 0.25
1 . 87 ± 0 . 24

11.09 ± 0.30
4.09 ± 0.38

gested that neutrons are slightly more effective. Differences in results
may stem in part from the difficulty of equilibrating doses of X rays and
neutrons. Marshak (1942) has presented data which suggest that neu-
trons are more effective than X rays in inducing chromosomal changes
in mouse tumors, but diagnosis was based on abnormal anaphase figures,
and presumably included single-event as well as two-break processes.

The frequency of two-break rearrangements induced by X rays, in
which each break would be caused by a separate ionizing particle, would
be expected to increase as the square of the dose in accordance with the
formula N/Nq = k'Dn, in which n indicates the number of independent
events. The observational data for two-break rearrangements conform

670

RADIATION BIOLOGY

in general with this expectation. Inversions, deficiencies, and translo-
cations detected by genetic methods by Timofeeff-Ressovsky (1939) in-
creased in frequency approximately as the square of the dose (Fig. 9-14a).
Data furnished by Makki and Muller (Muller, 1940) and by Catsch,
Radu, and Kanellis (1943; see also Catsch, Peter, and Welt, 1944; Catsch,
1948) for translocations between the second and third chromosomes
induced by X rays in the range from 400-4000 r, can be fitted to a
curve that represents the 1.5 power of the dose. This deviation from
the anticipated second power is attributed to the scoring of multiple-

col 0

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u

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<

r

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LU

yy

o

yy

a:

LU

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i

i

i i

(a)

1500 3000 4500 6000
DOSAGE, r

-• EXPERIMENTAL

1000 2000 3000 4000 5000
(6) DOSAGE, r

o o THEORETICAL

Fig. 9-14. Dose-frequency relations for viable types of chromosomal rearrangements
induced by irradiation of spermatozoa of Drosophila milanogaster. (a) Dose-frequency
relations determined by genetic tests for inversions, deficiencies, and translocations.
Broken line represents the theoretical exponential ("two-hit") curve. The vertical
bars indicate the range of the standard error for each of the experimentally derived
values. (Data from Timofeeff-Ressovsky, 1939.) (b) Dose-frequency relations deter-
mined by cytological tests for chromosomal rearrangements. Exponential curve
represented by broken line and limits of error by vertical bars as in Fig. 9-14a. (Data
from Bauer, 1939c.)

break rearrangements as two-break events, and to the disproportionately
greater chance for the production of inviable recombinations (dominant
lethals) when many breaks are available in the same nucleus.

The frequencies of inversions, deficiencies, translocations, and more
complex rearrangements, as determined by salivary-gland-chromosome
analysis of the progeny of males irradiated at various dosage levels, in the
studies made by Bauer, Demerec, and Kaufmann (1938) and Bauer
(1939b), are indicated in Fig. 9-14b. It will be seen that the frequency of
cells carrying the viable types of rearrangements increases more rapidly
than the first power of the dose, approaching the second power at the
lower levels of treatment.

The frequencies of complex rearrangements detected in these studies
appear to be much lower than expected at low doses, and to increase

CHROMOSOME ABERRATIONS IN ANIMALS 671

less rapidly than would be expected on the general assumptions of the
"hit theory." Data summarized by Fano (1941) are presented in
Table 9-5. Expected frequencies (shown in parentheses) have been
computed on the assumption that the relative frequencies do not depend
on the dose. It will be seen that the relative frequencies of the different
types of changes — two-break, three-break, four-break, of both the 2 + 2
and the 4 type — are nearly constant at all doses. As a possible explana-
tion of deviation from expectancy, Fano (1943b) has suggested that
potential breaks may be influenced, by mechanical perturbations, to
participate in rearrangement whenever two other breaks initiate the
process. There is also the possibility that these mechanical disturbances
may in themselves cause breakage that is not attributable to the action
of the ionizing radiations. Because of such factors it is not possible at
present to formulate any simple theoretical interpretation of the relation
of frequency of complex chromosomal rearrangements to dose.

In all these considerations it must be kept clearly in mind that an
observed rearrangement represents the climax of a series of processes
involving breakage and recombination. Efforts to separate these two
phases experimentally and thereby elucidate the process of structural
rearrangement have included studies of the relation between frequency
of aberrations and such factors as wave length, time, and intensity of the
ionizing radiations. It was shown by Muller and Ray Chaudhuri
(reported in Muller, 1940) that y rays and 50-kv X rays are equally
efficient per roentgen in inducing translocations. Thus 2000 r of 50-kv
X rays produced 2.9 + 0.3 per cent translocations between the second
and third chromosomes of D. melanogaster, and 2000 r of y rays produced
3.8 + 0.6 per cent. These values do not differ significantly, nor do those
obtained by Catsch, Kanellis, Radu, and Welt (1944) in a study of the
frequency of translocations between the second and third chromosomes
induced by doses of 1000, 2000, and 4000 r at wave lengths of 0.5, 0.4, and
0.15 A.

It has also been shown that variation of time or intensity of treatment
does not alter the frequency of viable types of rearrangements induced by
a given dose of radiation (Muller, 1940; Kaufmann, 1941a; Dempster,
1941a; Makhijani, 1945). Muller and Makhijani (reported in Muller,
1940) found by genetic tests that doses of 2000 r given at intensities
ranging from 0.05 to 250 r per minute induced similar frequencies of
translocations between the second and third chromosomes. In these
experiments spermatozoa were treated after they had been transferred to
females in insemination; and the translocation frequencies were similar
for a given dose of X rays, whether it was administered in a single treat-
ment, with immediate or delayed opportunities for egg laying, or in a
series of four treatments. A like absence of effect of discontinuous treat-
ment when males were irradiated was established by cytological examina-

672

RADIATION BIOLOGY

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CHROMOSOME ABERRATIONS IN ANIMALS

673

tion of salivary-gland chromosomes (Kaufmann, 1941a). This method
permits detection not only of rearrangements, but also of their types and
proportions, and of the total number of breaks involved. The results
are given in Table 9-6.

Table 9-6. Comparison of Frequencies of Induced Chromosome Breaks Result-
ing from Continuous and Discontinuous X-ray Treatment of Males

(Kaufmann, 1941a)
(No errors are furnished for the percentage of breaks, since the distribution of the
number of breaks is not given by a Poisson series; 1 X 3000 r and 1 X 4000 r refer to

continuous treatment.)

Dose, r

Total
glands

No. with

chromosomal

aberrations

Percentage
altered

No. of
breaks

Percentage
of breaks

1 X 3000"

595
132

184

271

44

140

184

112

28

43

79
15
39
54

18.82 ± 1.60
21.21 ± 3.50

23.37 ±3.12

29.15 + 2.76
34.09 +7.15
27.86 ± 3.79
29.35 ± 3.36

293

96
227

36
117
153

49 24

1 X 30006

3 X 1000 at 24-hr in-
tervals

^2 17

1 X 40006

83 76

4 X 1000 at 24-hr in-
tervals

2 X 2000 at 16-day
intervals

81.82
83 57

Total discontinuous
treatment

83 15

a Data from Bauer, 1939b.

b Includes data from Bauer, Demerec, and Kaufmann, 1938.

These findings have been interpreted as indicating that breaks produced
by irradiation of the mature spermatozoon do not participate in the
formation of new rearrangements until after the sperm has entered the
egg in fertilization; but, as will be discussed subsequently, this conclusion
can apply only to the viable class of aberrations that are available for
diagnosis by genetical or cytological techniques.

Experiments to determine the effect of differences in temperature on
the production of rearrangements have given conflicting results. The
earlier studies of Papalashwili (1935) and Mickey (1939), which indicated
that low temperature during irradiation results in a higher frequency of
chromosomal rearrangements than high temperature, were not confirmed
by the extensive work of Muller (1940) and Makhijani (1949), in which
similar frequencies of translocations between the second and third
chromosomes of D. melanogaster were obtained when temperatures as
different as 4.5 and 37.5°C were applied either at the time of irradiation
or at the time of fertilization. In another series of experiments (Kauf-
mann, 1946b, 1948a), it was found that there was no significant difference

674

RADIATION BIOLOGY

in frequency of rearrangements or in total number of induced breaks
when flies were exposed to temperatures of 8, 18, or 28°C before, during,
or after exposure to X rays. More recent studies of sex-linked lethals by
King (1947) and of translocations by Kanellis (1946) and Baker (1949)
indicate that irradiation at a temperature in the 0-to-4°C range gives a
higher frequency of aberrations than irradiation at 20 to 32°C. As an
illustration, Fig. 9-15a and b shows the frequencies of exchanges among
the second, third, fourth, fifth, and Y chromosomes of D. virilis deter-
mined at two temperature levels by Baker. The data indicate that not
only the frequency of rearrangements, but also the slopes of the dose-

1000

4000

1000

2000
DOSAGE,

3000 4000

2000 3000
(a) DOSAGE, r ((,)

Fig. 9-15. Dose-frequency relations for viable types of chromosome rearrangements
induced by irradiation of spermatozoa of Drosophila virilis. (a) Number of exchanges
per tested sperm, (b) Minimum number of breaks per tested sperm. The method
of determining the shapes of the theoretical curves presented in these figures is
described in detail in the original article. {From Baker, 1949.)

frequency curves are influenced by the temperature at the time of treat-
ment. Either more breaks are produced when irradiation is carried on at
a low temperature (as Baker concluded), or the properties of breakage
ends of chromosomes are modified so as to increase the number of viable
types of exchange (as Kanellis concluded). Evidence that such modi-
fication can occur has accumulated in recent years, and will be considered
in the next section.

3-le. Qualitative Differences among Breaks. There is considerable evi-
dence that the capacity of breakage ends to participate in recombination
varies greatly in different organisms and types of cells. In some cases,
as in the irradiated eggs of Ascaris, the ends of chromosomes bordering on
induced breaks appear to have no capacity for uniting with one another,
for chromosomal rearrangements do not occur (Bauer and LeCalvez,
1944; cf. the similar studies of Hughes-Schrader and Ris, 1941, and Ris,
1942, on Hemiptera). That differences may exist in the behavior of
breakage ends in different tissues of the same organism was clearly shown

CHROMOSOME ABERRATIONS IN ANIMALS 675

by McClintock (1939). If a broken chromosome is present in the endo-
sperm or gametophyte tissue of maize, union of sister chromatids will
occur at the breakage point, but no such sister-strand reunion occurs in
the tissues of the embryo.

Another aspect of the problem of qualitative differences among breaks
concerns the different fates of the breakage ends produced in cells of a
single type, such as the spermatozoa of Drosophila. Of the breaks pri-
marily induced in a cell by irradiation, some may be restituted, some may
participate in structural rearrangement, and others may remain "open"
or "unhealed." When breakage ends are capable of rejoining, the prob-
ability of restitution or participation in structural change "appears not
to depend on a difference in the breakage process, but mainly on whether
other breaks are available with which interchange can occur" (Lea, 1946).
Existing data suggest that restitution and recombination occur with
approximately equal frequencies (Lea and Catcheside, 1945; Baker,
1949).

If neither restitution nor recombination takes place, it is difficult to
determine whether the breakage ends are different in some way from
other breakage ends, or whether they are capable of joining but have been
prevented from doing so by chance circumstances (Lea, 1947a). Analysis
of the proportions of breakage ends that failed to recombine in experi-
ments treating Tradescantia chromosomes with different types of radia-
tion favored the former of these alternatives (Catcheside, Lea, and
Thoday, 1946a; Lea, 1946). It thus seems probable that some breaks
may be qualitatively different from others with respect to their capacities
for subsequent recombination.

Data obtained in studies of Drosophila suggest that such qualitative
differences in breakage ends may exist from the time of their origin
(Kaufmann, Hollaender, and Gay, 1946; Kaufmann and Wilson, 1949).
These studies involved treatment of spermatozoa with near-infrared
radiation before exposure to X rays. The near-infrared portion of the
spectrum, centering around wave length 10,000 A, was not in itself effec-
tive in inducing chromosome breaks or gene mutations; but when used
before X rays it significantly increased the frequency of viable types of
rearrangements as compared with the frequency in controls receiving
only the X rays. On the other hand, treatment with near-infrared radia-
tion after exposure to X rays did not increase the frequency of rearrange-
ments. The frequency of induced dominant or recessive lethals was not
modified by near-infrared pretreatment or posttreatment. The experi-
mental data are summarized in Table 9-7. Additional posttreatment
data were obtained by Kaufmann and Wilson (1949) from exposure of
females inseminated by X-ray-treated spermatozoa to near-infrared
radiation. As in the experiments summarized above there was no sig-
nificant increase in frequency of rearrangements or percentage of breaks

676

RADIATION BIOLOGY

as compared with values obtained from controls that received only the
X-ray treatment.

Comparable results were obtained when near-infrared radiation was
used in combination with nitrogen mustard in treating spermatozoa; pre-
treatment effected an increase of about 50 per cent in the frequency of
rearrangements, but there was no significant increase when near-infrared
radiation was used after the spermatozoa had been exposed to the
chemical (Kaufmann, Gay, and Rothberg, 1949).

Since pretreatment did not increase the frequency of single-break
dominant lethals, no general increase in number of all types of breaks was
involved in its effect. The action of near-infrared radiation in "sensitiz-

Table 9-7. Effects of Treatment with X Rays and with X Rays plus Near-
infrared Rays
(Kaufmann, Hollaender, and Gay, 1946)

Treatment of males

24-72 hr of infrared + 4000 r
of X rays

4000 r alone

4000 r of X rays 4- 24-72 hr
of infrared

Frequency of chromosomal breaks

Total
sperms

721
549

483

Rearrange-
ments

312
169

133

Per cent
rearrange-
ments

43.27 ± 1.8
30.78 ± 2.0

27.50 ± 2.0

No. of
breaks

850
435

386

Breaks
per 100
sperms

117.9
79.2

79.9

ing" the chromosomes therefore appears to be restricted to the control
of processes involved in the formation of chromosomal aberrations. In
other words, the subsequent behavior of some breakage ends is modified
by conditions created as a consequence of pretreatment, whereas that of
others is not. In explanation of this difference it has been suggested that
the single-break, dominant-lethal aberrations originate at the time of
irradiation of the spermatozoa, whereas the recombination of other break-
age ends is delayed until after fertilization. Supplementary lines of
evidence in support of this interpretation have been presented in detail
by Kaufmann and Wilson (1949).

In the light of these observations, the breaks induced by irradiation
have been classified as "complete," or thoroughgoing, and "potential"
(the latter in conformity with the concept developed by Muller, 1940,
1941; Kaufmann, 1941b; Fano, 1941). Various lines of experimental
evidence indicate that the potentialities of individual breaks to become
restituted or to participate in structural rearrangement can be modified
by various kinds of supplementary treatment (e.g., by treatment of
spermatozoa with radiation of wave length 2537 A after treatment with

CHROMOSOME ABERRATIONS IN ANIMALS 677

X rays, or by exposure of eggs to near-infrared radiation at the time of
their fertilization by X-ray-treated spermatozoa; see Kaufmann and
Hollaender, 1946, and Kaufmann, 1946b). Potential breaks that fail to
establish new contacts during the period of recombination presumably
undergo restitution soon thereafter, although some experimental evidence
suggests that recombination may be delayed until after the first cleavage
mitosis (Heifer, 1940).

3-lf . Break Recombination. The random distribution of breaks among
the chromosomes of Drosophila having been established by analysis of
viable rearrangements, the question remains whether breakage ends unite
with equal freedom to produce inviable types of exchanges. Informa-
tion on this question is available with respect to some of the alternative
possibilities of recombination within and among chromosomes.

Intrachromosomal rearrangements involving two breaks usually even-
tuate as either inversions or deletions (the symmetrical and asymmetrical
types diagramed in Fig. 9-3). Evidence from studies on Drosophila
suggests that deletions are produced about as frequently as inversions of
the same length (Fano, 1941). The frequency of X-chromosome inver-
sions used in this comparison was determined by salivary-gland-chromo-
some analysis; the frequency of X-chromosome deletions, which cannot
be ascertained accurately by that method, was determined by genetic
analysis of the exceptional females produced in a cross between an irradi-
ated wild-type male and a female carrying attached-X chromosomes with
recessive marking genes (Bishop, 1941).

Other analyses of two-break rearrangements indicate that recombina-
tion within a chromosome limb is essentially at random. When inver-
sions were grouped in classes, according to length in number of divisions
within the euchromatic portions of the chromosomes (ranging from 0-18
divisions), the numbers corresponded closely with those expected on the
basis of random recombination (Bauer, Demerec, and Kaufmann, 1938;
Bauer, 1939b). A more intensive study of length of X-chromosome two-
break rearrangements, in terms of subdivisions (ranging from 0 to 113
subdivisions, since each of the 19 divisions has 6 subdivisions), revealed
some departures from the values expected with random recombination
(Kaufmann, 1946a). Rearrangements measuring about 13 subdivisions
or multiples thereof were somewhat more frequent than those of other
lengths. This modal grouping suggests a pattern of coiling within the
X chromosome at the time of recombination that slightly increases the
opportunities for reunion of parts separated by the distance of the turn of
the coil, as compared with regions separated by shorter or longer
distances.

No accurate measurements can be made of the length of rearrange-
ments having one break in the proximal heterochromatic region (as in
division 20 of the X) and one elsewhere along the chromosome (as in

678

RADIATION BIOLOGY

divisions 1 to 19 of the X), because of uncertainty concerning the exact
position of the proximal break. These " heterochromatic-euchromatic "
rearrangements have been used, however, to determine the distribution of
breaks in divisions 1 to 19 of the X chromosome that combine with those
in division 20 (Kaufmann, 1946a). Breakage ends in division 20 com-
bined freely with other broken regions throughout the X; no evidence was
obtained of preferential recombination with the intercalary hetero-
chromatic regions that lie scattered along the X chromosome. Combined
data from this study are shown in Table 9-8.

Table 9-8. Distribution of 143 Breaks in Divisions 1 to 19 of the X Chrom-

20) Compared with Distribution of All

(Kauf-

Division number

Break

1

2

11
7.45

51 *
54.55

3

4

5 6"

7

Observed

5

8.65

67
63 . 35

14
10.93

1 77
80.07

6
9.49

73
69.51

10
10.57

78
77.43

X

8
9.37

70

68.63

2 = 15.5

Expected6

Observed

Expected^

11 Grouping required for x2 determinations.

6 Expected values are based on random recombination.

Salivary-gland-chromosome studies on Drosophila by Catcheside (1938,
1948) and Bauer (1939b) indicate that the frequency of inversions having
both breaks in the same arm of a chromosome (intrabrachial) is not sig-
nificantly different from that of inversions having the two breaks in
different arms (interbrachial) . However, in an extensive series of studies
in which the frequency of rearrangements of Drosopnila having two breaks
within one chromosome arm was compared with that of rearrangements
having one break in one arm and the other break in any of the other arms
of the chromosome complex, it was found that the former is much lower
and the latter is much higher than chance recombination would permit
(Bauer, Demerec, and Kaufmann, 1938, Catcheside, 1938, and Bauer,
1939b, on D. melanog aster; Heifer, 1941, and Koller and Ahmed, 1942, on
D. pseudoobscura). The X-chromosome data presented by Kaufmann
(1946a), for example, show that inversions within the X are about two
and a half times as frequent as expected on the basis of random recom-
bination (see Table 9-9).

The disparity between intra- and interchromosomal exchanges is even
more pronounced when unfertilized eggs are irradiated. It was found,
for example, that X-ray treatment of oocytes of Drosophila or Sciara pro-

CHROMOSOME ABERRATIONS IN ANIMALS

679

dueed inversions but practically no translocations (Glass, 1940; Bozeman,
1943; Grouse, 1950).

Two breaks in different chromosomes may also conceivably lead to the
formation of an asymmetrical exchange, for example, a dicentric chromo-
some and an acentric fragment (Figs. 9-3 and 9-4). Since such types
would not be perpetuated in Drosophila for salivary-gland-chromosome
analysis, a comparison cannot be made in that organism between the fre-
quencies of the viable or symmetrical two-break interchromosomal
exchanges and their asymmetrical counterparts. From studies on

osome Utilized in Exchanges with the Proximal Heterochromatin (Division
1048 Breaks Recorded in These Divisions
mann, 1946a)

distribution

8

9 10"

11

12

13

14 15"

16

17 18"

19

Total

8
8.17

60

59 . 83

N = 14

9
13.45

103
98.55
P = 0

12

11.89

87
87.11
.346

9
11. .17

84
81.83

10
7.57

53
55.43

9
8.89

65
65.11

15

8.89

59
65.11

12
9.61

68
70 . 39

5

6.97

53
51.03

143
143.07

1048
1047.93

Tradescantia it appears that the symmetrical and asymmetrical types
occur with about equal frequencies (Lea, 1946). In Carlson's study of
neuroblast chromosomes of Chortophaga, 14 chromatid interchanges were
analyzable, of which 5 were symmetrical and 9 asymmetrical.

Analyses of patterns of recombination have also been made for the
three- and four-break rearrangements. In computing expected values,
the proportion of viable to inviable types must be considered for each
possible type of break distribution (Bauer, 1939b; Kaufmann, 1946a).
Among the viable three-break exchanges, observed frequencies con-
formed closely to values expected on the basis of random recombination
(Bauer, 1939b, as shown in Table 9-10). There was in this study, how-
ever, and also in that of Kaufmann (1946a), a disproportionately large
number of rearrangements with all three breaks in the same chromosome
limb (in the latter, eight in the X as compared with the expected 2.5).
Among the four-break rearrangements, the type having two independent
exchanges (2 + 2) was more frequent than expected, and the single cyclic
rearrangement (4) much less frequent. Although the number of rear-
rangements falling into each of these two classes was small, the breaks
utilized in the rearrangements were distributed among the chromosomes
with frequencies that corresponded fairly well with random recombina-

680

RADIATION BIOLOGY

tion, as is illustrated by the data of Kaufmann (1946a) presented in
Table 9-11.

The low incidence of rearrangements of each of the complex types
involving five or more breaks precludes their analysis on a similar basis,
although Fano (1943b) and Kaufmann (1943) have shown that a con-
centration of the breaks in one or two limbs often occurs in these multiple-
break rearrangements. Fano suggested that in the recombination proc-

Table 9-9. Frequency of X-chromosome Inversions (X/X) and of Trans-
locations BETWEEN THE X AND THE AUTOSOMES (X/A)
(Data listed in first four lines involve only breaks in divisions 1-19, data in fifth
line involve breaks in division 20. Expected values are calculated on basis of random
recombination among five long limbs.)

(Kaufmann, 1946a)

Numbers of inversions and translocations

x2

Source material

X/X

X/A

P

Observed

Expected

Observed

Expected

All 2-break rearrange-

ments0

87

34.22

305

357.78

89.2

<0.0001

All 2-break exchanges6

131

48.36

423

505 . 64

154.7

<0.0001

2-break exchanges

with at least 1 X

break in intercalary

heterochromatim. . .

53

23.70

111

140.30

42.3

<0.0001

2-break exchanges with

X breaks in inter-

calary euchromatinrf

78

26.60

312

363 . 40

106.6

<0.0001

2-break exchanges with

1 break in division

20

53

24.90

142

170.10

36.4

<0.0001

° From nuclei having only one inversion or one translocation.
b From more complex rearrangements in addition to 2-break rearrangements.
c Regions included are IF, 3C, 4A, 4E, 7B, 9A, 11 A, 12D, 12E, 16F, 19E.
d X-chromosome breaks in subdivisions not listed in preceding line.

ess adjustments among the chromosomes originally involved may cause
mechanical disturbances of sufficient intensity to produce new breaks,
which then participate in the development of the complex rearrangement.
Another possibility, which does not require the participation of mechan-
ically induced breaks, is that heterochromatic breaks initiate the process
of recombination on an intrabrachial level, and the developing rearrange-
ment then incorporates adjacent potential breaks. This alternative is
based on the suggestion of Kaufmann (1946a) that potential breaks in

CHROMOSOME ABERRATIONS IN ANIMALS

681

Table 9-10. Distribution among the Chromosomes of Breaks Utilized in

3-break Rearrangements
(Bauer, 1939b)

Break distribution

1:1:1

2:1

3

Observed

14
13.89

25

30

27.79
30.

5

Expected

2.32
11

x2 = 0.005 N = 1 P = ca. 0.95

Table 9-11. Distribution of Breaks Utilized in 4-break Rearrangements

(Kaufmann, 1946a)

Distribution among the chromosome limbs

1:1:1:1

2:1:1

2:2

3:1

4

Single-contact 4-break
rearrangements

Observed
Expected

3
5.35

23

28.07

6
4.46

12
5.94

0
0.19

Independent 2+2
exchanges

Observed
Expected

x2 = 8.

19

19.11

10 N =
44
42.99

3 P = (

9
7.96

6
)044
8
10.61

13

1
0.33

x2 = (

)50 N --

= 3 P =

10.94
ca. 0.92

heterochromatin may be able to initiate the production of rearrangements
somewhat sooner than breaks in euchromatin.

4. DIFFERENCES IN SENSITIVITY TO IONIZING RADIATIONS

On the basis of the more general considerations of the process of
induced structural rearrangement outlined in the preceding pages, some
attention can be given to the factors responsible for differences in sensi-
tivity of cells to ionizing radiations.

It has been recognized since the early years of the century that the
X-ray sensitivity of a tissue is related to its reproductive capacity
(Bergonie and Tribondeau, 1906). It has also been reported that cells
with a short mitotic cycle and intermitotic period are more sensitive to
radiation and suffer greater damage than those with a longer cycle
(Roller, 1947; Knowlton and Widner, 1950). Since high reproductive

682 RADIATION BIOLOGY

activity of a tissue indicates that many of its cells are dividing, it seems
probable that cells in the course of mitosis are especially sensitive to the
deleterious effects of radiations. This is, in fact, clearly demonstrated by
experiments which show that a much larger dose of radiation is required
to kill a cell immediately than to cause its death at or after its next divi-
sion. As an example, 2500 r or more is required to kill chick tissue-
culture cells while they are in resting stages, but irradiation of these
cells with 100 r is sufficient to kill a high proportion of the cells when they
subsequently attempt to divide (Lasnitzki, 1943a, b). Irradiation of
Pandorina, a colonial member of the Volvocales, with doses of X rays
ranging from 3000 to 300,000 r did not kill the cells immediately, but
death occurred when they subsequently attempted to divide (Halber-
staedter and Back, 1942). In another study, hematopoietic cells of tad-
poles of the bullfrog exposed to 500 r of X rays showed visible damage
only upon entering prophase (Schjeide and Allen, 1950). Some of the
latent damage that causes cell destruction during division in these cases
is presumably due to chromosomal injury. Many of the physiological
disturbances effected in tissues and organisms by ionizing radiations have
also been referred to alterations in the structure of chromosomes. In
determining the basis for differences in the response of tissues and cells to
ionizing radiations it thus becomes essential to ascertain to what extent
the observations of such differences are dependent on differences in break-
ability of chromosomes, to what extent on opportunities for recombina-
tion of breakage ends, and to what extent on experimental procedures
that permit identification of aberrations more readily in some types of
cells than in others.

4-1. RELATIVE SENSITIVITY OF DIFFERENT ORGANISMS

A few selected illustrations will serve to indicate the considerable differ-
ences in frequencies of radiation-induced chromosomal aberrations in
different species. About 24 r of X rays will induce breaks in 1 per cent
of the chromosomes in the neuroblast cells of Chortophaga (Carlson,
1941a). About 400 r is required to produce breaks in 1 per cent of the
X chromosomes of the spermatozoa of Drosophila, as determined by fre-
quencies of dominant lethals (Fano and Demerec, 1941 ; Pontecorvo,
1942). At the other extreme it was reported by Cleveland and Day
(cited in Sax and Swanson, 1941) that doses of X rays in the range
between 3000 and 20,000 r did not produce any chromosome aberrations
in the protozoon Holomastigotoid.es.

Blumel (1950) reported a difference in the survival of eggs of D. virilis
and D. melanogaster exposed to comparable doses of /3 radiation from
P32 combined in H3P04 added to the culture medium. Pairs of mature
flies were placed in shell vials, and the cultures were subsequently
examined. Few flies hatched in the cultures of virilis, many in the

CHROMOSOME ABERRATIONS IN ANIMALS 683

melanogaster cultures. Parallel genetical and cytological studies of
fertile progeny in the virilis cultures indicated that radioactive P32 pro-
duces both mutations and chromosomal aberrations. The melanogaster
flies were not tested for mutations.

There are some reports that the rate of occurrence of aberrations may
differ in different stocks of the same species. For example, the frequency
of dominant lethals was found to be higher in the Oregon-R strain of
D. melanogaster than in the Swedish-b strain when both were exposed
simultaneously to X rays (Demerec, Kaufmann, and Hoover, 1938;
Dempster, 1941b). In a more recent study, however, Demerec and Fano
(1941) questioned the statistical significance of the observed differences
because of the considerable range of variability from experiment to
experiment in studies of frequencies of dominant lethals.

4-2. EFFECT OF PLOIDY

It has been shown in several experiments that the radiation sensitivity
of a cell is related to the number of sets of chromosomes it contains.
Haploid microspores of Tradescantia are about twice as sensitive as
diploid microspores with respect to the production of chromosomal aber-
rations by a given dose of X rays (Sax and Swanson, 1941). Tetraploid
seeds of barley are more resistant than diploid seeds to the effects of
irradiation, as determined by growth rates and vigor (Miintzing, 1941).
Haploid yeast plants are more sensitive than diploids, as determined by
rates of survival (Latarjet and Ephrussi, 1949). In contrast with these
results, it was reported by Lamy and Muller (1939) that the mortality
rates are not significantly different in triploid and diploid embryos of
Drosophila exposed to X rays. From this it was concluded that the
lethal effects of X rays on embryos must be "physiological" rather than
genetic in nature. However, it has been shown that in another insect,
the wasp Habrobracon, haploid male larvae are more sensitive than
diploid female larvae (Whiting and Bostian, 1931); and these observa-
tions have recently been extended by Clark and Kelly (1950). Lethal
effects were determined by Clark and Kelly by measuring rates of eclosion
after treatment of prepupae and pupae. Haploid males were found to
be more sensitive than either diploid males or diploid females.

4-3. RELATIVE SENSITIVITY OF CHROMOSOMES IN DIFFERENT TYPES

OF CELLS OF THE SAME SPECIES

The frequencies of aberrations induced by X-ray treatment in various
types of cells of the plant Tradescantia were determined by Sax and
Swanson (1941). The order of sensitivity in the cells examined, begin-
ning with the most sensitive, was as follows: microsporocytes, micro-
spores, root-tip cells, and the generative cell. Microspores and root-tip
cells of Allium were less sensitive than those of Tradescantia. Differ-

684

RADIATION BIOLOGY

ential sensitivity in these cases was attributed to differences in chromo-
some development, in speed of nuclear changes, and especially in degree of
freedom and capacity for chromosome movement.

Marshak and Bradley (1945) found that the chromosomes of the
Walker 256 rat carcinoma were more sensitive to X rays and neutrons
than those of a rat lymphosarcoma throughout the greater portion of the
resting stage. They concluded that there are physiological differences in

Table 9-12. Frequency of Translocations among Spermatozoa Utilized in
Insemination at Different Periods of Time after Irradiation

(Catsch and Radu, 1943)

Days

after

treatment

Total
sperm
tested

No. with
aberrations

Percentage of
aberrations

1

2559

306

11.96 +0.64

2-7

1483

92

6.21 ±0.63

8-13

1133

78

6.89 ± 0.75

14-19

1532

21

1.37 ± 0.30

20-25

1028

1

0.10 ± 0.10

Table 9-13. Frequency of Dominant Lethals among Spermatozoa Utilized in

Insemination at Different Periods of Time after Irradiation

(Timofeeff-Ressovksy, 1931)

Days

after

treatment

No. of

eggs
laid

No. of

larvae

emerging

Percentage

of egg
mortality

10-15

15-30

Control

1829
2172
4763

437
1463
3738

76.1
32.6
14.5

the chromosomes of two different tissues (lymphoid and epithelial) of the
same species.

In such experiments the aberrations were scored during a later stage of
the same mitotic cycle in which the cells were exposed to the ionizing
radiations. In other cases, where the method of assay determined the
frequency of aberrations surviving a series of cell generations, as in
Drosophila studies, the possibility must be considered that the survival
value may differ in different types of cells. When males of Drosophila
are irradiated, the frequency of induced rearrangements is fairly uniform
among spermatozoa transferred in the earlier copulations, but signifi-
cantly lower among those transferred in later matings. Typical data for
chromosomal rearrangements are shown in Tables 9-12 and 9-13; similar
data for sex-linked lethals have been summarized and discussed by
Schultz (1936).

CHROMOSOME ABERRATIONS IN ANIMALS 685

The marked reduction in frequency after 12 to 15 days has been
attributed to the fact that in actively copulating males the spermatozoa
that were mature at the time of irradiation are used up, and are replaced
by cells that then existed as spermatocytes or spermatogonia. Accord-
ing to this explanation, these cells are subject to germinal selection; that
is, they are eliminated from the germ line if they carry aberrations
that prove cell-lethal, and as a result the spermatozoa maturing after
irradiation are for the most part free of chromosomal rearrangements.
Although this theory may seem adequate to account for the decline in
frequency of aberrations, it does not take into consideration the fact that
the viable types of exchanges such as translocations and inversions, which
are perpetuated through cleavage and embryonic mitoses if induced in
mature spermatozoa, are eliminated if induced in spermatogonia or
spermatocytes. Histological examination of irradiated testes of Dro-
sophila showed some necrotic apical cells to be present, but no necrosis
of spermatogonia, spermatocytes, or spermatids was seen (Pontecorvo,
1944). On the basis of this observation the conclusion was reached that
no germinal selection takes place. Lea (1947b) suggested, however, that
germinal selection operates among spermatogonia. He attributed the
decline in dominant lethals in later matings to the elimination of affected
germ cells in spermatogonial stages. This explanation implied that
mature and immature germ cells are equally sensitive with regard to the
induction of the dominant-lethal type of aberration, and Lea stated that
there is little basis for assuming that chromosomes in spermatogonia are
less sensitive to lethal changes than those in the sperm. As regards the
induction of viable types of chromosome change, on the other hand, this
author concluded that "spermatogonia are much less sensitive than
mature sperm" (Lea, 1947b).

Because of this series of conflicting interpretations, the problem of
germinal selection and relative sensitivity of different stages in the male
germ line of Drosophila requires further consideration. Clarification may
possibly be obtained through the study of chemical-induced rearrange-
ments, since it was shown by Kaufmann, Gay, and Rothberg (1949) that
the frequency of translocations induced by nitrogen mustard treatment
of Drosophila males is higher among spermatozoa transferred 13-18 days
after treatment than among those utilized in earlier matings.

Differences in radiation sensitivity of cells of the testis of the grass-
hopper, Decticus, were reported by Cocchi and Uggeri (1944). Treat-
ment of males with moderate doses of X rays caused chromosomal damage
that could be detected after a few days, whereas high doses were required
to effect more rapid necrosis. A dose of 25 r affected cells in telophases
of the last spermatogonial division, the damage being detected by pyc-
nosis of these cells or of the primary spermatocytes into which they were
transformed. A dose of 100 r affected all secondary spermatogonia.

686

RADIATION BIOLOGY

Much larger doses were required to damage all spermatocytes and
spermatids; and primitive spermatogonia, mature spermatozoa, and the
epithelial cells of the testis were found to be even more resistant.

Differential sensitivity of cells of rodents in various stages of spermato-
genesis has been determined by fertility tests and by histological examina-
tion of the irradiated testes (see, for example, Snell, 1935, 1941; Snell and
Ames, 1939; P. Hertwig, 1938, 1941; Henson, 1942; von Wattenwyl and
Joel, 1941, 1944). Histological examination of testes of irradiated mice
by P. Hertwig (1938) showed that the sensitivity of the germ cells to
radiation decreased with their increasing maturity. The youngest cells
disappeared first, and the mature spermatozoa last (Table 9-14). Similar

Table 9-14. Disappearance and Reappearance of Various Types of Spermato-
genous Cells in the Irradiated Testis of the Mouse

(P. Hertwig, 1938)

Days after irradiation

Stage of development

2

4

6

7

9

11

13
X

16
X

*

18
X

21
X

*

26

X

*

32

X

*

39
X

*

46

Spermatogonia. . ....

X

0

*

Young spermatocytes

X

X

*

X

*

0

0

0

X

*

X

*

X

*

X

*

Large spermatocytes

X

X

*

X

*

X

X

*

*

0

*

X

*

X

*

X

*

Meiotic divisions

X

X

*

X

*

X

X

*

X

*

0

*

*

X

*

X

*

Prospermatids

X

X

*

X

*

X

X

*

X

*

0

*

X

*

X

*

Spermatids

X

X

*

X

*

X

X

*

X

X

*

X

*

X

0

*

X

*

Spermatozoa

X

X

*

X

*

X

X

*

X

X

*

X

*

X

X

*

0

*

0

X

*

The symbol X indicates the presence of cells after a treatment of 800 r; the symbol *
indicates the presence of cells after a treatment of 200 r; the symbol 0 indicates the
occasional appearance of the cells in question. Testes in the 200-r series were fixed
only 4, 6, 9, 13, and 21 to 46 days after irradiation.

results were obtained in studies in which mice were irradiated by injection
of P32 in isotonic saline (Warren, MacMillan, and Dixon, 1950). Females
inseminated by irradiated spermatozoa often produce abnormally small
litters (see, for example, Snell, 1935; Brenneke, 1937), which suggests
that the ionizing radiations induce dominant-lethal chromosomal aberra-
tions. Among the viable progeny fertility may be reduced. Snell (1935,
1941) attributed such semisterility to the presence of chromosomal aber-
rations, induced in the paternal gamete (see also P. Hertwig, 1941;
Roller and Auerbach, 1941).

CHROMOSOME ABERRATIONS IN ANIMALS G87

P. Hertwig (1941) found that doses of radiation sufficient to inhibit
spermatogenesis led to temporary sterility when the supply of mature
spermatozoa had been exhausted. When fertility was restored, the
newly formed spermatozoa produced litters of approximately normal
size. This result suggests either that few chromosomal breaks are
induced in the primordial spermatogonia, or that germinal selection in
the course of spermatogenesis eliminates most of the cells with chromo-
somal aberrations.

Evidence of differences of sensitivity in germ cells of female mice at
different stages of development was presented by Murray (1931). After
exposure to 150 r of X rays, all primary follicles disappeared within two
days, and no follicles were seen after 43 days. Details of these various
experiments and the extensive literature pertaining thereto have been
summarized by Glucksmann (1947).

Another aspect of the general problem concerns the relative sensitivity
of male and female gametes. Metz and Boche (1939) found that the
chromosomes of the spermatozoon and the oocyte of Sciara responded
differently to X rays. Numerous rearrangements were found among the
progeny of irradiated males, but none among the offspring of treated
females. Subsequent studies have shown that this difference is not a
matter of sex, but depends on the condition of the chromosomes at the
time of treatment (Crouse, 1950) . In Drosophila, dominant-lethal effects
may be induced by irradiation of the eggs. When oocytes are irradiated
before fertilization, a higher proportion of the zygotes fail to hatch than
when spermatozoa are irradiated with an equivalent dose (Sonnenblick,
1940) ; but as Glass (1940) has shown, irradiation of eggs produces many
fewer chromosomal aberrations — inversions but no translocations — than
irradiation of sperm. In the gnat, Phryne fenestralis, irradiation of
oocytes apparently produces only the dicentric single-break type of
aberration, whereas irradiation of spermatozoa also produces multiple-
break recombinations (Bauer and Lerche, 1943).

Such comparisons have at times been interpreted as indicating that
some of the dominant-lethal effects produced by irradiation of eggs may
not be attributable to induced chromosomal alterations (Muller, 1938).
Lea (1946) suggested, however, that dominant lethals induced in unferti-
lized eggs, as well as in sperm, can be explained by structural alterations of
chromosomes, if it is assumed that the probabilities differ in sperm and
egg that a breakage end will join with another breakage end in preference
to undergoing sister-union. Similar calculations suggest that chromo-
some aberrations are largely responsible also for lethals induced in
fertilized eggs (see Lea and Catcheside, 1945; Lea, 1946; Haldane and
Lea, 1947). It is suggested that if primary breaks occur at random, and
if joinable breaks unite at random, differences in the frequencies of viable
types of exchange induced by a given dose of radiation in mature sperms

688 RADIATION BIOLOGY

and eggs depend on the proportion of the primary breaks that are incapa-
ble of restitution or participation in exchange and accordingly eventuate
as dominant lethals. If this proportion is high, the treated cells will
readily be inactivated by radiations and will yield few structural
rearrangements (as happens when mature eggs are irradiated). If the
proportion is low, the cells will be less sensitive to the dominant-lethal
effects of radiation and will yield more structural rearrangements (as
happens when mature spermatozoa are irradiated).

4-4. CHANGES IN SENSITIVITY OF CHROMOSOMES
IN CELLS OF THE SAME TYPE

4-4a. Effect of Aging. The effect of age of mature spermatozoa on the
frequency of induction of dominant lethals has been studied in D. melano-
gaster. Mature spermatozoa present in the testes at the time of emer-
gence will be retained if the flies are withheld from copulation. By storing
males, and controlling the time of irradiation, it was found that in early
developing males the sensitivity of the chromosomes to irradiation, as
determined by dominant-lethal counts of eggs, increased with increase in
age (Str0mnaes, 1949; cf. Dempster, 1941b). It has been suggested that
this dependence on age of sperm and rate of development of the males
may explain the discrepancies between the dose-frequency relations for
dominant lethals reported by different workers (Str0mnaes, 1949).

4-4b. Effect of Stage of Mitosis. In 1906 Krause and Ziegler found evi-
dence that cells were most sensitive to irradiation at the time of organiza-
tion of the equatorial plate. During the second quarter of the century
extensive data assembled by a number of investigators suggested that
chromosomes were most sensitive to breakage during interphase or early
prophase stages. In some cases, studies of the same type of cell in the
same species led to different conclusions about relative sensitivities at
different mitotic phases. Obviously, the experimental methods were not
uniform, and no standard method had been developed for assessing the
full extent of radiation damage to the chromosomes. In more recent
years, critical studies of induced chromosomal aberrations have led to
conclusions agreeing with those of Krause and Ziegler. The extensive
literature bearing on the question has recently been surveyed by Whiting
(1945a), Bozeman and Metz (1949), and Sparrow (1951). Our attention
here will be restricted to a few illustrations of differences in sensitivity at
different phases of the mitotic cycle, as indicated by the production of
chromosomal aberrations.

Information concerning relative sensitivities of chromosomes at differ-
ent stages of gametogenesis has been obtained in studies of Habrobracon.
In this wasp, cells in various stages of oogenesis are arranged seriatim
along the length of each of the four ovarioles. Criteria have been devel-
oped for determining whether the chromosomes of eggs at the time of

CHROMOSOME ABERRATIONS IN ANIMALS 689

irradiation are in first meiotic metaphase, or in prophase. Eggs are
permitted to develop parthenogenetically so that the effect of the treat-
ment can be detected directly by its influence on hatchability (Whiting,
1940, 1945a). The results obtained in such experiments indicate that
eggs whose chromosomes are in metaphase of the first meiotic division
are much more sensitive than those in prophase stages of this division.
Embryonic abortion in these cases is presumably due in large measure to
single-break dominant-lethal chromosomal aberrations (Whiting, 1945b).
Germinal selection cannot explain the observed differences, since the
eggs continue meiosis and initiate the cleavage mitoses at rates not
noticeably different from the controls, after doses greatly exceeding those
necessary to inhibit embryonic development.

Apparently the situation in Drosophila is similar, although the stage of
development of the egg at the time of treatment cannot be determined
with as great precision as in Habrobracon. Patterson, Brewster, and
Winchester (1932) found a higher proportion of dominant lethals among
eggs laid within 24 hours after irradiation than among those laid on the
third day, which presumably were in an earlier stage of oogenesis at the
time of treatment. Sonnenblick (1940) also noted that the most mature
oocytes are especially sensitive to irradiation.

In Sciara the sensitivity of the oocyte increases as it passes from late
prophase to metaphase and anaphase, as determined by the percentage
of rearrangements detected and by egg hatchability (Reynolds, 1941;
Metz and Bozeman, 1942). In these fungus flies all oocytes develop
synchronously, so that the meiotic stage at the time of irradiation can
readily be distinguished. More detailed information regarding sensi-
tivity in *S. ocellaris is furnished in a later paper (Bozeman and Metz,
1949). Sensitivity is almost zero before the breakdown of the nuclear
membrane at the beginning of the first meiotic division. It then rises
rapidly to a peak in anaphase, and drops off in late anaphase, when
mitotic activity is arrested pending fertilization. The types of induced
rearrangements include inversions, duplications, deletions, and trans-
positions, but almost no translocations. Summarized data based on
salivary-gland-chromosome analyses are presented in Fig. 9-16.

The most exhaustive studies of differences in sensitivity during the
nuclear cycle have been made by Sparrow (1944, 1948, 1951) on Trillium.
Meiotic divisions are generally well synchronized in the anthers of the
buds of this plant. The stage during which irradiation was carried out
could thus be determined with a high degree of accuracy, and the effects
of the treatment could be studied during the course of meiosis or in either
of the two postmeiotic mitoses. Determination of fragment number in
successive mitoses gave a more adequate estimate of the frequency of
induced aberrations than had been possible with methods previously used.
Since other investigators did not use this double scoring technique, their

690

RADIATION BIOLOGY

data do not represent total sensitivity, but only a portion of the induced
effect. Breaks that are not perceptible in the condensed chromosome
immediately after treatment may appear after an intervening interphase
(as Reynolds, 1941, also showed in Sciara). Results of Sparrow's studies
showed that sensitivity increases during prophases of the first meiotic
division, reaching a peak in the interval between diplotene and first

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PHASE PHASE PHASE

Fig. 9-16. X-ray sensitivity during various stages of first meiotic division, as deter-
mined by salivary-gland-chromosome analysis of Fi progeny of irradiated Sciara
females. Left column (diagonally striped) represents calculations based on wild
stock; open column at right, based on yellow stock; solid black column, based on
totals of both stocks. (From Bozeman and Metz, 1949.)

metaphase, and then falls off through the stages of the second meiotic
division, to reach a low in the postmeiotic interphase (Fig. 9-17). More
recent studies by Sparrow and Maldawer (1950) indicate that part of the
observed difference in sensitivity is due to a difference in the frequency of
recombination of breakage ends, which is significantly higher in cells
irradiated in early interphase than among those irradiated in first meiotic
metaphase. However, these authors state that this difference in recom-
bination is not sufficient to account for the aforementioned differences in
frequency of fragments. The recent results thus indicate that chromo-
somes in the condensed form have a high proportion of "potential"
breaks, which subsequently produce chromosomal aberrations. These
results stand in vivid contrast to many of the earlier plant studies, from
which it was inferred that chromosomes are most sensitive to breakage in
resting or early prophase stages.

The factors responsible for changes in sensitivity during the cycle of

CHROMOSOME ABERRATIONS IN ANIMALS

09 1

mitosis cannot at this time be clearly denned. Movements of chromo-
somes incident to the coiling of the chromonemata and formation of the
equatorial plate may be of importance in modifying the reaction system.
As mitosis progresses, nuclear membrane and nucleoli normally disappear
and are later re-formed. Profound changes are thus effected in the
structure of the chromosome and its cellular environment. Chemical
changes that have been detected within chromosomes and nucleoli in
the course of these events involve the types, proportions, and patterns of
association of nucleic acids and proteins. Very little is known about the

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STAGE IRRADIATED
A SCORED AT ANAPHASE I • TOTAL FRAGMENTATION INDUCED
O SCORED AT MICROSPORE METAPHASE
Fig. 9-17. Graph showing the numbers of fragments induced by 50 r of X rays at
various stages of the nuclear cycle during microsporogenesis in Trillium erectum.
(From Sparrow, 1951.)

cyclic alterations in associated cellular materials. Structural and func-
tional modifications of cellular components may be effected by a multi-
tude of physical and chemical agents (listed in Sparrow, 1951). To the
extent that such modifications affect the reaction system initiated by ion-
izing radiations, the agents used will alter the radiosensitivity of cells.

5. CHEMICAL AND CYTOCHEMICAL STUDIES2

Numerous attempts have been made in recent years to correlate the
cytologically detectable effects of ionizing radiations with changes in

2 This section has been prepared in collaboration with Miss Helen Gay, to whom the
author expresses his sincere thanks.

692 RADIATION BIOLOGY

the chemical composition of cellular materials. The experimental pro-
cedures have involved cytochemical study of irradiated cells and chemical
analysis of materials extracted therefrom. Supplementary studies have
been made on nucleic acids and proteins that were irradiated after extrac-
tion from the cells. Attention has been focused, in all these studies, on
nucleic acids and proteins because they are considered to be the major
structural components essential to the synthetic and reparative processes
of the cell. It has been established that ionizing radiations effect
changes in the physical and chemical properties of these materials, and
in their quantities and proportions. The more important question of
the alterations effected in their patterns of organization remains largely
unexplored.

Cytochemical methods have shown that X rays and 7 rays, within the
dosage range of 40-4000 r, produce metabolic disturbances in prolifer-
ating and undifferentiated cells that are characterized by inhibition of the
synthesis of desoxyribonucleic acid in the nucleus, and accumulation of
ribonucleic acid, mainly in the cytoplasm (Mitchell, 1940, 1942, 1943,
1944). The two types of nucleic acid were identified in these studies by
digestion of the cells with the enzymes ribonuclease and desoxyribo-
nuclease. Determinations of the amounts of nucleic acid were made by
a method of ultraviolet photomicrography. Comparable areas of non-
irradiated and irradiated tissues were photographed with wave length
2537 A, which is absorbed by the purine and pyrimidine bases of the
nucleic acids. The sections were then digested with one of the nucleases
and rephotographed. Differences in the amount of blackening of the
photographic plate afforded an estimate of changes effected by irradiation
in the amounts of the nucleic acids.

Inhibition of the synthesis of desoxyribonucleic acid by ionizing radia-
tions has been confirmed in other experiments. Cytochemical studies
have used spectrophotometric methods to determine the amount of
nucleic acid in irradiated and control sections of the same tissue. Absorp-
tion of ultraviolet rays (wave length 2537 A) by purines and pyrimidines
was determined on unstained preparations (Ely and Ross, 1948a, b), and
absorption of visible light (around wave length 5460 A) was determined
on preparations stained by the Feulgen reagent (Stowell, 1945; Petrakis,
Ashler, and Ferkel, 1949). Radioactive phosphorus has also been used
to determine the amount of newly formed desoxyribonucleic acid in
X-ray-treated and nonirradiated tissue (e.g., studies by von Euler and
von Hevesy, 1942, 1944; Hevesy, 1945 — see Table 9-15; and Holmes,
1947). Hevesy (1945) reported that doses of X rays at or above the
therapeutic level inhibited the formation of desoxyribonucleic acid about
equally in growing and differentiated tissue. Approximately 75 per cent
of the inhibiting action disappeared within 2 hours after irradiation.

Inhibition of synthesis of desoxyribonucleic acid after irradiation has

CHROMOSOME ABERRATIONS IN ANIMALS

693

been attributed to failure of conversion of ribose into desoxyribonucleic
acid (Mitchell, 1940, 1942, 1943, 1944), since ribonucleic acid accumu-
lates in the irradiated cell. The greatest accumulation seems to be in the
cytoplasm, but an increase has also been detected in the nucleus by
Mitchell (1944), and by Roller (1947), who noted that nucleoli, which
are known to contain ribonucleic acid, are formed precociously in tumor
cells after their irradiation.

In contrast with the foregoing observations, there have been some
reports that the amount of ribonucleic acid in the cell decreases after
its irradiation (e.g., Holmes, 1947; Ely and Ross, 1948a, b). The prob-
lem was examined by Petrakis et al. (1949), who made a quantitative

Table 9-15. Ratio of Newly Formed Desoxyribonucleic Acid Molecules
before and after irradiation of jensen's sarcoma of the rat

(von Hevesy, 1945)

Dosage, r

Time interval

between

irradiation

and injection

Time elapsed
between

injection and
sacrificing
rat, hours

Ratio of newly
formed nucleic

acid in the

controls and in

the irradiated

sarcoma

750-1500
335-1500
450-1500

1500
1230-1500

Few minutes
Few minutes
Few minutes
Few minutes
3-7 days

0.5

1

2
4-6

2

3.2
2.4
2.2
2.8
1.7

histochemical study of rat-liver epithelium over a 6-day period after
irradiation of the animals with either 600 or 1500 r. The results of these
investigators showed that the detectable amounts of ribonucleic acid
varied with the dose and the interval of time elapsing between irradia-
tion and fixation of the tissue. After the 600-r dose there was an increase
during the first 24 hours in the concentration of ribonucleic acid in cyto-
plasmic granules and nucleoli, as determined by their stainability with
the basic dye methylene blue; but subsequent examination indicated that
a marked decrease occurred between the second and sixth days. After
the 1500-r dose, there was a decrease in cytoplasmic basophilia during the
first 3 hours, followed by an increase during the next 3 hours to about 40
per cent in excess of the control value, and again a marked decrease
detectable at 24 hours after irradiation. Similar results, indicating that
the concentration of ribonucleic acid depends on the dose and on the time
interval between treatment and fixation of the cell, have been obtained
by the author and his associates in cytochemical studies of sections of
irradiated plant and animal tissues stained with various basic and acidic
dyes (unpublished data). In these studies the ribonucleic acid was

694

RADIATION BIOLOGY

identified by using purified, crystalline ribonuclease. It is suggested that
the differences in the reports of various workers concerning the accumula-
tion or loss of ribonucleic acid after irradiation may be due in part to
differences in dosage and time between treatment and examination, and in
part to differences in the response of the tissues being studied.

Hevesy (1945) has noted that cells are highly radiosensitive if the
synthesis of nucleic acids and associated materials is proceeding rapidly,
but are more resistant if synthetic activity is low. Differences in sensi-
tivity between actively dividing and differentiated tissues are explained
on the assumption that cells in actively growing tissues do not have time
to recover from the deleterious effects of radiation before initiating divi-
sion, whereas there is ample time for recovery in differentiated cells.
The high content of desoxyribonucleic acid in tumor cells may be a
factor in their radiosensitivity (Sparrow, 1944).

Another problem that has been attacked by chemical and cytochemical
methods concerns the nature of the changes induced by ionizing radia-
tions that lead to the adhesion and deformation of chromosomes (the
"primary" or "physiological" effect). Adhesion of chromosomes sug-
gests that changes have occurred in the viscosity of their constituent
materials. It has frequently been assumed that such changes in viscosity
are due to depolymerization of desoxyribonucleic acid (see, for example,
Darlington, 1942). There is little question that X rays can depolymerize
salts of desoxyribonucleic acid in vitro, as has been shown, for example,
by Sparrow and Rosenfeld (1946) ; Taylor, Greenstein, and Hollaender
(1948); G. C. Butler (1949); Scholes, Stein, and Weiss (1949); Limperos
and Mosher (1950) ; and Smith and Butler (1951). The data of Sparrow
and Rosenfeld for sodium thymonucleate are given in the last column of
Table 9-16.

Table 9-16. Relation of X-ray Dosage to Relative Viscosities of Solutions of

Thymonucleohistone and Sodium Thymonucleate

(Sparrow and Rosenfeld, 1946)

Relative

viscosity

Dosage, r

Nucleohistone

Sodium
thymonucleate

0

3.47

3.97

7,500

3.27

3.50

15,000

3.10

3.13

30 , 000

2.72

2.26

45,000

2.54

1.99

60 , 000

2.21

1.61

90,000

1.79

1.25

120,000

1.74

1.15

CHROMOSOME ABERRATIONS IN ANIMALS G95

High doses of radiation were used in all such experiments ; in the studies
just cited the minimum dose required to abolish structural viscosity
completely in a 0.2 per cent solution was 22,400 r. It may well be ques-
tioned, therefore, whether the comparatively low doses required to effect
adhesion and pycnosis of chromosomes in living cells — in some cases as
little as 25 r — operate in a similar manner. It may also be asked whether
depolymerization, which leads to a decrease in viscosity of solutions of
nucleic acid in vitro, would bring about an increase in stickiness of the
materials of the chromosomes.

In an effort to answer these questions, cytochemical studies have been
made of the effect of the exposure to ionizing radiations on methyl green
stainability of chromosomes. Purified methyl green, when used under
suitable experimental conditions, stains polymerized but not depolymer-
ized desoxyribonucleic acid (Kurnick, 1947, 1950; Pollister and Leuchten-
berger, 1949). Methyl green stainability of meiotic cells of Trillium
erectum was not impaired, as determined by spectrophotometric methods,
when the living buds were exposed to doses as high as 20,000 r, although
such doses reduce the viscosity of solutions of desoxyribonucleic acid
(Moses, DuBow, and Sparrow, 1951). It is apparent that depolymeriza-
tion of desoxyribonucleic acid (which can be produced by treatment of
fixed cells with hot water or with the enzyme desoxyribonuclease) is not
induced in the living cells of this plant by exposure to X rays. Similar
results were obtained by the author in studies of plant and animal cells
in which X-ray treatment had produced deformed and adherent meta-
phase and anaphase chromosomes. No reduction of methyl green stain-
ability of these chromosomes was effected by doses as high as 16,000 r.
These results agree with those of Himes (1950), who found that the
"stickiness" of chromosomes caused by the "sticky" gene in maize and
by chemical treatment of root tips of onion is not due to depolymerization
of the desoxyribonucleic acid.

In the light of these experimentally derived data it appears that the
attempts to interpret the effects of X rays on chromosomes in terms of
depolymerization of desoxyribonucleic acid are premature and inconclu-
sive. Attachment and detachment of desoxyribonucleic acid are not
simple coupling and uncoupling phenomena, since it has been shown that
the chromosome is a complex aggregate of both desoxyribose and ribose
nucleic acids, intimately associated with each other and with histones and
more acidic proteins (Kaufmann, McDonald, and Gay, 1951). Some
studies have also indicated that there is very little increase in the amount
of desoxyribonucleic acid between mid-interphase and metaphase (e.g.,
Swift, 1950; Lison and Pasteels, 1950 ; Pasteels and Lison, 1950), although
other experiments (e.g., those of Ogur et al, 1951, on Lilium) suggest that
the amount of desoxyribonucleic acid increases during mitosis as well as
during interphase. It has also been reported that changes occur during

696 RADIATION BIOLOGY

mitotic prophases in the amount and type of chromosomal ribonucleic
acid (Kaufmann, McDonald, and Gay, 1948).

Since in the living cell nucleic acids exist in association with proteins,
the possible effects of ionizing radiations on proteins and nucleoproteins
must also be assessed. Changes occur in the viscosity of solutions of
gelatin, egg albumin, and serum albumin exposed to X rays or radium
(see, for example, the review of Arnow, 1936). The action of X rays in
lowering the viscosity of a solution of nucleohistone is shown in Table
9-16. When nucleoproteins prepared from chicken erythrocytes and
carp spermatozoa in the form of stiff gels were irradiated, the gels gradu-
ally liquefied (Errera, 1946). If the cells were first subjected to intense
irradiation (50,000 r), and the nucleoproteins were then extracted, the
latter formed liquid solutions rather than gels. Von Euler and Hahn
(1946) irradiated nuclei isolated from calf thymus with 65,000 r but did
not detect any change in the nucleoproteins. Cytological examination
of cells of the testis of the salamander, Triturus, that had been irradiated
with heavy doses of X rays (from 5,000 to 50,000 r) revealed chromosomal
material that had spread out along the spindle fibers (Rugh, 1950). It
was suggested that denaturation of protein might have been responsible
for the lowered viscosity leading to this dislocation.

In the light of these various lines of experimental evidence it is apparent
that a satisfactory understanding of the action of ionizing radiations on
the materials of the chromosome must await the accumulation of informa-
tion about the types and patterns of association of nucleic acids, proteins,
and related materials during the various phases of the mitotic cycle.

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CHROMOSOME ABERRATIONS IN ANIMALS 697

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CHROMOSOME ABERRATIONS IN ANIMALS ()99

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700 RADIATION BIOLOGY

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702 RADIATION BIOLOGY

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CHROMOSOME ABERRATIONS IN ANIMALS 703

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704 RADIATION BIOLOGY

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CHROMOSOME ABERRATIONS IN ANIMALS 705

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700 RADIATION BIOLOGY

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Manuscript received by the editor May 1, 1951

CHAPTER 10

Radiation-induced Chromosome Aberrations in Tradescantia1

Norman H. Giles, Jr.

Osbom Botanical Laboratory , Yale University, and
Biology Division, Oak Ridge National Laboratory

Introduction. Methods; aberration types and their relationships: General experimental
techniques — Types of aberrations — Ratio of symmetrical and asymmetrical exchanges —
Distribution of breaks in chromosomes — Distribution of aberration types among nuclei —
Spontaneous chromosome aberrations — Time of chromosome division. Variations in
radiosensitivity during the nuclear cycle. Quantitative radiation results: The relation of
aberration yield to dose — The relation of aberration yield to intensity — The space factor in
reunion — Proportion of breaks that undergo restitution — Relative efficiencies of various
radiations — Aberration production by absorbed radiosotopes — Aberration production
by slow neutrons. Recapitulation. Effects of modifying factors: Centr if ligation;
Sonic vibration — Colchicine — Ultraviolet — Infrared — Temperature — Oxygen. Conclu-
sions. References.

INTRODUCTION

Studies on the effects of ionizing radiation on the chromosomes of
species of the spiderwort genus Tradescantia have all appeared since the
publication of the volumes edited by Duggar (1936), "Biological Effects
of Radiation," and, as a consequence, the present review does not consti-
tute in any way a revision and extension of earlier observations presented
in these volumes. There have appeared, however, within the past few
years a number of more or less complete treatments of the early radiation
studies on Tradescantia. In particular, the discussions by Lea (1946)
and by Catcheside (1945, 1948) have been quite extensive. Under the
circumstances, it does not seem desirable to review in as great detail as
would otherwise be necessary the material already covered in these
publications. In order to make the present discussion reasonably com-
plete, however, an attempt will be made to present the essential details
of earlier observations, together with references to selected original
papers and review articles on these subjects. In addition, a particular
effort will be made to provide a more detailed discussion of recent research
in this field, with emphasis on those results which appear to necessitate a

1 Work at Oak Ridge performed under Contract No. W-7405-eng-26 for the Atomic
Energy Commission.

713

714 RADIATION BIOLOGY

modification of earlier concepts or which have contributed to a better
understanding of the basic mechanisms by which ionizing radiations
produce chromosome aberrations.

A complete evaluation of the significance of current research in this
field, especially as it relates to earlier results, is not an easy task. As a
consequence, many of the conclusions drawn at the present time may
require considerable revision in the near future. This is particularly
true with respect to the recent discovery of the significance of oxygen in
increasing the frequency of aberrations. It seems clear that an under-
standing of this effect is fundamental to any interpretation of the physico-
chemical mechanism of aberration production by ionizing radiations.
At the present time, however, it is obvious that many questions concern-
ing the nature of the oxygen effect remain to be answered.

The first utilization of Tradescantia for quantitative X-ray experiments
is that of Sax (1938). Previous workers (Riley, 1936; Husted, 1936) had,
however, demonstrated the suitability of this material, especially the
developing microspores, for the cytological analysis of radiation effects.
The initial quantitative X-ray observations of Sax were soon extended by
him and by his students at Harvard and elsewhere utilizing X rays as
well as other types of radiation. Sax presented the early X-ray results
in terms of a comprehensive theory of the effects of radiation on the
chromosomes of Tradescantia in 1940. The Tradescantia investigations
were initiated shortly thereafter in England by Catcheside, Lea, and
others. These investigators, on the basis of the earlier studies together
with their own observations, developed a detailed, quantitative theory of
chromosome aberration production in Tradescantia by ionizing radiations.
Certain aspects of this theory are presented in a review by Catcheside
(1945), and a comprehensive account is given in the volume by Lea (1946).
A later review, including additional literature appearing since 1945, has
been written by Catcheside (1948).

METHODS; ABERRATION TYPES AND THEIR RELATIONSHIPS

GENERAL EXPERIMENTAL TECHNIQUES

Most of the observations on radiation effects in Tradescantia have been
carried out on diploid species in which the haploid chromosome number is
6. Several different species have been studied including T. gigantea,
T. bracteata, T. canaliculata, and especially T. paludosa. Although cer-
tain differences in quantitative results have been noted when supposedly
equivalent irradiation treatments have been applied in different labora-
tories, such differences appear not to result from marked differences in
species sensitivity (Sax and Swanson, 1941), but rather from differences
in exposure conditions and dosage measurements. It is possible, how-

chromosome aberrations in Tradescantia 715

ever, that a certain degree of genetically controlled difference in sensi-
tivity among species or within a given species may exist.

The stage in the life cycle of Tradescantia which has proved most
favorable for observations of radiation effects on chromosomes is the
first postmeiotic mitosis in the developing microspores. Following
meiosis, the haploid microspores (having six chromosomes) go through a
regular mitotic cycle, and the chromosomes can be observed at metaphase
or anaphase, utilizing simple acetocarmine or other smear techniques.
The various mitotic stages are usually fairly well synchronized in the six
anthers of one bud, and a smear preparation of an appropriately selected
single bud will usually contain from 100-400 cells in a stage of mitosis
suitable for chromosome observations. Since the different buds on a
single inflorescence contain microspores in successive stages of develop-
ment, it is possible with one radiation exposure to treat cells in different
stages of mitosis. Observations of nuclei at metaphase or anaphase can
then be made at intervals after irradiation and the types and frequencies
of various chromosome aberrations determined. Since the duration of
the various stages in microspores has been determined (Sax and Edmonds,
1933) , it is possible to make certain correlations of aberration types and
frequencies with particular stages in the mitotic cycle, especially with the
resting stage and prophase.

When inflorescences are exposed to penetrating radiation and the cells
in metaphase and anaphase examined immediately afterward, the chromo-
somes can be seen to exhibit a type of stickiness during which certain
chromosome parts, especially ends, tend to adhere. This adherence may
lead occasionally to bridging and fragmentation at anaphase. Such a
condition has been attributed to an effect of the radiation on the nucleic
acid forming the matrical material of the chromosome (Darlington, 1942).
This type of transient aberrant behavior is soon replaced, as seen in cells
examined a few hours after exposure, by the typical permanent aberration
types resulting from chromosome breakage and the reunion of broken
ends with which the subsequent discussion will be concerned.

TYPES OF ABERRATIONS

Observations at increasing intervals of time following irradiation have
shown the existence of two major categories of aberration types differing
in whether they are derived from chromosomes which are effectively
single or double at the time of irradiation.

Chromatid Aberrations. The first category consists of types produced
in chromosomes which are effectively double at the time of irradiation.
Chromatid aberrations appear first, within a few hours after irradiation,
reach a peak 12-20 hours later, and are the only types present up to about
26 hours (under spring and summer growing conditions; see Sax, 1941).
Following a relatively brief transition period, in which both chromatid

716

RADIATION BIOLOGY

and chromosome types may occur in the same cell — or rarely, even in the
same chromosome (Sax and Mather, 1939)— chromosome types replace
the chromatid types and are present exclusively from about 31 hours on.
The frequency of chromosome aberrations rises rapidly to a maximum
shortly after this type appears, and this frequency is maintained for
several days after irradiation. The typical periods for observation of

CHROMOSOME ABERRATIONS

TYPE

IS

o

in hi

cc _J

uj i±J

I- Q

Z

INTERPHASE

PRO-
METAPHASE

ANAPHASE

CHROMATID ABERRATIONS

TYPE

2 UJ

O a:

CC CD

I

o
1-

CD

PROPHASE

PRE-
METAPHASE

ANAPHASE

UJ

o _

28

o

cr

cc o

0^

0

z

I-
z

o

vv

Fig. 10-1. Diagrammatic representation of the origin and types of aberrations, both
chromosome and chromatid, induced in Tradescantia microspores by ionizing radia-
tions. Only the types of major importance which can be recognized at metaphase
and anaphase in cytological analyses are shown. Original break positions at inter-
phase or prophase are indicated by short diagonal lines. {Modified from Sax, 1940.
Reproduced by permission of the author and the editor of Genetics.)

chromatid and chromosome aberration frequencies are 20-24 hours and
4-5 days following irradiation, respectively. The time and duration of
the transition period appears to depend rather markedly on environ-
mental factors.

The various kinds of chromatid and chromosome aberration types pro-
duced in Tradescantia microspores have been diagramed and discussed
in considerable detail by Sax (1940), Lea (1946), and especially by
Catcheside, Lea, and Thoday (1946a) and Catcheside (1948). The
present discussion will consider in detail only certain selected types which
occur most frequently and which have been of major importance in the

chromosome aberrations in Tradcscantia 111

quantitative data on radiation effects. The mode of origin of these
types is shown diagrammatically in Fig. 10-1, and photomicrographs of
the various types are presented in Plate I.

There are three major chromatid aberration types: (1) chromatid
breaks, in which one of the two chromatids in a metaphase chromosome
has a transverse fracture, giving rise to a free acentric fragment at
anaphase (Plate I-E, F) ; (2) isochromatid breaks (chromatid dicentrics),
in which the two sister chromatids of a single chromosome are broken at
the same locus, the broken ends typically undergoing lateral union to
produce a dicentric and a U-shaped fragment (Plate I-B). At the suc-
ceeding anaphase a bridge and a free acentric fragment result (Plate
I-C). The interpretation of the manner in which ionizing radiations
produce isochromatid breaks has been a controversial one. For example,
Darlington and La Cour (1945) have maintained that this aberration
type arises as a result of chromosome breakage before effective division,
apparent sister union resulting from a reproductive error in the chromo-
some thread and not from true reunion. Catcheside (1948) has sum-
marized the evidence against this view and in favor of the alternative one,
that isochromatid breaks, in general, arise as a result of the simultaneous
breakage at the same level of the two chromatids of a single chromosome.
(3) Chromatid exchanges are derived from two chromatid breaks, either in
two separate chromosomes, yielding interchanges, or in the same chromo-
some, yielding intrachanges. The commonest category is the interchange
in which the union of broken ends gives rise to symmetrical (reciprocal
translocation, Plate I-D) or to asymmetrical (dicentric and acentric frag-
ment, Plate I-E) types. Additional exchange types, which are in general
much less frequent, involve recombination between two isochromatid
breaks or between one isochromatid and one chromatid break, and have
been described and diagramed by Catcheside, Lea, and Thoday (1946a).

Chromosome Aberrations. This second major category of aberration
types is made up of those arising in chromosomes which are effectively
single at the time of exposure. There are also three major chromosome
aberration types: (1) chromosome breaks (terminal deletions), in which
the unsplit chromosome suffers a transverse fracture giving rise at meta-
phase to two shortened chromatids and two acentric fragments (Plate
I-G) ; (2) interstitial deletions (minute, isodiametric fragments), involving
two breaks close together in one arm of an unsplit chromosome, the broken
ends uniting to produce a small acentric ring which appears following
division as a pair of dotlike fragments at metaphase (Plate I-G, N). This
aberration is actually a type of intra-arm exchange (intrachange) but is
listed separately because of the frequency with which it occurs. (3)
Chromosome exchanges, which arise from two chromosome breaks either in
two different chromosomes, yielding interchanges, or in different arms of
the same chromosome, yielding large intrachanges. In general, only the

718

RADIATION BIOLOGY

WC

J 1

Z

B

''V<

D

E

G

H

«*5*

^>

K

\

V

M

7r

*

N
Plate I

o

chromosome aberrations in Tradescantia 719

Description of Plate I

Photomicrographs of Tradescantia microspore chromosomes showing the major
types of aberrations induced by ionizing radiations (from Sax, 1940). Figures A-F
are chromatid aberration types; Figs. G-0 are chromosome aberration types. (Repro-
duced by permission of the author and the editor of Genetics.)

Fig. A. Untreated. Six normal chromosomes at metaphase.

Fig. B. An isochromatid break at metaphase. Acentric fragment (U-shaped) at left.

Fig. C. Similar isochromatid aberration at anaphase. Partially straightened frag-
ment to right.

Fig. D. Symmetrical chromatid interchange at metaphase.

Fig. E. Asymmetrical chromatid interchange (at top) and a chromatid break (center).

Fig. F. Chromatid break at anaphase, shortened arm and acentric fragment visible
at upper center. Also probable chromatid-isochromatid intrachange giving duplica-
tion-deficiency chromatids (to left).

Fig. G. A chromosome break (upper center) and a chromosome interstitial deletion
(to right).

Fig. H. An asymmetrical chromosome interchange (dicentric); paired acentric frag-
ments at lower right.

Fig. I. An asymmetrical chromosome interchange (dicentric) at upper right, and an
asymmetrical chromosome intrachange (centric ring) at lower center. Two pairs of
acentric fragments at top, center and left.

Fig. J. A symmetrical chromosome interchange.

Fig. K-N. Types of separation at anaphase of asymmetrical chromosome inter-
changes (dicentrics) . An interstitial deletion is also present in Fig. N.

Fig. O. Anaphase separation of an asymmetrical chromosome intrachange as a
continuous ring. Somatic nondisjunction ha"s also occurred.

720 RADIATION BIOLOGY

asymmetrical types in these two categories — dicentrics (Plate I-H, I, K,
L, M, N) and centric rings (Plate I-I, 0), respectively — can be regularly
scored, although symmetrical interchanges can be detected occasionally
when they are very unequal (Plate I-J).

RATIO OF SYMMETRICAL AND ASYMMETRICAL EXCHANGES

Symmetrical and asymmetrical chromatid interchanges appear to be
about equally frequent according to Catcheside, Lea, and Thoday (1946a),
although the data of Sax (1940) suggest that asymmetrical interchanges
are more frequent. These latter results are probably biased somewhat,
since cells at anaphase as well as at metaphase were scored, and sym-
metrical exchanges cannot be detected at anaphase. The observations of
Catcheside et al. (1946a) also indicate that symmetrical and asymmetrical
chromatid intrachanges are equally frequent and these authors state that
such results argue against the existence of any polarization in chromo-
somes which would prevent the random joining of breakage ends.

DISTRIBUTION OF BREAKS IN CHROMOSOMES

It is not possible to determine directly in Tradescantia the distribution
of initial breakage, since the processes of restitution and reunion, to be
considered later, intervene. There is, in fact, evidence that more aber-
rations tend to arise from breaks produced in the proximal rather than in
the distal regions of a chromosome (Sax, 1940). Although such evidence
may indicate a nonrandom distribution of initial breaks, it appears more
likely that it indicates a nonrandom reunion of broken ends, resulting
from the operation of secondary factors influencing reunion and restitu-
tion. The more proximal distribution of aberrations has been attributed
by Sax (1940) to stresses imposed in the region of the centromere by the
various coiling mechanisms.

DISTRIBUTION OF ABERRATION TYPES AMONG NUCLEI

The distribution of various chromatid and chromosome aberration
types among nuclei following X irradiation and neutron irradiation has
been studied by Catcheside, Lea, and Thoday (1946a) and Rick (1940).
The distributions observed are in accordance with the Poisson formula,
and this evidence, especially that for chromatid breaks, is taken as com-
patible with the view that single breaks in a given chromosome are pro-
duced by the action of single ionizing particles and are unaffected by the
presence or absence of other breaks in the cell.

SPONTANEOUS CHROMOSOME ABERRATIONS

Of interest in connection with radiation experiments are observations
on the types and frequencies of spontaneous aberration types in micro-
spores. Such data are in fact necessary as controls for comparison with

chromosome aberrations in Tradescantia 721

experimental material. Studies of spontaneous chromosome aberrations
have demonstrated that the same aberration types occur in unirradiated
material, but that the frequencies are usually very low (Giles, 1940a;
Sax and Luippold, 1952). Even this low rate is much too high to be
accounted for in terms of natural radiation. Further, there is evidence
that the spontaneous rate may be considerably higher in material of
hybrid origin (Giles, 1940a, 1941; Darlington and Upcott, 1941).

TIME OF CHROMOSOME DIVISION

Observations on the kinds and sequence of aberration types have been
used to draw certain conclusions about the time and degree of chromo-
some duplication during mitosis (Riley, 1936; Mather, 1937; Sax, 1940,
1941 ; Catcheside, 1948). The transition from chromosome to chromatid
aberrations, which corresponds in general to the late resting stage, has
been taken to indicate the time in the nuclear cycle at which duplication
occurs (Mather, 1937). One of the difficulties with this interpretation,
however, is the observation that chromosome breaks may occur in the
same cell with chromatid breaks. Some investigators (Darlington and
La Cour, 1945) have, in fact, maintained that chromosomes are broken
by radiation only when they are undivided and that reunion then takes
place after division, giving rise to either chromosome or chromatid break
types. Catcheside (1948) has summarized the evidence favoring the
view that chromatid aberrations actually arise as a result of breaks in
divided chromosomes, and has concluded that, while a small proportion of
chromatid breaks may be derived from chromosome breaks, most are not.
It thus seems likely that the average time of chromosome splitting should
be about halfway between the peaks of chromatid and chromosome break-
age, and thus does correspond in general to the transition from chromo-
some to chromatid types.

It can still be argued, however, that actual duplication has occurred
earlier, but that the chromosome or chromatid is the unit of reunion.
There is, in fact, some evidence from radiation studies (Swanson, 1947)
for a quadripartite condition of the chromosomes at prophase in pollen
tube nuclei. This condition appears to be the exception rather than the
rule, however, since even in pollen tubes, chromatid rather than half-
chromatid break types predominate in nuclei where the chromosomes can
be seen microscopically to be divided at the time of treatment (Swanson,
1943).

VARIATIONS IN RADIOSENSITIVITY DURING THE NUCLEAR CYCLE

The original studies of Sax (1938, 1940) demonstrated that more aber-
rations were produced for the same X-ray dose in prophase as compared
with resting stage chromosomes of Tradescantia microspores. It was

722 RADIATION BIOLOGY

later shown (Sax and Swanson, 1941), that the sensitivity apparently was
greatest shortly before mid-prophase, somewhat less in early prophase,
and only about one-third the prophase maximum during the microspore
resting stage. Comparative studies with diploid microspores from tetra-
ploid Tradescantia species indicated a similar sensitivity cycle, but also
demonstrated that the diploid spores were only one-half to one-third as
sensitive as the haploid, in terms of aberrations per chromosome. Fur-
ther evidence from the tetraploids indicated that meiosis was much more
sensitive than mitosis. Chromosomes in root tip mitoses in the diploid
were found to be less sensitive than those in the haploid microspores.
Bishop (1950) has studied the radiosensitivity of chromosomes in the
postmicrospore mitosis (in the generative nucleus of the pollen grain)
which is initiated after the microspore division and completed in the
pollen tube, at which time cytological analyses at metaphase can be
made. He finds evidence for two sensitivity peaks, one at 3 days and
one at 5 days before the dehiscence of the anthers. These peaks are con-
sidered to correspond, respectively, to the time of chromosome doubling
during prophase of the pollen tube mitosis (cf. Roller, 1946), and to
metaphase-anaphase of the microspore mitosis. The range of sensitivity,
from the low point at the postmitotic resting stage, is estimated to be
approximately 2.4 times as great for the prophase maximum and ten
times as great for the metaphase-anaphase maximum.

The extensive studies of Sparrow (1951) on the radiosensitivity of
Trillium chromosomes are in general agreement with the Tradescantia
data. The more detailed data on Trillium, particularly at meiosis, indi-
cate that late prophase and metaphase of the first meiotic division are
over fifty times as sensitive as the least sensitive stage, early interphase in
the microspore. Complete data are not yet available for the entire
microspore cycle.

The factors responsible for such marked changes in sensitivity during
the nuclear cycle are not yet completely elucidated (Sparrow, 1951). It
is clear that any change in sensitivity (i.e., any change in the frequency
of aberrations produced by the same dose) may be attributed either
(1) to an actual difference in the number of initial breaks, or (2) to a
change in the proportion of breaks which undergo reunion and/or restitu-
tion, or (3) to both these factors. There is evidence in Trillium (Sparrow
and Maldawer, 1950), from a comparison of the ratios of fragments to
dicentrics and rings induced at metaphase of meiosis and at microspore
interphase, that a greater amount of reunion occurs at interphase, the
stage of lowest sensitivity. However, the increase in reunion is not of
sufficient magnitude to account for the observed decrease in fragmenta-
tion. It thus appears that both factors mentioned may be involved in
changes in sensitivity.

Many possible reasons for variations in the incidence of primary break-

chromosome aberrations in Tradescantia 723

age or of restitution and reunion during the nuclear cycle have been sug-
gested (Sparrow, 1951). Differences in primary breakage may reflect
variations in the chemical composition of the chromosomes during
mitosis and meiosis, for example, in their desoxyribonucleic acid (DNA)
content, as suggested by Darlington and La Cour (1945). [However,
quantitative measurements by Ris (1947) and by Sparrow, Moses, and
Steele (1950) indicate that there is no significant change in the amounts
of either desoxyribonucleic acid or pentose nucleic acid during meiosis
and mitosis.] It is also possible that changes in the cellular environ-
ment (e.g., changes in oxygen tension during mitosis and meiosis) may
be responsible for modifying the relative efficiency of ionizing radiations
in breaking chromosomes. Among the factors that have been suggested
as possibly influencing reunion and restitution are the type of coiling
present in the chromosomes and the degree of chromosome duplication
(plus the relative proximity of the duplicated sister strands) at the time
of irradiation (Sax and Swanson, 1941; Bishop, 1950).

QUANTITATIVE RADIATION RESULTS

THE RELATION OF ABERRATION YIELD TO DOSE

The initial development of a quantitative theory of chromosome aber-
ration production in Tradescantia microspores by radiation resulted
primarily from studies of the relation between the yield of various aberra-
tions with dosage and intensity of X rays and neutrons. The early
experiments of Sax (1938; 1940) with X rays demonstrated two types of
relationship: (1) a linear relation with dose for certain aberration types,
e.g., for isochromatid breaks, and (2) a nonlinear (geometric-exponential)
relation (the exponent of the dose being greater than one) for other aberra-
tions, such as chromatid and chromosome exchanges. In the first experi-
ments of Sax, in which the radiation dose was administered at a constant
intensity, the exponent of the dosage curve for exchanges was approxi-
mately 1.5. In later experiments, when the time of irradiation was kept
constant and the intensity varied, the exponent of the dosage curve for
exchanges was approximately 2 (Fig. 10-2).

These results were interpreted as indicating that with X rays certain
aberration types (one-hit) are produced by single events, whereas, other
aberrations (two-hit types) are produced by two separate events (which
must be related in time and space, as will be discussed later). The
nature of the event producing the break, whether related to a single
ionization or to the passage of a single ionizing particle, could not be
determined from the initial X-ray experiments alone.

Subsequently, experiments with fast neutrons (Giles, 1940b) showed
that with this radiation the frequencies of all aberration types studied,

724

RADIATION BIOLOGY

including chromatid and chromosome exchanges, were linearly related to
dose (Fig. 10-3). Further, when equal doses of X rays and neutrons were
compared, neutrons were found to be much more efficient than X rays in
producing aberrations. These results were interpreted as indicating that
with fast neutrons all aberrations are produced by single events, and that
the passage of an ionizing particle, rather than the occurrence of a single
ionization, constitutes the event, or hit.

30 -i

(/)

_i
_i

LU
O

Q

O

tr

LU
Q.

en

O

h-
<

20

LU io-

50 100

X-RAY DOSE, r

O ISOCHROMATID BREAKS
• CHROMATID EXCHANGES

Fig. 10-2. Relation between X-ray dosage and frequency of isochromatid breaks
(one-hit) and chromatid exchanges (two-hit). Time of exposure constant. (Data
of Sax, 1940. Dosage values multiplied by correction factor, cf. Catcheside, Lea, and
Thoday, 1946.)

These observations on X-ray and fast-neutron effects on Tradescantia
chromosomes have been subsequently confirmed and extended in numer-
ous independent experiments. The following general conclusions relating
the frequency of various aberration types to dose can be made. Simple
(one-hit) break types, i.e., chromosome and chromatid breaks and iso-
chromatid breaks, increase in approximately linear proportion to dose for
all types of radiations tested — X and y rays (Sax, 1938; 1940; 1941;
Thoday, 1942; Newcombe, 1942; Rick, 1940; Catcheside et al, 1946b),
neutrons (Giles, 1940b; 1943; Thoday, 1942), and a particles (Kotval and

chromosome aberrations in Tradescantia

725

Gray, 1947). Break types involving exchanges, i.e., chromatid inter-
changes and chromosome dicentrics and rings, increase more rapidly
than the first power of the dose when X rays or 7 rays are used. The
actual exponent obtained in a given experiment depends on the intensity
of the radiation. With high intensities (e.g., ca. 150 r/minute) aberra-
tion frequency is approximately proportional to the square of the dose
(Sax, 1941; Thoday, 1942; Sax and Brumfield, 1943). At intermediate
and low intensities (e.g., ca. 20 r/minute and 3 r/minute), the exponent

400 r ( X RAY )
40 n (NEUTRONS)

A FAST NEUTRONS (OAK RIDGE REACTOR)

• X RAYS (190 kv- MEMORIAL HOSPITAL)

Fig. 10-3. Relation between frequencies of chromosome interchange (dicentric and
centric rings) and dosages of fast neutrons and X rays. 1 n unit = ca. 2.5 r. (Data
of Giles, unpublished.)

becomes progressively less than 2 (Fig. 10-4) (Sax, 1941). A dosage-
squared relationship is also obtained if the time of irradiation is kept
constant and the intensity varied (Sax, 1940, 1941), provided the total
irradiation time is relatively short (Sax, 1950b). It is concluded from
these results that, fundamentally, exchange aberration frequencies
increase as the square of the dose with X rays, the two breaks involved
being produced by separate X-ray hits. Modifications of the dosage-
squared relationship are attributed indirectly to an effect of intensity,
such that at low intensities relatively greater restitution, as compared
with reunion, occurs and hence proportionally more exchanges arise as a
result of single hits.

726

RADIATION BIOLOGY

Although the majority of the X-ray-induced interchanges appear to
result from two separate hits, a certain fraction has been shown to
originate as a result of single hits. This proportion has been investigated
for chromatid exchanges, by Catcheside, Lea, and Thoday (1946b) in
experiments in which two similar dosage curves were obtained at con-
stant, but widely different, exposure times. The results indicate that
one-hit exchanges are of relatively minor consequence at high intensities,
but make a substantial contribution to the total aberration yield at low

lOO-i

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o

Q.

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o

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<.

K

cc

Ul

m

<

100

300
DOSE OF XRAYS, r

500

J 160 r/min

INTENSITY A 20 r/min

O 2.7 r/min

Fig. 10-4. Effect of dosages of X rays at different intensities upon the yield of chromo-
some exchanges. (Data from Sax, 1941.)

intensities. Similar conclusions have been arrived at by Sax for chromo-
some exchanges (1950b).

Rick (1940) has shown that the yield of interstitial deletions increases
as the 1.5 power of the dose when irradiations are performed at constant
time and at relatively high intensities, and has concluded that these
types represent a mixture of one- and two-hit aberrations.

With fast neutrons (Giles, 1940; 1943; Giles and Conger, 1950; Thoday,
1942) and a particles (Kotval and Gray, 1947), all types of exchange
aberrations increase linearly with dose. As indicated previously, these
results are compatible with the view that such two-break aberrations are
produced by the passage of a single ionizing particle. The special case of
slow-neutron effects will be considered later.

chromosome aberrations in Trade scantia

727

THE RELATION OF ABERRATION YIELD TO INTENSITY

Some of the results just discussed under dosage relationships indicated
the existence of an intensity effect on the yield of certain aberration
types in Tradescantia. This effect has been investigated in detail in
X-ray experiments by Sax (1939, 1940, 1950a, b, 1952), Faberge (1940),
Marinelli, Nebel, Giles, and Charles (1942), Catcheside, Lea, and Thoday
(1946b), and Lane (1951).

The most direct way in which an intensity effect can be demonstrated is
in experiments in which the same X-ray dose is administered at several

60-,

DURATION OF EXPOSURE, minutes

• X RAY, 300 r (MARINELLI & Ol., 1942)

A X RAYS, 320 r( SAX, 1939)

O FAST NEUTRONS, 26 n (GILES, 1943)

Fig. 10-5. Effect of intensity of radiation dose on yield of chromosome interchanges
induced by X rays and neutrons.

different intensities. When this is done, it is found that there is a marked
decrease in the frequencies of exchange-type aberrations, both chromo-
some and chromatid; whereas, the yields of chromatid and isochromatid
breaks are not affected (Figs. 10-5, 10-6). The general conclusion
drawn from such experiments is that the two independently produced
breaks necessary for the production of an exchange aberration must occur
within a certain time in order for reunion, and thus aberration formation,
to result. This is the case, since restitution is constantly taking place;
and if the second break is not produced before restitution of the first
break occurs, no exchange can result. Consequently, when a given
X-ray dose is administered at a high intensity, most breaks are simul-
taneously present in a given nucleus and numerous reunions can occur;

728

RADIATION BIOLOGY

when the same dose is extended over a long period of time, restitution
intervenes to reduce the number of breaks which can participate in
reunion. On this view, absence of an intensity effect with chromatid
and isochromatid breaks is expected, since these are one-hit aberration

types.

The data from neutron experiments (Giles, 1943) indicate that, con-
trary to the X-ray results, there is no intensity effect with either exchanges

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40-

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-AO

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4

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16

DURATION OF EXPOSURE, minutes

• CHROMATID BREAKS (XRAYS, 150/)- CATCHESIDE et a/., 1946
A ISOCHROMATID BREAKS (X RAYS,'l50 r)- CATCHESIDE eta/., 1946
A ISOCHROMATID BREAKS (X RAYS, 130 r)- GILES, 1943
O ISOCHROMATID BREAKS (FAST NEUTRONS, 10 nj-GlLES, 1943

Fig. 10-6. Effect of intensity of radiation dose on yield of chromatid and isochromatid
breaks induced by X rays and neutrons.

or simple break types (Figs. 10-5, 10-6). The absence of an intensity
effect with exchanges affords strong support for the previously outlined
mechanism of aberration production, since these aberrations have been
shown by dosage data to be one-hit types, both breaks being produced by
a single proton path; consequently, an intensity effect is not anticipated.
Lea and Catcheside (1942), Lea (1946), and Catcheside, Lea, and
Thoday (1946b) have used the experimental data on the intensity effect
for exchanges to calculate the average restitution time (t), i.e., the
average time elapsing between breakage and restitution. Using the
original data of Sax (1939, 1940), the value of r is approximately 4

chromosome aberrations in Tradescantia 729

minutes for both chromosome and chromatid breaks. In later experi-
ments (Catcheside et al., 1946b), when irradiation was prolonged beyond
about 30 minutes, however, the experimental results were not in very
good agreement with the theoretical predictions, since there was an
unexpected persistence of interchanges even in experiments in which the
irradiation extended over several hours. These investigators studied this
situation in an attempt to determine whether the discrepancy could be
accounted for on the basis that some exchanges are one-hit rather than
two-hit aberrations. Although a certain proportion of exchanges were
found to be one-hit types, this value was not sufficient to account for the
observed discrepancy. Consequently, it was concluded that, in addition
to a short-term component (rl), there is also a long-term component (t2)
involved in restitution and that the value of t2 is of the order of hours,
not minutes. It is suggested that biologically this situation may result
from the circumstance that, if two broken ends of a given break do not
restitute within a few minutes, they may then become separated beyond
the spatial limits of restitution, and a considerable period may elapse
before they are accidentally brought together again.

In addition to experiments with varying intensity, other experiments
with fractionated doses have been performed to investigate the mechan-
ism of aberration production. In general, the results of these experiments
(Sax, 1939, 1940; Faberge, 1940) are in agreement with those of the
intensity experiments. When a given dose is divided into two or more
fractions, the yield of exchange aberrations decreases, but that of one-hit
types is not affected. By utilizing increasing time intervals between
doses, Sax (1939, 1940) concluded that some breaks may remain open
and capable of reunion for as long as 1 hour, but that most breaks undergo
restitution or reunion in a considerably shorter period.

The results of experiments by Lane (1951) on dose fractionation are
similar to those of Sax in showing a pronounced reduction in exchange
aberrations when intervals up to 4 hours between doses are employed.
However, when longer intervals of 6 or 8 hours were used, a recovery
effect was noted, the aberration yield returning, with an 8-hour interval
between fractions, to almost the level obtained with an equivalent dose
delivered without fractionation. This result, plus the observation that
the aberration yield from a given dose delivered in two equal fractions
often appeared to be less than twice the yield obtained when half that
dose was used, led to the conclusion that the so-called intensity effect
was actually to be interpreted in terms of a temporary inhibition of
chromosome breakage by radiation, such that an initial dose rendered
the chromosomes more resistant to breakage by a subsequent dose.
That this inhibition was temporary was shown by the subsequent
recovery effect with time intervals between doses longer than 4 hours.
These over-all results and interpretations are clearly incompatible with

730

RADIATION BIOLOGY

much of the theory of aberration production as outlined previously.
The experiments of Lane have, however, been repeated by Sax (1952) —
the only difference being that a higher radiation intensity was used — and
on the basis of a much more extensive analysis in terms of numbers of
cells scored, no evidence of an inhibition or recovery effect is found, even
with an interval between doses as long as 12 hours (Table 10-1). Thus

Table 10-1. Effect of X-ray Dose Fractionation on Yield of Chromosome

Interchanges, 180 r/Minute
(Data from Sax and Luippold, 1952)

Time interval

No., of

No. of

Interchanges

Dose, r

between doses, hr

cells

interchanges

per cell

360

0

2802

2055

0.732

180 + 180

4

2970

1534

0.516

180 + 180

8

1665

847

0.509

180 + 180

12

1000

487

0.487

180

0

2817

726

0.258 (X 2 = 0.516)

the rejection by Lane, on the basis of his experiments, of the theory of
relatively rapid reunion and restitution, appears not to be warranted.

THE SPACE FACTOR IN REUNION

Sax (1940) pointed out that the observed ratio of certain types of
chromosome inter- and intrachanges (i.e., dicentric-centric rings = 3.3 : 1)
was not as expected (10 : 1) on the basis of random reunion of broken ends,
intrachanges being favored. These data are interpreted as indicating
that space limitations influence the reunion process, and that most
breaks must be relatively close together in order for reunion to occur.
The further conclusion is drawn that the limitations to reunion imposed
by this factor of proximity must mean that most of the breaks induced by
radiations do not lead to the production of aberrations. Rather, the
two broken ends simply reunite in the original position (the process of
restitution) and no structural alteration is visible at succeeding meta-
phases or anaphases.

Lea (1946, 1947) has developed several quantitative methods for
estimating the average distance over which reunion occurs. One of
these depends on the use of biological and physical data on exchanges
induced by fast neutrons and X rays. It is possible to calculate the
number of ionizing particles, either protons or photoelectrons, which will
pass through a given Tradescantia microspore nucleus irradiated with
equivalent physical doses of neutrons of X rays. Since neutron-induced
exchanges are primarily one hit, the data on protons can be used to set
an upper limit to the distance from a given break at which a second break
normally taking part in an exchange must occur. Since X-ray-induced

chromosome aberrations in Tradescantia 731

exchanges are primarily two hit, the data on photoelectrons can be used
to set a lower limit to the distance from a given break at which the second
break must occur. Calculations based on physically equivalent neutron
and X-ray doses, e.g., at 50 r, indicate that 23 protons and 303 photo-
electrons pass through a nucleus of diameter 12 n and set these limits as
<1.3 and >0.9 jjl, respectively. Thus the average distance over which
reunion occurs is concluded to be approximately 1 n. Independent
estimates derived from the ratio of interstitial deletions to asymmetrical
exchanges and from the most frequent size of interstitial deletions are in
agreement with this calculation.

PROPORTION OF BREAKS THAT UNDERGO RESTITUTION

Evidence has been presented in the preceding section that reunion is
not at random, and that breaks separated by distances greater than
approximately 1 /z usually do not undergo reunion. From this evidence,
plus the results of the intensity experiments, it can be concluded that
the majority of breaks undergo restitution. Further general evidence
for restitution comes from data on the frequencies of different kinds of
breaks observed in individual cells. For example, as pointed out by
Catcheside (1948), it seems clear that chromatid exchanges and chromatid
breaks arise from a similar initial event, a chromatid break, reunion
occurring in one case and not in the other. If, in fact, chromatid
exchanges do arise in this manner from reunion of two chromatid breaks,
and if all chromatid breaks not participating in such reunions were pre-
served, it is apparent that the occurrence of chromatid exchanges should
affect the distribution of surviving chromatid breaks. However, in cells
with exchanges, there is no evidence of an excess of odd as compared with
even numbers of surviving chromatid breaks. Lea (1946) and Catche-
side, Lea, and Thoday (1946b) have developed several quantitative
methods for calculating from the number of observed aberrations of
various types the number of breaks primarily produced, and hence the
proportion of breaks which restitute. These include (1) a comparison of
the frequencies of incomplete reunion and restitution in various chromatid
aberration types; (2) a consideration of the departure at high doses from
the dose-squared relation for X-ray-induced exchanges; (3) comparisons
of expected and observed break frequencies with fast neutrons; (4) com-
parisons, based on the observed ratio of chromatid exchanges to chro-
matid-isochromatid exchanges, and of chromatid to isochromatid breaks,
of the relative proportions of primary chromatid and isochromatid breaks
which persist; and (5) a consideration of the relative frequencies of
chromatid and isochromatid breaks. All these calculations agree in
indicating that only a minority of the breaks primarily produced in
Tradescantia microspores are scored and that the majority restitute and
escape recognition. The estimates of the fraction (f) of unjoinable

732 ' RADIATION BIOLOGY

breaks are 0.09, 0.09, and 0.5 for X rays, neutrons, and a particles,
respectively.

The over-all evidence thus indicates that a substantial fraction of
originally induced breaks undergo restitution. Only failure of restitution
permits new reunion and the two processes of restitution and reunion are
thus competing ones in which both space and time factors are important.

RELATIVE EFFICIENCIES OF VARIOUS RADIATIONS

Comparisons of the relative efficiencies of various ionizing radiations
and of X rays of various wave lengths in producing breaks and aberra-
tions have been of considerable importance in the development of the
theory of aberration production in Tradescantia. As indicated pre-
viously, the greater efficiency of fast neutrons as compared with 160-kv
X rays provided part of the initial evidence that single ionizations were
insufficient to produce chromosome breaks. The conclusion that several
ionizations (or related chemical events, as will be discussed later) are
necessary for chromosome breakage arises from the following considera-
tion: Ionizations are much more densely spaced along a proton track
(derived from neutron radiation) than along an electron track (derived
from X radiation) . Hence, when a proton traverses a chromosome, many
ionizations will be produced within its volume ; if only one ionization were
required for breakage, neutrons should be less rather than more efficient
than X rays, since additional ionizations produced within the chromosome
would be wasted, contributing to the physical dose but not to the bio-
logical effect.

Utilizing the neutron-X-ray comparisons, Lea and Catcheside (1942)
attempted to determine the approximate number of ionizations required
to produce a break. Their conclusion was that seventeen ionizations, on
the average, are required for breakage to occur. Further, with the usual
X-irradiation procedures, in which relatively hard X rays are employed,
it appears that breakage arises primarily from the concentrated ionization
produced at the end (tail) of the electron path. The conclusion that the
tail of the electron path is responsible for break production with X rays
was tested experimentally by comparing the efficiencies of X rays of
different wave lengths, particularly in the soft X-ray range. For this
purpose, the effects on pollen tube chromosomes were examined and the
experimental results were found to agree well with the theoretical predic-
tions (Catcheside and Lea, 1943).

The calculations of Lea and Catcheside (1942) indicated that the prob-
ability of break production when a proton (from a fast neutron) traverses
a chromosome is somewhat less than one. On this basis a particles,
because of their much greater ionization density, should be actually less
efficient than fast neutrons. The studies of Kotval and Gray (1947),
however, indicate that a particles are actually more efficient than neu-

chromosome aberrations in Tradescantia 733

trons. It is possible to conclude from these results that a particles need
not always traverse a chromosome in order to produce a break, but that
breaks may also arise when the particle passes in the immediate vicinity
of the chromosome. Such a view implies an indirect action of the radia-
tion in producing chromosome breaks, a topic which will be considered in
detail later.

ABERRATION PRODUCTION BY ABSORBED RADIOISOTOPES

The abundant production by atomic reactors of radioactive isotopes of
various elements has stimulated interest in the use of these substances,
not only as tracer, but also as radiation, sources. Experiments have been
performed with two different isotopes, P32 and C14, both producing /3 rays,
to determine the effects of these substances in producing chromosome
aberrations in Tradescantia microspores (Giles, 1947; Giles and Bolomey,
1948). When cut stems of inflorescences were placed in solutions of
various activities, containing one or the other of the two isotopes, aberra-
tions similar to those produced by X rays were observed in the micro-
scopes. The types, frequencies, and temporal sequence of aberrations
were recorded over a period of several days following the initial treat-
ments. Quantitative measurements of /3 activity from disintegrations of
P32 molecules in individual half-anthers showed good correlations with
the relative frequencies of aberrations detected cytologically in sister
half-anthers (Giles and Bolomey, 1948). It has not been possible, how-
ever, in the Tradescantia anthers to make sufficiently accurate calculations
of dosages from internally distributed isotopes to determine whether
aberrations arise as a result of recoil and/or transmutation events in addi-
tion to ionization events.

ABERRATION PRODUCTION BY SLOW NEUTRONS

The cytogenetic effects of slow neutrons are being discussed separately
from those of other radiations because of certain unique features associ-
ated with the interaction of this type of radiation with biological mate-
rials. Fast neutrons arising from uranium fission can be moderated by
elastic collision with graphite or heavy water in a nuclear reactor until
their velocities are reduced to thermal energies (average 2200 meters/
second = 0.025 ev). Such neutrons do not behave like fast neutrons,
whose principal reaction of significance in biological materials is the
ejection of recoil protons from hydrogen atoms. Rather, thermal neu-
trons, because of their very low velocities and energies, are normally
captured by various atoms in biological materials. Such a capture reac-
tion produces an unstable compound nucleus with an excess of energy.
This unstable nucleus may then (1) emit a y ray immediately to form a
stable isotope, e.g.,

H1 + n -» [H2] -> H2 + 7

734 RADIATION BIOLOGY

(2) emit a heavy particle immediately to form a stable isotope, e.g.,

B10 + n-> [Bu]-»Li7 + a

or (3) emit a capture radiation immediately, forming a radioactive
daughter which then emits /3 or 7 rays at a rate characteristic of the
isotope formed, e.g.,

half life
N14 + n -> [N15] -> C14 + p ca. 6000 years -> N14 + /3~

The radiations of biological importance are the immediate "capture"
radiations and the delayed or "decay" radiations, since these produce
ionization. Thermal neutrons do not themselves appear to produce any
effect, since they are uncharged particles of low energies.

The reactions outlined indicate that there are two principal classes of
radiations arising from slow neutrons which should be of biological sig-
nificance, 7 radiation and heavy-particle radiations. As has been shown,
exchanges in Tradescantia microspores are linearly related to doses of
protons and a particles, but show a geometrical relation to doses of 7 rays.
Thus it should be possible, by determining the kind of dosage curve
obtained for slow neutron-induced exchanges, to decide the relative bio-
logical importance in chromosome aberration production of these two
classes of radiations arising from slow neutron exposures. Such experi-
ments have been performed, utilizing a special exposure facility in the
thermal column of the Oak Ridge reactor (Conger and Giles, 1950). The
results obtained indicate that the relationship with doses of slow neutrons
is linear for all types of aberrations, giving biological evidence that cap-
ture reactions resulting in the emission of particulate radiations are of
major consequence in producing aberrations. Physical calculations indi-
cate that the boron and nitrogen reactions are responsible for approxi-
mately 32 and 52 per cent of the total rep of ionization absorbed in
Tradescantia tissue, while the hydrogen reaction accounts for 16 per cent,
practically all the remainder.

Simultaneous X-ray exposures were made in order to compare the
relative biological efficiencies of thermal neutrons and X rays for the
production of chromatid and isochromatid breaks. After appropriate
corrections were made for 7-ray contamination in the neutron exposure
chamber, the observed thermal neutron-X-ray ratio for chromatid
aberration production was found to be approximately 11:1. If only the
nitrogen protons and boron a particles from thermal neutron capture are
considered, their efficiency compared with X rays seems to be about 15 : 1.
The expected ratio, calculated on the basis of previous radiation results
from external sources, was ca. 5.2:1. Thus it appears that particulate
radiations originating internally from capture reactions are considerably
more efficient (about three times) in break production than are similar
radiations from external sources. Since some evidence indicates a non-

chromosome aberrations in Tradescantia 735

random distribution of boron and possibly of nitrogen in cells, with higher
relative concentrations in the nucleus and possibly in the chromosomes, it
is suggested that this may account in part for the observed efficiency
differences. In addition, recoil and transmutation effects may also be of
importance. It thus seems probable that the greater effectiveness of
particulate radiations originating internally from capture reactions results
from both the site and mode of origin of these radiations.

RECAPITULATION

Before considering the effect of modifying factors, it seems appropriate
to summarize briefly the general theory of aberration production by ion-
izing radiation as discussed in the previous sections.

On this theory, as originally proposed by Sax and Lea and Catcheside, a
break in either a single (resting stage) or divided (prophase) chromosome
is due to the direct action on the chromosome of the ionization produced
by the passage through the chromosome of a single particle, such as an
electron, a proton, or an a particle. Such a break may then remain as
such giving rise to a terminal deletion, rejoin in the original position (the
process of restitution) and thus be undetected, or join with another
adjacent break in the same or in a different chromosome (the process of
reunion) to produce various types of aberrations.

Comparative experiments with different radiations under various con-
ditions indicate that several ionizations (average ca. 17) are required to
produce a chromosome break; that the majority of breaks produced
undergo restitution; that restitution is, for most breaks, a relatively rapid
process (the average restitution time being ca. 4 minutes, although some
breaks may remain "open" for considerably longer periods); and that
reunion is not at random, since for an exchange to occur the two breaks
involved must be produced with an initial separation of not more than
ca. 1 ft.

The quantitative relationship between a dose of radiation and the
yield of aberrations depends on both the type of aberration and the kind
of radiation. With X and y rays, certain types of breaks (simple, one-hit)
are linearly related to dose and are apparently produced by the relatively
concentrated ionization at the tail of a single electron track. Other
types, principally those involving breaks in separate chromosomes
(exchanges, two-hit types) increase as some power of the dose greater than
one. These aberrations apparently arise from the production of the two
separate breaks by the tails of two separate electron tracks. An intensity
effect with X rays exists for exchange breaks, as indicated by the fact that
the yield for exchanges, but not for simple breaks, is lower when the same
dose is delivered at a low as compared with a high intensity. If the dose
is given in a short time, all the initial breaks are present in the nucleus
simultaneously, reunion is favored, and the exchange yield is proportional

736 RADIATION BIOLOGY

to the square of the dose. If the dose is extended over a considerable
period (at a low intensity), restitution is favored over reunion and the
exchange yield is reduced.

With fast neutron and a-particle experiments, in which many fewer
ionizing particles traverse the nucleus than do electrons in comparable
X-ray exposures, all aberrations exhibit a linear relationship with dose.
Thus exchange aberrations apparently arise as the result of the simul-
taneous production by the same ionizing particle of the two breaks taking
part in the exchange. Confirmation of this fact comes from the evidence
that there is no effect of neutron intensity on aberration yield.

The interpretation of chromosome aberration production as just out-
lined has been quite generally successful in accounting for most of
the quantitative results of radiation experiments with Tradescantia.
Recently, however, experimental data of two sorts have been obtained
which indicate that this theory in its simplest form is not entirely ade-
quate. The first evidence was that obtained by Kotval and Gray (1947)
in their studies with a particles. On the basis of comparative ionization
distribution and particle numbers, the theory predicts that a given
amount of ionization produced by a particles should be considerably less
efficient in producing chromosome breaks than an equal ionization dose
produced by fast neutrons; whereas, the experimental results indicate
that, for equal ionization doses, a particles are somewhat more efficient.
It was concluded that a proportion of the breaks produced by a particles
arise from ionization produced in the immediate vicinity of, but not
within, a chromosome, thus suggesting the involvement of an indirect as
well as a direct mechanism. The second, and even more striking, evi-
dence was that obtained by Thoday and Read (1947), who noted a pro-
nounced effect of oxygen on the frequency of X-ray-induced aberrations
in the root-tip mitosis of the broad bean, Vicia faba. Their experiments
indicated that the absence of oxygen during irradiation resulted in a
marked decrease in aberration frequency and consequently supplied
additional evidence for the probable presence of indirect mechanisms
responsible for chromosome breakage. Extensive studies of the oxygen
effect on aberration production in Tradescantia microspores have also
been performed and the results of these experiments will be discussed in
one of the next sections on modifying factors. In that discussion an
attempt will be made to evaluate the significance of all these more recent
findings in relation to the mechanism of aberration production as pre-
viously described.

EFFECTS OF MODIFYING FACTORS

The effects of various modifying factors applied before, during, or after
exposures to radiations have been extensively studied in attempts to

chromosome aberrations in Tradescantia 737

elucidate further the mechanisms of chromosome breakage and reunion.
In certain instances, for example centrifugation effects, such studies have
served primarily to support previous hypotheses as to the process
involved. In other cases (e.g., infrared effects) the investigations have
revealed pronounced effects which may require modification of original
views, but for which no clear explanations are as yet available. Finally,
certain studies (the oxygen effect) have yielded results of paramount
significance for any interpretation of the biophysical and biochemical
mechanism of aberration production and appear to necessitate a definite
revision of certain aspects of older hypotheses. In view of the past and
probable future importance of studies of modifying factors, it appears
desirable to devote a rather considerable amount of space to a considera-
tion of these effects. Since it is felt that the oxygen effect is of special
significance, a major portion of the discussion will be devoted to the
available evidence relating to this effect. It should be borne in mind,
however, that many of the conclusions arrived at in this latter discussion
must be, of necessity, tentative ones, since such investigations have only
recently been initiated and much work remains to be done to clarify
various aspects of the problem.

CENTRIFUGATION; SONIC VIBRATION

Experiments of Sax (1943) have shown that centrifugation (2080 rpm)
during X irradiation approximately doubles the yield of chromosome and
chromatid interchanges as well as of isochromatid aberrations. When
centrifugation followed irradiation (the interval between cessation of
irradiation and initiation of centrifugation being 5 minutes) there was no
increase in aberration frequency. In control experiments, centrifugation
alone produced no effect. In analogous experiments in which sonic
energy (9100 cycles/second) was applied during X irradiation (Conger,
1948), an increased frequency of both chromosome interchanges and dele-
tions resulted, although the magnitude of the effect was less than that
found with centrifugation. In control experiments, no effect of this
treatment alone was detected, although chromosome breakage following
supersonic treatment (400,000 cycles/second) alone has been reported in
other plant material (Wallace, Bushnell, and Newcomer, 1948). These
experiments are interpreted as indicating that factors such as mechanical
stresses or vibrations can cause movement of broken ends produced by
irradiation, and thus promote new reunions yielding aberrations, as
opposed to restitution.

COLCHICINE

Brumfield (1943) has studied the effect of pretreatment with colchicine
on the frequency of X-ray-induced aberrations in Allium root tips. The
experimental roots were placed in a 0.05 per cent solution of colchicine for

738 RADIATION BIOLOGY

45 minutes immediately before exposure to 300 r of X rays. Subsequent
to irradiation, both experimental and control roots were kept in a colchi-
cine solution and a cytological analysis was made 48 hours later. There
was a marked reduction, by about two-thirds, in the frequency of X-ray-
induced chromatid aberrations in the root tips pretreated with colchicine;
whereas, chromosome aberrations were about equally frequent in the
experimental and control roots. These results are interpreted as arising
from an effect of colchicine in reducing the amount of chromosome move-
ment in prophase, where chromatid effects are induced, thus favoring
restitution as opposed to recombination. Since chromosome movement
at the resting stage is presumably at a minimum, the absence of an effect
of colchicine on chromosome breaks is as expected.

ULTRAVIOLET

The investigations of Swanson (1940, 1942) utilizing pollen tube
chromosomes (in Trade scantia) have shown that ultraviolet differs from
X rays in that this radiation produces only chromatid breaks and no
isochromatid breaks or chromatid exchanges. Subsequent to these
studies of the comparative effects of ultraviolet and X radiation indi-
vidually, experiments were performed (Swanson, 1944) in which ultra-
violet was used in combination with X rays. Pretreatment with ultra-
violet (2537 A) 1 hour before X irradiation produces an inhibition of all
types of visible X-ray breaks. Chromatid breaks and translocations are
relatively more affected than are isochromatid breaks. The degree of
inhibition depends on the dosage of ultraviolet, but the nature of this
proportionality was not established. The suggestion is made that the
inhibition resulting from ultraviolet pretreatment arises from an effect on
the chromosome matrix. This effect may result from a greater resistance
of the matrix to subsequent X-ray breakage, or from the failure of broken
ends having an ultraviolet-treated matrix to undergo new reunion, thus
presumably favoring restitution.

Posttreatment with ultraviolet has no inhibitory effect on isochromatid
breaks. However, translocations are inhibited, the effect decreasing as
the time after radiation at which the ultraviolet is applied increases (up to
1 hour). Chromatid breaks are inhibited even when ultraviolet is
applied as late as 1 hour after X irradiation. These posttreatment
inhibitory effects on chromatid translocations and deletions are inter-
preted as indicating an effect of ultraviolet in facilitating restitution as
opposed to reunion.

Similar experiments in Drosophila (Kaufmann and Hollaender, 1946)
involving posttreatment with ultraviolet of X-rayed spermatozoa also
resulted in a decrease in the frequency of gross chromosomal rearrange-
ments. Dominant lethals, however, were not affected. These results
are interpreted in the same way as those obtained in Tradescantia — as

chromosome aberrations in Tradescantia 739

indicating an effect of ultraviolet in favoring restitution as opposed to
recombination.

INFRARED

The original investigations of Kaufmann, Hollaender, and Gay (1946)
demonstrated a marked effect of pretreatment with infrared in increasing
the frequency of X-ray-induced chromosomal rearrangements in Dro-
sophila. This effect was subsequently confirmed in experiments with
Tradescantia microspores by Swanson and Hollaender (1946) and Swan-
son (1949). Pretreatment of whole inflorescences was carried out with
infrared (between 6000 and 11,500 A with maximum energy transmission
at 10,000 A) for varying periods of time (up to 96 hours) before exposure
to X rays, and cytological analyses were made at 22 hours following X-ray
exposure. All such pretreatments resulted in an increase in all types of
chromatid rearrangements with the increase in interchanges being most
marked. Posttreatments with infrared also resulted in an increase in the
frequency of chromatid breaks and exchanges. (In the initial pretreat-
ment experiments no effect on isochromatid breaks was found, but in
later experiments these also showed an increase.) As with pretreatment,
the effect on interchanges was most marked, and increased yields were
obtained even when posttreatments were delayed as much as 18 hours
after X irradiation. In order to explain the fact that both pre- and post-
treatments are effective, the hypothesis is proposed that both infrared
and X rays alone are capable of weakening the chromosome structure (in
addition to the normally produced X-ray breaks) and that such changes
only become realized as complete breaks upon the addition of the other
type of radiation.

Yost (1951) has investigated the effect of infrared pre- and posttreat-
ments on the frequency of chromosome breaks induced at the resting
stage. Again, both these treatments resulted in an increase in the fre-
quency of the aberrations scored — dicentrics and centric rings. Pre-
treatments were followed immediately by X irradiation and cytological
examinations made 96 hours later. The frequency of aberrations in
buds pretreated with infrared was, on the average, approximately
doubled. For tests of posttreatment effects, buds were X-rayed and
samples then given a single exposure to infrared at several successive
intervals following the X irradiation, up to 96 hours, at which time cyto-
logical analyses were made. As indicated, increases were noted in each
test and these were of approximately equal magnitude at all intervals
(and of the same order of increase as for the pretreatments). Further, the
unexpected result was obtained that all the aberration types observed
were chromosome types, even in instances where the infrared exposures
were made just before the cytological examination at 96 hours, when the
chromosomes were obviously in the prophase stage where they respond

740 RADIATION BIOLOGY

to X-irradiation breakage as double structures. It is concluded that the
type of aberration (whether chromosome or chromatid) is determined by
the time of X irradiation and is independent of the time at which the
infrared is applied. It is not yet clear why the additional reunion taking
place during infrared treatment does not correspond to the singleness or
doubleness of the chromosome as shown by X irradiation.

The Drosophila results obtained when infrared is used as a supple-
mentary treatment with X irradiation differ in certain ways from those
obtained in Tradescantia. In an extensive series of experiments, Kauf-
mann and his co-workers (Kaufmann, Hollaender, and Gay, 1946;
Kaufmann and Hollaender, 1946; Kaufmann, 1946; Kaufmann and Gay,
1947; Kaufmann and Wilson, 1949) found that pretreatment results in an
increased frequency of translocations, but not of dominant lethals
(analogous to isochromatid breaks in Tradescantia) or of recessive lethals.
Further, posttreatment of mature sperm has no effect. If, however,
posttreatment is carried out at the time of fertilization (when chromo-
some recombination is usually assumed to occur), the frequency of trans-
locations is increased. The action of infrared in Drosophila thus appears
to be restricted to an effect on processes involved in the formation of
chromosomal interchanges. The failure to detect an increase in the
single-break type of dominant lethal indicates that the higher frequency
of translocations is not simply a result of an over-all increase in chromo-
some breakage. These observations, together with the evidence that
infrared alone has no detectable effect, have led to the hypothesis that
this radiation acts in Drosophila to facilitate recombination (as opposed
to restitution) among breaks produced by X radiation. It also appears
that breaks may fall into two qualitatively different classes, since the
subsequent behavior of the broken ends of some is modified by pretreat-
ment while that of others remains unchanged. On this basis the pro-
posal is made that dominant lethals in Drosophila may arise at the time
of irradiation as a result of sister chromatid reunion; whereas, breaks
resulting in translocations do not undergo recombination until the time of
fertilization.

It is at present difficult to provide a comprehensive and unified inter-
pretation of the supplementary effects of infrared applicable to both
Drosophila and Tradescantia. The evidence that in Tradescantia all types
of breaks are increased in frequency by pretreatment appears to con-
stitute a real difference from Drosophila. However, the major effect of
pretreatment in Tradescantia seems to be on translocation frequency.
Again the major increase of posttreatment effects in Tradescantia, on the
basis of available data, appears to be on translocation frequency.

Evidence against the idea of a weakened chromosome structure giving
rise to true breaks upon subsequent irradiation as postulated by Swanson
and Hollaender (1946) is available from the experiments of Sax (1942).

chromosome aberrations in Tradescantia 741

In these experiments the X-irradiation of microspores during the resting
stage did not increase the subsequent sensitivity of the chromosomes in
these same cells when they were X-rayed in prophase. In view of these
results it appears improbable that infrared treatments following X
irradiation would be more likely to produce additional true breaks.

It is possible that these difficulties may be resolved and a better under-
standing of the mechanisms of the infrared effect be achieved as a result
of future research based on experiments briefly reported by Swanson and
Yost (1951). These authors find that a heat shock applied after infrared
exposure, and before X irradiation, will remove the infrared effect in
Tradescantia microspores. They suggest that infrared may act to pro-
duce a metastable state in the chromosome which renders it more sensitive
to X-ray breakage, and that this state is replaced by the normal state
following heat shock. They also state that a similar effect is obtained
when the order of treatment is X rays, heat, infrared and suggest that
X rays, too, may produce, in addition to the usual breaks, incipient breaks
or metastable states that are acted upon by infrared.

TEMPERATURE

As might perhaps be expected, temperature was the first modifying
factor whose effect was studied in Tradescantia microspores. Sax and
Enzmann (1939) demonstrated that the yield of chromatid aberrations of
all types was greater at low than at high temperatures for a given X-ray
dose. They interpreted these results as indicating an effect of tempera-
ture on the recovery process, such that restitution is favored at high and
reunion at low temperatures, since it was supposed that at low tempera-
ture a break would remain open longer allowing the broken ends a greater
chance, following chromosome movement, of undergoing reunion as
opposed to restitution. Although there was evidence in the early experi-
ments of an effect of postirradiation temperature changes, later experi-
ments failed to demonstrate this effect, and Sax (1947) concluded that
the temperature of the cells at the time of irradiation was the factor of
major importance. The general observations of Sax and Enzmann were
confirmed in independent investigations by Faberge (1940) on X-ray-
induced " fragments" and by Catcheside, Lea, and Thoday (1946b) on
chromatid aberrations, although the latter investigators failed to find
much temperature effect on chromatid deletions. Darlington and
La Cour(1945) have maintained that most or all of the temperature
effect in modifying aberration frequency is an indirect one resulting from
an influence of this factor on the timing of the nuclear cycle, and thus on
chromosome sensitivity to radiation. This objection has been met by
Sax (1947) and especially by Catcheside (1948), who showed that the
relatively brief exposures to high or low temperatures at the time of
irradiation had little or no effect on the relative temporal positions of the

742

RADIATION BIOLOGY

peak frequencies of various chromatid aberrations although the magni-
tudes of the peaks was greater at low than at high temperatures. These
results indicate that there had been no major modification by tempera-
ture of the timing of the nuclear cycle.

The discovery of the oxygen effect (to be discussed later) served to
reopen the problem of the temperature effect, especially the interpreta-
tion of this effect as operating solely on the recovery mechanism. Since
X-ray-induced aberration frequencies are positively correlated with
oxygen concentration and oxygen is more soluble in water at low than at
high temperatures, it seemed possible that all the apparent temperature

1.0-

0.10 0.20 0.30

VOLUME PER CENT DISSOLVED OXYGEN

040

O DISSOLVED 02 FROM PARTIAL PRESSURE OF 02 - 27°C

• DISSOLVED 02 FROM 5% 0? AT VARIOUS TEMPERATURES
Fig. 10-7. Comparative effects of temperature and dissolved oxygen on chromosome
interchange yield induced by 400 r of X rays. (Giles, Beatty, and Riley, unpublished.)

effects might be due to an indirect effect on oxygen availability. How-
ever, experiments of Giles, Beatty, and Riley (1951; and unpublished)
indicate that the relation between chromosomal aberration yield and
temperature can be attributed only in part to an effect of temperature on
oxygen solubility. In addition, there appears to be a rather large effect
of low temperature alone in increasing aberration frequency, such that
chromosome interchanges and interstitial deletions are much more fre-
quent than expected on the basis of an oxygen solubility effect alone at
low as compared to high temperatures as long as oxygen is present during
irradiation (Fig. 10-7). Whether this additional effect of temperature —
beyond that portion probably associated with oxygen solubility and thus
very likely with the initial breakage frequency — is to be attributed to an
influence on the recovery mechanism, in accordance with the earlier
interpretations, is not clear. It is also possible that the additional effect
may result from an influence of temperature on the formation or effective-

chromosome aberrations in Tradescantia

743

ness of substances responsible for actual breakage (to be discussed in detail
later). If, for example, the "half life" of such mutagenic substances is
increased at low temperature, this might permit these substances to
increase their relative spheres of effectiveness, within the nucleus, in
aberration production at low temperatures.

Additional evidence regarding the temperature effect is provided by
experiments (Giles, Beatty, and Riley, 1951; and unpublished) per-
formed in the absence of oxygen (in helium). The relation between

15-1

i i-

o

o:

u
a.

<

£ 0.7i
m

03-

0

r-

25
TEMPERATURE,°C

— i

40

• ISOCHROMATID BREAKS ( IN HELIUM - 200 r)

O ISOCHROMATID BREAKS ( IN OXYGEN- 150 f)
Fig. 10-8. Effect of temperature on yield of chromosome interchanges when X irradia-
tion is performed in 5 per cent oxygen (400 r at 50 r /minute), helium (900 r at 300
r /minute). (Data of Giles, Beatty, and Riley, unpublished.)

temperature and chromosome aberration frequency under these circum-
stances is just the reverse of that found when oxygen is present, more
interchanges and deletions being present in helium at high than at low
temperatures. The same relationship holds for all types of chromatid
aberrations as well. Data for isochromatid aberrations are shown in
Fig. 10-8. How this result is to be interpreted is not yet clear. If the
action of X rays in the absence of oxygen is considered to be largely
direct, as some evidence indicates (Giles and Beatty, 1950), then the
temperature effect would presumably be on the recovery process, such
that restitution is favored at low and reunion at high temperatures.
However, if the production of aberrations in the absence of oxygen is
assumed to result largely from the production of OH radicals, which is

744 RADIATION BIOLOGY

certainly a possible interpretation (to be discussed later), then the
temperature effect may result from an influence on either the production
or, more likely, the effectiveness of the OH radicals. The discovery of
this reversal of the temperature effect in the absence of oxygen makes it
unlikely that the effect of temperature is to be attributed solely to a
modification of the behavior of broken chromosome ends during the
recovery process. Hollaender et al. (1951) have reported preliminary
experiments indicating a similar reversal of the temperature effect with
X-ray-induced killing of bacteria.

Faberge (1948, 1950) has investigated the effect of X irradiation of
pollen at very low temperatures and finds that the breakage frequency
at — 192°C is only about one-fifth that at +25°. Furthermore, the sensi-
tivity curve for intermediate temperatures resembles, in general character,
that for hydrogen peroxide production plotted against temperature when
water containing oxygen is X-rayed (Bonet-Maury and Lefort, 1948).
Experiments have been performed also to investigate the combined effects
of temperature and of nitrogen (Faberge, 1950 and unpublished). At
+ 25°C the aberration frequency is reduced in nitrogen, as compared with
air. At — 192°C there is a reduction in aberration frequency in air com-
pared with +25° as previously observed; and in nitrogen, sensitivity is
still further reduced by about one half. Faberge feels that nitrogen and
very low temperature both reduce the sensitivity by the same amount,
but through independent mechanisms.

OXYGEN

The experiments of Thoday and Read (1947), using root tips of Vicia
faba, demonstrated that oxygen has a marked effect on chromosome sensi-
tivity to X radiation as measured by induced aberrations visible at
anaphase. The absence of oxygen during X irradiation reduced the
aberration frequency to about one-third that observed when the irradi-
ation was performed in the presence of air. Subsequent studies by the
same investigators (1949) showed that the oxygen effect was markedly
less when a-particle radiation was used.

A series of investigations (Giles and Riley, 1949, 1950; Giles and
Beatty, 1950; Giles, Beatty, and Riley, 1951; Riley, Giles, and Beatty,
1952; Giles, Beatty, and Riley, 1952; and Giles, 1952, and unpublished)
has been carried out utilizing Tradescantia inflorescences in order to con-
firm and extend these observations. In most of the experiments special
exposure chambers have been used to make possible the rapid removal or
introduction of various gases either before, during, or after irradiation
and to control the temperature of the inflorescences during exposures.
Most of the cytological observations have been confined to chromosome
aberrations — interchanges and interstitial deletions. However, some data
are also available for chromatid effects. The major experimental con-

chromosome aberrations IN Tradescantia

745

elusions which have emerged from these studies may be summarized as

follows.

1. There is a pronounced effect of oxygen in increasing the frequency
of X-ray-induced aberrations of all types. Results of experiments with
chromosome interchanges are shown in Fig. 10-9. The possibility that
other gases are responsible for the effect has been excluded.

90 180

X-RAY DOSAGE, r

270

360

EXPOSED IN 02
EXPT. I
EXPT. E

EXPOSED IN AIR

A EXPOSED IN N2
Fig. 10-9. X-ray dosage curves for chromosome interchanges induced in atmospheres
of oxygen, air, or nitrogen. (Giles and Riley, 1949. Figure reproduced by permission
of the authors and the editor of the Proceedings of the National Academy of Sciences.)

2. Oxygen must be present during the actual X irradiation to be
effective. Pre- and posttreatment exposures of cells to pure oxygen or
to anaerobic conditions for periods of time up to 1 hour have no effect.
The results with pure oxygen in microspores contrasts with the effects
observed by Conger and Fairchild (1951), who found that oxygen alone
can produce aberrations in pollen grain chromosomes.

3. Even in the complete absence of oxygen, in so far as this gas can
be completely removed from microspore cells, there is still an appreciable
aberration frequency.

746

RADIATION BIOLOGY

4. The percentage of increase in aberration frequency above this base
line depends on the percentage of oxygen present during irradiation.
This relationship is approximately linear between 0 and 10 per cent oxy-
gen, after which the increase in aberration frequency becomes more
gradual, until an approximate plateau is reached at 21 per cent. Data
for chromosome interchanges at a constant dose are given in Fig. 10-10.
Similar results have been reported by Baker (Hollaender, Baker, and
Anderson, 1952) for reciprocal translocations detected genetically in
Drosophila virilis.

5. The effect of oxygen in increasing the yield of X-ray-induced
aberrations is an immediate one. This has been demonstrated by experi-

O

or

UJ
Q.

to

LlJ

<
O

1.1 -

0.9

0.7

0.5

o

o

o
q:
i
o

0.3

0.1

AA A 7, A I I " I ""

0 10 20 30 40 50
(AIR)

60 70 80 90

— TT

100

PERCENTAGE OF OXYGEN IN EXPOSURE CHAMBER,
NORMAL ATMOSPHERIC PRESSURE

Fig. 10-10. Relation between percentage of oxygen and yield of X-ray-induced
chromosome interchanges (400 r at 50 r/min). (Giles and Beatty, 1950. Figure
reproduced by permission of the authors and the editor of Science.)

ments in which a rapid introduction of oxygen into cells during irradi-
ation is effected. It has also been shown that the removal of oxygen
during irradiation results in a decreased aberration yield. Data for
chromosome interchanges are presented in Table 10-2.

6. The effect of pressure during irradiation depends on the amount of
oxygen present in the gas mixture used. These studies provide further
evidence that the increase in aberration frequency is related to the actual
amount of oxygen dissolved in the cells.

7. Irradiation in pure hydrogen at normal pressure and under three
atmospheres of pressure effects little or no significant decrease in aber-
ration yield.

chromosome aberrations in Tradescantia

747

8. In agreement with previous studies on the temperature effect, an
increased aberration yield at low temperatures is observed when irradi-
ations are performed in the presence of oxygen in the range between
approximately 1° and 35°C (see Fig. 10-7), but this result cannot be
attributed entirely to an effect of temperature on oxygen solubility.

Table 10-2. Experiments Demonstrating that the Effect of Oxygen on the

Yield of X-ray-induced Chromosome Interchanges in Tradescantia Is Confined

to the Period of X-ray Exposure. (All Doses: 300 r at 300 r/Minute)

(Data from Giles and Riley, 1949)

Condition of buds

No.

Inter-

Interstitial

Series

of
cells

changes
per cell

deletions

No.

Pre-

treatment

Exposure

Posttreatment

per cell

1

Vacuum

Vacuum

Vacuum — 10 min

880

0.12 + 0.01

0.11 ± 0.01

2

Vacuum

Vacuum

Oxygen introduced
(within 3 sec) to 1500
mm Hg — 10 min

700

0.09 + 0.01

0.10 ± 0.01

3

Oxygen

Oxygen at 1500 mm Hg

Oxygen at 1500 mm Hg
— 10 min

150

0.70 ± 0.07

0.83 ± 0.07

4

Oxygen

Oxygen at 1500 mm Hg

Evacuation (within 25
sec); vacuum — 10 min

200

0.72 + 0.06

0.85 ± 0.07

5

Vacuum

1st 30 sec: vacuum
2d 30 sec: oxygen intro-
duced (within 3 sec) to
1500 mm Hg

Evacuation (within 25
sec); vacuum — 10 min

350

0.39 ± 0.03

0.50 + 0.04

6

Oxygen

1st 30 sec: oxygen at

1500 mm Hg
2d 30 sec: evacuation

(within 25 sec) to 1-2

mm Hg

Oxygen introduced
(within 3 sec) to 1500
mm Hg — 10 min

518

0.61 ± 0.03

0.59 ± 0.03

9. In the absence of oxygen, a reversal of the temperature effect is
noted, aberration frequencies being higher at high than at low temper-
atures (see Fig. 10-8).

10. When fast neutron radiation is used, an oxygen effect is also found,
but the magnitude of the effect is considerably less for all types of aber-
rations than it is with X radiation.

11. Experiments on the intensity effect with X rays in oxygen and in
nitrogen, using dosages of equivalent biological effect, have shown similar
decreases in the yields of chromatid exchanges in the two gases at low
intensities.

As a working hypothesis, to explain the oxygen effect, the view will be
taken that this effect results from an increased frequency of initial chro-
mosome breakage as a consequence of the action of certain active sub-
stances produced in the aqueous cellular medium when oxygen is present
during irradiation. Before discussing the evidence for this view and for

748 RADIATION BIOLOGY

the possible identity of the intermediate substances involved, it is neces-
sary to consider the evidence against other possible explanations for the
oxygen effect. The three other principal explanations for this effect on
chromosome aberration production, especially in Tradescantia, appear to
be the following: (1) an effect of oxygen, as compared with oxygen lack,
on some metabolic system or systems in the cell resulting in increased
radiosensitivity ; (2) an effect of oxygen itself on the recovery process
such that the reunion of broken ends, which results in aberrations, is
favored over restitution, which restores the original conditions; and (3) an
effect of oxygen, when present during irradiation, not on the breakage
process, but on the subsequent recovery process. Such a situation could
arise either from the production in the presence and absence of oxygen of
qualitatively different types of breaks with respect to their subsequent
behavior in restitution or reunion, or from the effect on the recovery pro-
cess of some product arising when oxygen is present during irradiation.

There would appear to be a distinct possibility that oxygen lack might
influence radiosensitivity by modifying certain aspects of cellular metabo-
lism, thus causing a decrease in chromosome aberration frequency. For
example, there is good evidence that radiosensitivity varies with different
stages in the mitotic cycle (cf. Sparrow, 1951), and an effect of oxygen (as
compared with nitrogen) in modifying the timing of this cycle might
result in an effect on aberration frequency (cf. Gaulden, Carlson, and
Tipton, 1949). Further, it seems possible that modifications in the rate
of cellular respiration might influence radiosensitivity. Other effects on
cellular metabolism (e.g., on nucleic acid synthesis) of a lowering or
increase in oxygen tension might also be anticipated. The major evi-
dence against the view that the oxygen effect operates by way of dis-
turbed cellular metabolism comes from the comparative studies of this
effect with different kinds of radiations, as emphasized by Thoday (1950).
Although there is a marked oxygen effect with X rays on chromosome
aberration frequency in Vicia root tips, there is little or no effect with
a particles. Similar observations have been made in Tradescantia (Con-
ger, unpublished) and, in addition, the oxygen effect with fast neutrons
has been found to be intermediate (Giles, Beatty, and Riley, 1952)
between that for X rays and a particles. If the effect of oxygen were a
metabolic one, there is no reason to suppose that the resulting modifica-
tion in radiosensitivity would vary with different kinds of radiations.

Further evidence against the possibility that a disturbance of the
mitotic cycle can explain the oxygen effect comes from the experiments
(Giles and Riley, 1950; Giles, 1952) in which it was shown that this
effect is an immediate one, since the introduction of the gas during irradi-
ation results in a marked increase in aberration frequency. That oxygen
could have influenced the timing of the mitotic cycle in these experiments
is clearly impossible. Furthermore, the experiments were performed on

chromosome aberrations in Tradescantia 749

chromosomes in the resting stage, a part of the mitotic cycle where there
is relatively little variation in radiosensitivity (Sax, 1938; Roller, 1946).
It also appears highly improbable that the introduction of oxygen during
irradiation could have modified nucleic acid synthesis rapidly enough to
have affected chromosome radiosensitivity if, in fact, the course of such
synthesis does influence radiosensitivity (Darlington and La Cour, 1945).
The possibility must also be considered, especially in Tradescantia,
that the effect of oxygen is on the recovery mechanism — the reunion, or
restitution of broken chromosome ends — rather than on the initial
breakage mechanism. Experiments have been performed (Giles and
Riley, 1950) which appear to eliminate the possibility that oxygen itself
may be influencing the recovery process. In these experiments, buds
were X-irradiated in a vacuum, and oxygen was introduced immediately
following the exposure. There was no increase in aberration frequency
over control experiments in which buds were irradiated in vacuum and
maintained in this condition for ca. 15 minutes following the exposure.

These experiments do not completely rule out the possibility that the
oxygen effect may still operate by way of the recovery process, since as
noted in possibility (3) above, it is conceivable that some substance pro-
duced in the cells when oxygen is present during X irradiation might
influence the reunion of broken chromosome ends or that broken ends
produced in the presence and absence of oxygen might be qualitatively
different with respect to their subsequent behavior during the recovery
process. Evidence against this interpretation is provided by intensity
experiments in oxygen and in nitrogen (Riley, Giles, and Beatty, 1952),
in which the restitution times for breaks giving rise to chromatid
exchanges in the presence and absence of oxygen are found to be
essentially identical. If the oxygen effect were operating by way of the
recovery process, a difference in the average restitution time might well
be anticipated. Further general support of the view that a major effect
on the recovery process is not involved comes from the comparative oxy-
gen effect with different types of radiations. It is found that the effect
of oxygen on aberration frequencies is inversely correlated with the
specific ionization of the radiation used (X rays, fast neutrons, and alpha
particles). There appears to be no reason to assume that breaks pro-
duced by these radiations, especially by X rays and fast neutrons, should
differ qualitatively with respect to a possible oxygen effect on the recovery
process. It is possible, however, to provide a reasonable hypothesis to
explain these observations on the basis of differential initial breakage,
as will be discussed later. It should be noted that, despite the evidence
just presented, the possibility cannot yet be excluded that the oxygen
effect, even though operating primarily on the breakage mechanism, may
also exert some influence on the recovery process.

The preceding evidence is taken to indicate that the effect of oxygen in

750 RADIATION BIOLOGV

modifying X-ray-induced chromosome aberration frequencies does not
arise as a result of general metabolic disturbances within cells produced
by the presence or by the absence of this gas. Further, the evidence
favors the view that the effect of oxygen is not on the recovery mecha-
nism, involving the restitution or reunion of broken chromosome ends.
It thus appears that the increased aberration frequencies observed in
the presence of oxygen probably result from an increased frequency of
radiation-induced chromosome or chromatid breaks when this gas is pres-
ent during irradiation. Consequently, the problem remains to determine
what mechanism or mechanisms can explain this increased frequency of
chromosome breakage by radiation, especially by X radiation, in the pres-
ence of oxygen. Further, consideration must be given to the way in
which the results of the oxygen-effect studies can best be fitted into an
over-all interpretation of the mechanism of chromosome aberration pro-
duction by ionizing radiations.

During the past few years, increasing evidence has accumulated which
indicates that many radiation effects in aqueous systems are largely
indirect, mediated by active substances resulting from the radiodecom-
position of water. Weiss (1944) concluded that these initial substances
are OH radicals and H atoms and has discussed their mode of origin in
irradiated water. Weiss (1944, 1947) and others (e.g., Allsopp, 1944;
Lea, 1946; Allen, 1948; Bonet-Maury and Lefort, 1948; Sparrow and
Rubin, 1952; Gray, 1952) have discussed the subsequent interactions of
these primary products with one another and with dissolved substances,
especially with oxygen, when water is irradiated with various radiations.

When pure water is irradiated the following reactions are believed to
occur:

radiation

H20 . . . H20 > H20+ . . . H20- (1)

(ionization)

H20+ > H+ + OH (2)

H20- > H + OH" (3)

The result of the electron transfer in reaction (1) and the subsequent
decompositions in reactions (2) and (3) are to produce H atoms and OH
radicals. The relative magnitude of the various reactions which follow
depends upon the type of radiation and the resulting geometric distribu-
tion of these primary products. The following reactions are all thought
to occur (Allen, 1948) :

H + OH^ H20 (4)

OH + OH -> H202 -> H20 4- O (5)

H + H -» H2 (6)

OH + H2 -> H20 + H (7)

H + H202 -> H20 + OH (8)

chromosome aberrations in Tradescantia 751

If oxygen is present in the water, as is normally the case in most bio-
logical systems, the following reactions can also occur :

H + 02 -> H02 (9)

H02 + H — H202 (10)

The experimental evidence, particularly that of Bonet-Maury and
Lefort (1948) and Allen (1948), indicates that in the absence of oxygen
there is very little decomposition of water by X rays, since little or no
H2, 02, or H202 can be detected. However, pure water is readily decom-
posed by a particles. The amount of H202 formed [presumably by
reaction (5)] is directly proportional to the dose. There are few experi-
mental data for other radiations, but Allen's results (1948) indicate that
neutrons also produce appreciable amounts of H202 in oxygen-free water.
When dissolved oxygen is present, H202 is produced by X rays, presum-
ably by way of reactions (9) and (10). The influence of such factors as
dose rate, pH, temperature, and amount of dissolved oxygen have been
investigated by Bonet-Maury and Lefort (1948). With a particles, the
yield of H202 when oxygen is present is approximately the same as in the
absence of this gas. This absence of an oxygen effect is apparently
related (Allen, 1948) to the closely spaced distribution of the H atoms and
OH radicals, such that reactions (4), (5), and (6) are favored, and conse-
quently H atoms are unavailable to participate in reaction (9). With
X rays, the more widely spaced distribution of the H atoms and OH radi-
cals does not favor reaction (6) and consequently reaction (9) does occur.
When dissolved hydrogen is present in water in place of oxygen, a back
reaction to form water, and thereby remove the OH radicals, takes place
[reaction (7)].

As a result of their observations that there is a marked effect of oxygen
on chromosome aberration production by X rays, and little or no such
effect with a particles, Thoday and Read (1949) suggested that the active
substance responsible for aberration production might be H202. Further
striking parallelisms between H202 production in water under various
conditions of irradiation, such as temperature and pH, and chromosome
aberration production under similar conditions, are discussed by Giles
(1952). Much of the radiochemical and biological evidence may be
interpreted as furnishing indirect support to the view that H202 is
important in aberration production. There is also direct evidence that
H202 is mutagenic in such organisms as bacteria (Wyss et al, 1948) and
molds (Neurospora) (Wagner et al, 1950; Jensen et al, 1951). The
difficulty of introducing this substance into cells such as microspores has
precluded a direct test of aberration production by H202 in Tradescantia.

Although the H202 hypothesis has considerable evidence to support it,
the radiochemical data, especially for X irradiation, suggest that other

752 RADIATION BIOLOGY

active substances are probably involved in aberration production in the
presence of oxygen. It seems particularly likely that the H02 radical
[resulting from reaction (9)] would itself be highly active biologically and
should produce effects similar to H202. Furthermore, recent evidence
from experiments on the oxygen effect with fast neutrons has made it
clear that there is an inverse relationship between the specific ionization
(ionization density) for a given radiation and the magnitude of the
oxygen effect (Giles, Beatty, and Riley, 1952). The best data, for iso-
chromatid breaks, indicate that the nitrogen-oxygen dose ratios (the
ratio of the dose of a given radiation in nitrogen to that in oxygen produc-
ing an equal aberration yield) for a particles, fast neutrons, and X rays
are, in order, 1.0, 1.4, and 2.6. These data serve to reemphasize the
importance of ionization (and subsequent radical) distribution as a factor
in radiobiological effects.

This inverse relationship between specific ionization and the oxygen
effect apparently can be interpreted best as arising from differences in the
distribution, and consequently in the interactions, of the primary radia-
tion products in water (H atoms and OH radicals) and dissolved oxygen.
Thus with a particles, the OH radicals and H atoms are closely spaced
and reactions (4), (5), and (6) are favored. Even with oxygen present
there is little opportunity for reaction (9) to occur, since H atoms are
rapidly removed by reaction (6). Thus no H02 radicals are produced
and no oxygen effect is noted. With X rays, however, the primary
radiation products are more widely spaced and dissolved oxygen can
react with H atoms to produce H02 [reaction (9)]. With radiations
having intermediate specific ionization, such as recoil protons produced
by fast neutrons, some oxygen effect might be anticipated, since reaction
(6) would be less favored in comparison with reaction (9) than is the
case with a particles.

It seems quite possible that reactions produced by OH and H02
radicals (and possibly by H atoms) rather than by H202 molecules, are
principally effective in producing chromosome aberrations and that a
certain average concentration of such radicals may be required for a
chromosome break to result. It should be noted, however, that the
probability also exists that H202 molecules are involved in a-particle
effects, even if they prove to be relatively unimportant in X-ray effects,
since the close proximity of OH radicals along the paths of these particles
should favor their rapid reaction to form H202. At the present time it
does not appear possible to decide whether OH radicals (and possibly
also H atoms) or H202 molecules are primarily responsible for the a-par-
ticle effects. Further, there is the additional possibility that active
products of secondary reactions, such as organic peroxides which are
known to be mutagenic (Dickey et al., 1949), are involved. This seems
rather unlikely, however, in view of the apparent localization of the

chromosome aberrations in Tradescantia 753

effects to the immediate vicinity of particle paths, a point which will be
discussed later.

It will be recalled that in the absence of oxygen, i.e., in X irradiations
performed in other gases such as nitrogen or helium or in a vacuum, there
is still an appreciable frequency of aberrations. The question may be
raised as to whether this residual frequency is due largely to indirect
radical effects even in the absence of oxygen, as has been assumed in the
immediately preceding discussion, or whether this effect arises from the
direct absorption of the radiation energy by the nucleoprotein structure
of the chromosome. The whole problem of distinguishing between direct
and indirect effects, particularly in a situation such as this one, becomes
exceedingly difficult. The distinction may in fact become largely mean-
ingless in instances where such complex structures as chromosomes are
involved, in which, for example, water molecules giving rise to active
radicals upon irradiation may occur within the volume of the chromo-
some and probably even to some extent bound to it.

Despite these difficulties, it appears worth while to consider the avail-
able evidence bearing on the question of the mechanism of the radiation
effect in the absence of oxygen. On a priori grounds, it seems reasonable
to expect that an indirect effect by way of OH radicals, and possibly H
atoms, would occur in an aqueous system such as a cell in the absence of
oxygen. The difficulty of demonstrating such an effect experimentally is
considerable, however. In the first place it is difficult to prove that all
the oxygen has been removed from cells by the evacuation procedure
employed. If it is assumed that the oxygen is in fact removed, then the
existence of an indirect effect in the absence of oxygen can best be demon-
strated by the efficacy of some protective substance in decreasing this
effect by reacting with the intermediate active radicals or atoms before
they can produce their biological result, in this instance, a chromosome
break. A major difficulty in testing for such an effect arises from the
problem of ensuring that the protective substance is actually penetrating
the cell. For Tradescantia inflorescences, penetration by gases has been
shown to be effective, hence an attempt was made to test for a protective
effect using H2, which is known on radiochemical evidence (Allen, 1948)
to promote the back reaction to form H20 in X-irradiated water by com-
bining with the OH radical [reaction (7)]. However, no clearly sig-
nificant decrease in aberration frequencies was found following irradiation
in hydrogen at normal pressure or at three atmospheres above normal,
compared with irradiation in a vacuum or in nitrogen at three atmos-
pheres above normal pressure (Giles and Beatty, 1950).

Evidence that a hydrogen effect can be detected, at least in a chemical
system possibly similar to the one in this biological experiment, is indi-
cated by the experiments of Scholes and Weiss (1950) who found a
decreased effect of X rays in disrupting nucleic acid (as measured by the

754 RADIATION BIOLOGY

ammonia yield) when hydrogen was present during irradiation, as com-
pared with irradiation in a vacuum or in oxygen. The negative results
of the Tradescantia experiment may be taken to indicate that OH radicals
are in fact not effective in producing chromosome breaks, and that the
X-ray effect in the absence of oxygen is primarily a direct one. However,
there is no unequivocal evidence that hydrogen is actually present in the
cell in sufficient amounts or in the appropriate locations to ensure that the
back reaction will occur, although the evidence indicating a rapid penetra-
tion and effectiveness of oxygen would suggest that a similar situation
exists for hydrogen. Furthermore, it is also possible that H atoms rather
than OH radicals are responsible for the biological effect. That this may
be true for certain chemical reactions is again suggested by the experi-
ments of Scholes and Weiss (1950), in which evidence is presented that the
liberation of phosphate from X-irradiated nucleic acid may result from
reactions involving H atoms rather than OH radicals. The conclusion is
thus not yet warranted that all or most of the X-ray effect on chromo-
somes in the absence of oxygen is a direct one. In other biological studies,
such as those involving the killing of bacteria by X rays (Hollaender,
1952), there is evidence that chemical protection in the absence of oxygen
can occur.

Further general evidence for the probable importance of the indirect
effect comes from observations on the relatively greater radioresistance of
dry as compared with soaked seeds (cf. Gustaffson, 1947), even though
these experiments have been performed in air and not in the absence of
oxygen. Regardless of the relative magnitude of the direct and indirect
effects in the absence of oxygen, it appears to be quite clear that the
indirect effect is of major importance when oxygen is present.

Even if the identity of the intermediate substances responsible for
radiation-induced chromosome breakage were unequivocally established,
the problem of determining the chemical structures involved and the
chemical reactions leading to breakage would remain to be elucidated.
It is not known, for example, whether the protein or the nucleic acid or
both components of chromosomes are ruptured in the initial breakage
reactions. Certain studies on the effects of irradiation on nucleic acid
and proteins in vitro and in vivo are pertinent to this problem, however.
The experiments of Sparrow and Rosenfeld (1946) demonstrated that
X rays can induce depolymerization of thymonucleohistone and of
sodium thymonucleate. The studies of Scholes and Weiss (1950) already
referred to indicate that this effect on nucleic acid may result largely from
indirect radical action. This conclusion is supported by the observation
(G. C. Butler, 1949) that the presence of glucose or methanol in the solu-
tion during irradiation partially protects the nucleic acid. J. A. V.
Butler and Smith (1950) conclude that the degradation of DNA can be
produced by the action of OH radicals. Taylor et al. (1948) demon-
strated that there is a continuing depolymerization of nucleic acid after

chromosome aberrations in Tradescantia 755

the cessation of radiation. The recent work of J. A. V. Butler and Con-
way (1950) indicates that this latter effect occurs only if oxygen is present
in the nucleic acid solutions at the time of irradiation. The immediate
depolymerization effect of X radiation is independent of oxygen con-
centration. It is noteworthy that the radiosensitivity of nucleic acid to X
rays increases by a factor of ~3 in the presence of oxygen over that found
under anaerobic conditions. This numerical increase is substantially the
same as that found for chromosomal aberrations induced in Vicia root tips
and Tradescantia microspores. Additional experiments are described
(ibid.) from which the conclusion is reached that H202 is not the effective
agent when irradiation is performed in the presence of dissolved oxygen.

In addition to the experiments on irradiation of nucleic acid in vitro,
others have been reported (Limperos and Mosher, 1950) on the effects of
irradiation in vivo. These workers conclude that substantial depoly-
merization occurs in living cells during irradiation, since they were able
to isolate almost completely depolymerized DNA from the thymus of
recently X-rayed rats.

Although these experiments on the effects of X radiation on nucleic
acid are important in suggesting possible mechanisms of chromosome
breakage, they do not establish that depolymerization is necessarily
involved in this process. It is still not at all clear, for example, that
changes in the nucleic acid of chromosomes, as opposed to those in the
protein component, are the significant ones in chromosome breakage.
These experiments do indicate, however, that the structure of one impor-
tant component of chromosomes can be markedly modified by indirect
radiation effects mediated by radicals or related active substances.

CONCLUSIONS

In concluding this discussion of radiation-induced chromosome aber-
rations in Tradescantia, it appears desirable to consider to what extent
recent evidence, particularly that derived from the oxygen effect, necessi-
tates a modification of earlier conceptions of the mechanism of chromo-
some aberration production by ionizing radiations. Prior to the dis-
covery of the oxygen effect, the view was usually taken that the effect of
radiations on chromosomes was a direct one resulting from the ionization
of the bonding electrons of the molecules composing the chromosomes by
the passage of the ionizing particles (Catcheside and Lea, 1943; Lea,
1946). However, the demonstration of the marked effect of oxygen in
modifying the frequency of X-ray-induced aberrations, plus the evidence
that this effect is probably on the breakage and not on the recovery proc-
ess, appear to invalidate this opinion that direct molecular ionization is
the only mechanism involved. Rather, it now seems most probable that
the major fraction of the radiation effect on chromosomes is an indirect
one, resulting from the action of active radicals, or of related substances

756 RADIATION BIOLOGY

derived from them, produced in the aqueous medium of the cell. Despite
this indicated modification of earlier views on the basis of the recent
results, it must be recalled that the major emphasis in the theory of
radiation-induced chromosome changes in Tradescantia has been that a
particular chromosome break results from several ionizations produced by
one particular particle (whether an electron, a proton, or an a particle)
and not from a single ionization or from a cumulative effect of several
particles. Thus an essential feature of this theory, for which the major
evidence has been derived from comparative experiments with X rays
and fast neutrons, has been the localization of the biological effect along
particle tracks, and the influence on this effect of differences in the pat-
terns of ionization distribution along such tracks with various radiations
(Lea, 1946). Since the original experiments were all performed in the
presence of oxygen (in air), the evidence that chromosome breakage
stems from events produced by single particle tracks remains valid,
whether such events result from direct ionization of the molecules of the
chromosome or from indirect effects produced by radicals arising in the
water along the particle track. The types of dosage curves for inter-
changes induced by X rays and fast neutrons in the absence of oxygen are
similar to those found when oxygen is present, again indicating a localized
breakage mechanism involving the passage of single particles, regardless
of whether the breakage in the absence of oxygen arises from direct or
indirect effects. Thus the major modification required of earlier opinions
is simply concerned with the nature of the chemical events involved in
chromosome breakage. Consequently, it is only necessary to replace the
concept of characteristic columnar patterns of ionization along various
particle tracks, with one of columns of active radicals having similar
patterns (or columns of their immediate products arising from radical
interactions or from reactions with other solute molecules, such as
oxygen). As emphasized by Thoday (1950) and Allsopp and Catcheside
(1948), the principal point at issue is one of the relative localization of
effects to the immediate vicinity of particle tracks. Even though
indirect effects involving radical formation are involved, the evidence
indicates that the effective diffusion of such substances is very limited.
As a consequence, although a particle involved in producing a chromo-
some break by way of indirect radical effects may not actually traverse
the chromosome thread, it seems clear that it must at least pass in the
immediate vicinity of the site of breakage.

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758 RADIATION BIOLOGY

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7G0 RADIATION BIOLOGY

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Manuscript received by the editor Jan. 5, 1952

CHAPTER 11

Immediate Effects on Division, Morphology, and Viability

of the Cell

J. Gordon Carlson

Department of Zoology and Entomology, The University of Tennessee,1 and
Biology Division, Oak Ridge National Laboratory2

Introduction. Action of high-energy as compared with ultraviolet radiations. Meas-
urement of effect. High-energy radiations: Mitotic effects — Morphological effects — Cell
viability effects. Ultraviolet radiations: Mitotic effects — Cell morphology effects —
Viability effects. References.

INTRODUCTION

Among the earliest known effects of ionizing and ultraviolet radiations
on the living organism were their capacities to produce changes in cell
morphology that were frequently followed by death of the cell and to
reduce the mitotic activity in tissues. The last two decades have seen a
great increase in interest in this field and in the publication of papers deal-
ing with the results of research into these effects. As a natural conse-
quence, the subject of cytological effects has been subdivided in these
volumes into several topics, each delimited by somewhat arbitrary and
often overlapping boundary lines, more often than not determined by
the special interests and the particular points of view of the authors. The
subject matter of this chapter will be confined to a review of those radia-
tion-induced cytological effects that are ordinarily evident within a
single mitotic cycle of treatment. Omitted are such important subjects
as chromosome aberrations, protective agents, photorecovery, develop-
mental effects, physiological effects, and effects on microorganisms, which
are dealt with in other chapters.

For discussions of certain aspects of the effects considered in this
chapter the reader is referred to reviews by Packard (1931), Duggar
(1936), Warren (1942), Lea (1946), Spear (1946), Giese (1947), Rajewsky
and Schon (1948), and Carlson (1950).

1 Contribution No. 67 from the Department of Zoology and Entomology.

2 Work performed under Contract No. W-7405-eng-26 for the Atomic Energy
Commission.

763

764 RADIATION BIOLOGY

ACTION OF HIGH-ENERGY
AS COMPARED WITH ULTRAVIOLET RADIATIONS

The cytological effects of treatment with high-energy radiations are
probably the results of chemical and physical changes induced in certain
molecules by the release of energy inside the living cell. This assumedly
results directly or indirectly from ionizations induced by the radiation.
Through the selection of the kind of radiation one can control the manner
in which the energy is distributed within the cell. Gamma rays, /3 rays,
and short wave-length (hard) X rays, for example, which have a low
specific ionization in air, release their energy in relatively scattered rather
than clustered loci. The ionizing particles produced by long wave-length
(soft) X rays, however, give off energy in dense clusters, while a rays and
protons release their energy in very densely ionizing columns. The
quantity of energy absorbed by different structures in the cell depends on
the nature of the ionizing particle and not on the chemical composition of
the cell part affected. For this reason, different kinds of high-energy
radiation give biologically detectable effects that are quantitatively, but
not qualitatively, different.

On the other hand, the different wave lengths of ultraviolet radiation of
interest to the biologist, i.e., those in the range of 2250-3650 A, induce
cytological changes that may be both quantitatively and qualitatively
different. Ultraviolet quanta are selectively absorbed by the different
molecules of the cell, a given molecule absorbing a certain quantum only
if it has a corresponding energy level. The energy of a given ultraviolet
wave length will, therefore, be absorbed with a resulting change in certain
types of molecules. Wave lengths around 2600 A, for example, will be
highly absorbed by the nucleic acids of the cell, while those around
2800 A will be more highly absorbed by certain proteins. The
absorbing molecules are altered in the process, and these changes may
be reflected in morphological and functional alterations of the cell
constituents.

The kind of radiation determines the types of biological materials and
the methods of study that can be used. The greatest latitude is possible
with deeply penetrating radiations, such as y rays, neutrons, and hard X
rays. Beta rays, a rays, and soft X rays, however, impose definite
limitations, for their paths in tissue are very short. If the cells to be
studied for effect are single cells or if they are situated at the surface of a
tissue, the amount of energy reaching them can be determined with
reasonable accuracy. If deeper lying cells are to be studied, however,
more penetrating radiation must be used. An alternative is the method
used by Gray and Read (1942), wherein root tips were immersed in
radon solution, which readily penetrates the material and emits a par-
ticles within the cells.

CELL DIVISION, MORPHOLOGY, VIABILITY 765

MEASUREMENT OF EFFECT

In determining the relation of a dose of radiation to the biological
changes induced, it is essential that allowance be made for two factors
that characterize this kind of experiment: (1) latency in the appearance
of the effect and (2) the extent to which recovery may have occurred
between treatment and observation. Each warrants a more detailed
analysis.

Biological effects do not become observable immediately on the cessa-
tion of treatment, even though the ionizations and excitations produced
by radiations occur within a fraction of a second after the impingement of
the photons. Different biological effects may become detectable in
minutes, hours, days, or even years after treatment. In part, this is due
to the time required for the series of physical and chemical changes that
must intervene between the photon absorption and the morphological
change. For example, in alteration of the form of the nucleolus by ultra-
violet the maximum observable change occurs at about 35 minutes after
treatment (Carlson and McMaster, 1951). In part the latency is due to
the series of biological changes that must run their course before the
appropriate stage for detecting the change is reached. Changes in rate
of mitosis can be determined only at the end of an interval of time during
which the rate is measured. Chromosomal aberrations are first detect-
able hours or days after treatment and then only at certain mitotic
stages and in certain kinds of tissues, while gene mutations may have to
pass through two or three generations of individuals, and so may not be
detected for months or years after they have been produced. Though
the present chapter deals only with what we term immediate effects,
these may not be observable until several hours after treatment.

A striking characteristic of living substance is the capacity to repair
deleterious changes produced within it by external agents. The effects
dealt with in this chapter, with the exception of cell lethality and pos-
sibly the chromosome "stickiness" evident soon after treatment, are
subject to repair within the cell.

The intermediaries by which ionizations within the cell lead to changes
in the rate of mitosis are not known. Probably the initial effect may be
viewed as either the destruction of something within the cell — perhaps an
enzyme or substrate that is necessary for mitotic progression — or the
production of a substance that exerts a toxic effect on mitosis. Recovery,
then, would result either from the replacement of the destroyed enzyme or
substrate or from the removal of the toxic substance. From the time
that treatment begins and the first effects are induced, therefore, the
recovery processes will begin and the cell will act to restore the original
conditions. At any given time between the start of irradiation and the
completion of recovery, the amount of radiation effect within the cell will

766 RADIATION BIOLOGY

be the initial effect less the amount of recovery. This is referred to as the
residual effect. The dose of radiation that would be required at any
instant to produce an effect quantitatively identical with the residual
effect is known as the cumulative dose. Ideally the residual effect should
be expressed in terms of a deficiency of an essential substance or in the
amount of toxic material that is acting to produce the biological effect,
but this is not possible at present because of our lack of information on
the radiation-induced chemical and physical changes responsible for the
effect observed.

It is these two factors, latency and recovery, that often make it difficult
to determine accurately the immediate effect of a given dose of radiation.
The measurement of mitotic delay, for example, depends on the timing of
irradiated cells or the making of cell counts at certain intervals of time
following treatment. Since recovery may be assumed to be in progress
during this period and since the maximum observed biological effect
appears some time after the primary changes are induced, the accuracy
of our determination of initial and residual effect has definite limitations,
and these must not be overlooked in choosing a material and a method
and in interpreting results.

HIGH-ENERGY RADIATIONS

MITOTIC EFFECTS

Methods. The type of material selected by the researcher for study
determines to a large extent the type of mitotic problem that can profit-
ably be studied. The most direct approach to the problem of how radia-
tions affect mitosis is offered by observations on individual living cells.
Hanging-drop preparations of neuroblasts of the grasshopper, Chorto-
phaga viridifasciata, in artificial culture medium are very useful for such
studies. These cells are large, the internal structures are clearly visible
in the living cell, and all the mitotic stages are readily identifiable. It is
possible, therefore, to treat a cell at a known mitotic stage and then record
its behavior and time its progress subsequent to irradiation. A present
shortcoming of this method is our inability to maintain these cells at a
normal rate of division for more than 6-8 hours after the preparation is
made up; as a result, experiments designed to test recovery from large
doses, which may delay mitosis for several hours, are not feasible.

The marine invertebrate egg, such as that of Arbacia, also makes
possible treatment at a known stage of division and accurate timing of
subsequent mitotic progress, but the means of obtaining the data is quite
different. All of a lot of eggs from a single female will progress mitotically
at about the same rate after fertilization. Since stages in the mitotic

CELL DIVISION, MORPHOLOGY, VIABILITY 767

cycle of the zygote can be identified accurately only in fixed material, the
procedure is to determine the mitotic stage at any desired time by fixing a
sample of the eggs for future examination. Cleavage delay is customarily
expressed as the difference in time required by the treated and the control
eggs to progress from the stage of treatment to cleavage of 50 per cent of
the zygotes. If sperm or unfertilized eggs are treated, cleavage time is
measured from insemination. In addition to giving precise data on the
relation of dose to cleavage delay, this material is useful in obtaining
quantitative data on the degree and rate of recovery from mitotic
retardation.

Mitotically active parts of living plants or animals in vivo or in vitro
may be irradiated and fixed at the end of a certain time interval. The
effect on mitosis can then be determined by comparing the proportions of
cells in different mitotic stages with those of controls. Since the fre-
quency of different stages after treatment depends on both the dose and
the time after treatment at which the material is fixed, an accurate
picture of mitotic events following treatment is secured only by the
analysis of a series of preparations fixed at short time intervals after
irradiation. Unlike the marine invertebrate egg, this kind of tissue will
ordinarily consist of cells in interphase and various stages of mitosis at the
time of irradiation. The results are therefore expressed as changes in the
proportion of cells in different stages at different times after irradiation.
Part of the error inherent in the use of different preparations for different
counts can be eliminated by the use of culture preparations in which suc-
cessive counts can be made of the same group of cells.

It cannot be emphasized too strongly, however, that valid conclusions
regarding the effect of a given treatment on mitosis cannot be drawn from
counts alone of cells in different stages of the mitotic cycle at intervals
following treatment. For example, an increase in the number of cells in
middle prophase following a certain treatment must be the result of cells
entering middle prophase at a faster rate than they leave it. This may
be brought about in different ways, none of which could be confirmed or
negated by counts alone. It could result from: (1) an increase in the
mitotic rate of cells in early prophase, so that a number greater than
normal enters middle prophase in a given time; (2) a decrease in the
mitotic rate of cells in middle prophase, so that a number smaller than
normal leaves middle for late prophase in a given time interval; (3) a
reversal of mitotic progress in late prophase, so that late prophase cells
regress to become middle prophase cells ; or (4) any combination of these.
It may be seen, therefore, that a simple conclusion that treatment has
accelerated division, or that it has retarded it, cannot be reached merely
by comparing mitotic counts made at intervals after treatment. All too
often one comes across the wholly unjustifiable conclusion that, because

768

RADIATION BIOLOGY

the proportion of dividing cells in the tissue under consideration is greater
under one set of experimental conditions than another, the mitotic
activity has been increased or mitosis has been stimulated.

Nature of Effect. Direct observations of mitosis in hanging-drop
preparations of living Chortophaga neuroblasts following X irradiation
have demonstrated that there exists in these cells what we may term a
critical period between late and very late prophase (Fig. 11-1) (Carlson,

SUPH5!

Fig. 11-1. Mitotic cycle of Chortophaga neuroblast in vitro at 38°C (from Carlson and
Hollaender, 1948).

1941, 1950). Irradiation of a cell before it has reached this period usually
causes it to stop or even to revert mitotically; irradiation after it has
passed this period affects its progress through mitosis little if at all. The
cell passes through this critical period about 5 minutes before the nuclear
membrane disappears. At this time the chromosomes have almost
reached the end of prophase contraction, the shape of the cell is changing
from concavo-convex to spherical, the nucleoli are disappearing, and the
cytoplasmic viscosity is falling rapidly (Carlson, 1946). Neuroblasts that
have passed this stage at the time of treatment complete mitosis with

CELL DIVISION, MORPHOLOGY, VIABILITY 769

little or no delay, if the dose is less than 250 r. Larger doses produce a
delay that increases with the dose. This seems to result partly from
delay in the breakdown of the nuclear membrane and partly from chromo-
some "stickiness," which prolongs the anaphase separation of daughter
chromosomes. The more recently the irradiated cell has passed the
critical period, the greater is the delay in completing mitosis. If the cell
is in prophase at the time of treatment, but has not reached the critical
period, the effect on mitosis is determined both by the dose of radiation
and by the nearness of the cell to the critical period. After a very small
dose, such as 8 or 16 r, most cells in middle or late prophase,3 which have
not reached the critical period, are stopped mitotically for a length of time
that increases with increase of dose and decreases with rise of temperature.
The cell treated in interphase or early prophase is gradually slowed as it
approaches the critical period. Larger doses, 250 r, for example, produce
mitotic stoppage of those prophase cells that are nearing the critical
period. This is followed by reversion or simulated reversion to a stage in
which the chromatin resembles that of interphase (Carlson, 1940, 1941,
1950). Prophases are entirely absent from the tissue until recovery has
occurred; then these cells, together with those retarded in interphase,
again progress mitotically.

The most extensive series of studies yet made to determine the effect of
irradiation on the mitotic rate of cells treated at a known stage of mitosis
have been carried out by Henshaw and his co-workers on the marine
invertebrate egg. If Arbacia eggs or sperm are X-rayed and used immedi-
ately afterward in insemination, the first cleavage division is delayed, and
the larger the dose of radiation to which they have been exposed, the
greater is the delay (Fig. 11-2) (Henshaw, 1932, 1940a). Irradiation of
either sperm or egg is about equally effective in retarding cleavage
(Henshaw and Francis, 1936; Henshaw, 1940b; Henshaw and Cohen,
1940). When sperm alone are treated, it is found that the amount of
delay varies linearly with the logarithm of the dose (Henshaw, 1940a).
If the eggs and sperm are both treated, the zygote shows a greater delay
in the first cleavage division than if one or the other is treated (Henshaw
and Francis, 1936), but the effects are not additive in the sense that the
delay in the zygote formed from irradiated egg and irradiated sperm is
equal to the sum of the delays produced in zygotes formed from irradiated

3 Recent unpublished studies in our laboratory by Nancy D. Wolfson indicate that
appreciable numbers of neuroblasts treated at late prophase with 32 r of X rays pass
the critical period and complete mitosis with little or no delay. Since mitotic activity
falls to zero after this dose, however, cells in middle prophase at treatment must suffer
complete temporary blockage at some stage between middle prophase and the break-
down of the nuclear membrane. After so small a dose perhaps a certain amount of
time is necessary for the physical and chemical changes induced to be translated into
mitotic blockage.

770

RADIATION BIOLOGY

egg and untreated sperm and from untreated egg and irradiated sperm.
An additive effect in this sense would hardly be expected, unless recovery
from the irradiation effects took place successively in the maternal and
paternal parts of the zygote. It seems more likely that it would occur
concurrently in both. If such is the case, cleavage of the fertilized egg
would always have to await the recovery of the gamete with the greater
radiation-induced effect. The combining at random, therefore, of irradi-
ated eggs and sperm, which show individual variation with respect to
their radiation-induced damage, would in itself result in a greater average
delay than if eggs or sperm alone had been treated.4 Lea (1938b, 1946),
concluded that one would expect the cumulative doses, but not the effects

Q 100-

LU
Q

>

Q
CO
O

e>

LU

60

Ld

O

cr

LU

20'

MIN EXPO. 0 (CONTROLS)

O

/
O

o

20

O O-fCP P

60

100 140 180 220

MINUTES AFTER FERTILIZATION
Fig. 11-2. Relation of time after insemination to per cent of Arbacia eggs cleaved for
several doses of X rays administered at 7800 r/min to sperm before fertilization {after
Henshaw, 1940a).

as measured by delay in cell division, to be additive. He found that the
curve calculated on this basis fitted the experimental observations
satisfactorily.

In order to determine whether cleavage delay is due to a retardation of
one stage or is spread over the whole period from insemination to cleavage,
Henshaw (1938, 1940b) fixed samples of fertilized eggs, in which sperm or

4 1 am indebted to Mr. Jack Moshman for some calculations bearing on this point.
Using the variation in cleavage time after treatment of the sperm (Henshaw, 1940a.
Table I, 4-minute exposure), which shows a normal distribution, and assuming that
the range and delay would be similar in the egg, we find that a random combination
of the gametes would increase the time for 50 per cent cleavage by about 3J^ minutes.
This is approximately half the increase in delay produced by treatment of both
gametes over sperm alone, as indicated in another paper (Henshaw and Francis, 1936,
Fig. 1) for a comparable dose. It should be pointed out that this test, as set up, gives
a maximum increase in delay for irradiation of both gametes over either alone. If one
gamete is more sensitive than the other, the effect for irradiation of both will be closer
to that resulting from irradiation of the more sensitive one.

CELL DIVISION, MORPHOLOGY, VIABILITY

771

egg had been X-rayed, at short time intervals after insemination to com-
pare their progress with that of untreated controls. He found no retarda-
tion from entrance of the sperm head through fusion of the pronuclei;
considerable retardation from crescent formation (early prophase)
through late prophase, and slight retardation at metaphase, anaphase,
and telophase (Fig. 11-3).

The relative radiosensitivities of the different stages of the sea urchin
egg from insemination to first cleavage were tested by Yamashita, Mori,
and Miwa (1939) and Henshaw and Cohen (1940). Their results are in

MITOTIC TIME SCHEDULE

NS + NE 123
NS + IE

123

IS + NE

123

5 678

6 7 8

6 7 8

a

NS + NEH23

4

5

6 7 8

__---~

NS + !Erf23

4

5

6 7 8

4

5

6 7 8

IS +NE'LI_2_3

Fig. 11-3. Diagram to show the effects of X radiation on the duration of the different
stages of the first cleavage division of Arbacia. At top is a semidiagrammatic repre-
sentation of the mitotic time schedule of Arbacia {patterned after Fry, 1936). 1,
Entrance of sperm head into egg; 2, elaboration of sperm aster; 3, fusion of pronuclei;
4, crescent formation and disappearance; 5, prophase; 6, metaphase; 7, anaphase; 8,
telophase. N, normal ; I, Irradiated ; S, sperm ; E, egg (after Henshaw, 1940b).

close agreement. The former investigators, using Pseudocentrotus, found
that maximum delay of the first cleavage division results when irradiation
(X, 7, or (3 rays) takes place during the period extending from fusion of
the pronuclei into early prophase. Treatment during approach of the
sperm to the egg nucleus was somewhat less effective, while treatment
during late prophase, metaphase, anaphase, and telophase caused little
delay. Henshaw and Cohen (1940) X-rayed similar samples of Arbacia
zygotes at 5-minute intervals during the first 45 minutes after insemina-
tion. Using the Arbacia mitotic time schedule of Fry (1936) (see Fig.
11-3) to determine the stage at which each sample of zygotes was irradi-
ated, these investigators concluded that sensitivity as measured by
cleavage delay was greatest in zygotes treated during fusion of pronuclei.

772

RADIATION BIOLOGY

Stages arranged in successively decreasing order of sensitivity were:
entrance of sperm head into egg cytoplasm and its approach to the egg
nucleus, early prophase, and late prophase. No delay resulted from
treatment at metaphase.

Yamashita, Mori, and Miwa (1939) also determined the effect of
irradiating different stages of one- and two-celled Pseudocentrotus eggs on
the length of the second cleavage division. They found that it was
delayed only slightly by treatment of the one-celled stage at metaphase or
earlier, markedly by treatment at anaphase and telophase, and maximally
by treatment at the beginning of the two-celled stage.

<

Q

<r
a:

IS)

LU

(-
Z>

5

60 min
exposure

o

20 60 100 140 180

MINUTES AFTER IRRADIATION BEGAN WHEN EGGS WERE INSEMINATED
Fig. 11-4. Relation of time intervening between beginning of X irradiation and
insemination of Arbacia eggs to the cleavage delay, showing recovery from effect;
dosage rate, 520 r/min (after Henshaw, 1932).

The rate of recovery in the Arbacia egg has been shown by Henshaw
(1932) to be exponential. Eggs exposed to different doses of X rays
at the same dosage rate were inseminated at different times after the end
of treatment. For a series of exposures ranging from 5-60 minutes at
520 r/minute he found that the longer the delay between irradiation and
insemination, the less the first cleavage was delayed (Fig. 11-4). When
minutes delay on a logarithmic scale was plotted against minutes inter-
vening between irradiation and insemination, a straight line was obtained.

The doses and dosage rates used in these experiments on the sea urchin
are large, ranging from 2600 to 249,600 r delivered at rates of 120 to
7800 r/minute. These doses are of a much greater order of magnitude
than those used with most other kinds of material. Cleavage delays
recorded are from a few minutes to somewhat less than 3 hours.

Both Chortophaga neuroblast and sea urchin egg experiments indicate

CELL DIVISION, MORPHOLOGY, VIABILITY 773

that prophase is the mitotic stage most prolonged when the cell is irradi-
ated at an earlier stage. Exact comparisons, however, are not possible
at present, for Henshaw has not tested the immediate effect of irradiation
on the duration of the stage treated, and we as yet have little data on the
times required for neuroblasts X-rayed in different stages of mitosis to
complete mitosis.

In dealing with a tissue in which the cells are in different mitotic stages
and in which the stage of any given cell is not known at the time of irradia-
tion, the experimental procedures are quite different and the interpreta-
tion of the results considerably more involved than when the individual
cells or groups of cells are in known stages during irradiation. A brief
survey of the postirradiation changes in the proportion of cells in different
mitotic stages will be followed by a comparison of the results and conclu-
sions of different investigators who have studied different kinds of tissues.
Exposure of a tissue to ionizing radiations is followed by a decrease in the
number of prometaphases, metaphases, anaphases, and telophases pres-
ent, the amount of the decrease being positively correlated with the dose
of radiation to which the material is exposed. If the dose is sufficient to
reduce the numbers of cells in these stages to zero, the length of time
during which they will remain at zero is directly related to the dose. In
such experiments the order of disappearance of the stages is prometa-
phases first, then metaphases, then anaphases, and finally telophases
(Carlson, 1942). During recovery from the irradiation effect these stages
reappear in the same order. As the number of cells in these stages
increases, it will temporarily exceed the normal if the dose is so small as to
produce mitotic delay only during part of the mitotic cycle. If the dose
is large enough to affect cells at all periods of the mitotic cycle, however,
the number of cells in these stages as a rule never exceeds the original.
With this general picture in mind of the events that follow irradiation of a
mitotically active tissue, we can undertake a more detailed analysis of
the radiation-induced changes.

While there is general agreement that cells in prometaphase, meta-
phase, anaphase, and telophase at the time of irradiation subsequently
complete mitosis with little or no delay and that cells in interphase at
treatment may be prevented from entering mitosis by sufficiently large
doses of radiation, difference of opinion exists about the immediate reac-
tion of prophase cells to irradiation. Because the number of cells in
prophase is reduced as a result of irradiation, it has generally been
assumed that cells in this stage, like those in prometaphase-through-
telophase, are not very radiosensitive and so complete mitosis after
irradiation with little or no delay. This conclusion has been reached
from studies of mammalian tumors (Mottram, Scott, and Russ, 1926;
Warren, 1937), chick fibroblasts in vitro (Strangeways and Hopwood,
1926; Canti and Spear, 1929; Spear, 1931, 1932; Love, 1931; Lasnitski,

774 RADIATION BIOLOGY

1940), rat retina cells (Tansley, Spear, and Gliicksmann, 1937), crocus
root tips (Stone, 1933), and Tradescantia pollen grains (Roller, 1943). In
the Chortophaga neuroblast, on the other hand, middle and late prophase
cells show the greatest radiosensitivity, with early prophases and inter-
phases exhibiting successively less sensitivity (Carlson, 1941, 1950).
Deufel (1951) has also described a slowing down of prophases in the
Vicia root tip soon after treatment. Three possible explanations of these
apparently contradictory results may be considered.

First, the method used to determine the stage of greatest sensitivity
may not be adequate. Decrease in number or disappearance of prophase
cells from an irradiated tissue does not necessarily mean that blockage
is at late interphase and that the prophases have continued through
mitosis unchecked. As pointed out in an earlier section (p. 709), X
irradiation may lead to a disappearance of prophases by causing them to
revert to a condition resembling interphase, the real point of blockage
being in late prophase.

Second, the mitotic stage of greatest sensitivity may differ in different
kinds of cells, so that in certain cells blockage occurs at late interphase or
early prophase while in others it takes place at late prophase. If this
were true it would mean that the mitotic radiosensitivity of the cell
depends on factors other than those we commonly use in distinguishing
interphases and early, middle, and late prophases, namely, chromosome
morphology.

Third, the appearance of the prophase chromosomes of different kinds
of cells may be so different, because of differences in size and visibility,
that early prophase in one tissue may resemble late prophase in another
(Carlson, 1942), especially when examined in the living condition.
According to the description of Strangeways (1922) of the chick fibroblast
in vitro the beginning of prophase is preceded by spheration of the cell
and followed immediately by the disappearance of the nucleoli, the dura-
tion of prophase averaging about seven minutes at 39°C. In the Chorto-
phaga neuroblast culture, on the other hand, both the rounding-up of the
cell and the disappearance of the nucleoli occur within a few minutes of
the end of a prophase that extends over an average period of 102 minutes
at 38°C (Carlson and Hollaender, 1948; Carlson, 1950). It would seem,
therefore, that the long early and middle prophases of the grasshopper
neuroblast, which are characterized by slender, highly convoluted intra-
nuclear chromosomal threads, are passed over very briefly or even
omitted, perhaps because of the invisibility of the fine threads in the
smaller fibroblast, and what is termed late prophase in the neuroblast may
correspond to what is thought of as the whole prophase in the fibroblast.
In their study of the effects of y rays on the rat retina, Tansley et al.,
(1937) classed as prophase "all the early changes in the nucleus up to the
appearance of discrete chromosomes," and as metaphase, the stage from

CELL DIVISION, MORPHOLOGY, VIABILITY 775

the "appearance of chromosomes to the beginning of the journey to the
poles." The fact that their "prophases" are about half as abundant as
their "metaphases," as determined by counts in untreated preparations,
suggests that their "prophase" represents a portion of the mitotic cycle
comparable in extent to that in the chick fibroblast. Regardless of the
terminology adopted by the particular worker, until contradictory evi-
dence from observations of the mitotic history of living, irradiated cells
other than grasshopper neuroblasts is forthcoming, the conclusion seems
justifiable that the mitotic radiosensitivity is greatest shortly before
breakdown of the nuclear membrane when the chromosomes are clear
and well formed, and that whether or not an irradiated cell will continue
through mitosis or be stopped mitotically depends on whether or not it
has passed this stage.

Comparison of the results obtained from studies by different investi-
gators of the effects of ionizing radiations on the mitotic activity of differ-
ent tissues is difficult, especially in certain of the earlier studies, because
investigators (1) failed to control the temperature following treatment,
(2) used the term "mitosis" to include different and often undesignated
division stages, (3) made mitotic counts at very infrequent and irregular
intervals, or (4) did not make accurate measurements of the doses given.
Nevertheless, some general conclusions can be reached from an analysis of
the various studies.

The rate of decrease of mitotic activity immediately after irradiation is
greater in animal than in plant tissue. This is probably correlated with
the length of the mitotic cycle, which apparently is usually shorter in
animals (Lewis and Lewis, 1917; Strangeways, 1922; Wright, 1925; Carl-
son, 1941; Carlson and Hollaender, 1948) than in plants (Gray and
Scholes, 1951). The times after treatment for the number of dividing
cells to reach a minimum are shown in Table 11-1. The mitotic activity
of the animal cells reaches a minimum 3^-6 hours and that of plants 9-21
hours after treatment. An exception is the value of 3 hours found by
Marshak (1937) for root tips of several plant genera and by Gray et at.,
(1940) for root tips of Vicia after small doses. The former paper contains
so little information on this particular point that it is not possible to
evaluate the evidence. In the latter paper it is actually stated that
"mitosis is not far from its minimum 3 hours after irradiation."5 The
small dose used may also have some bearing on this low value (see Table
11-2).

When the exposure time is short, the duration of the postirradiation
period of mitotic decrease is positively correlated with the dose; the larger
the dose, the more extended is the time interval between treatment and
the period of minimum mitotic activity (Table 11-2, Fig. 11-5). The

6 Gray in a personal communication to me states that "the true value could easily
have been as late as 6 hours, since the minimum tends to be rather flat."

776

RADIATION BIOLOGY
Table 11-1

Interval

Organism

Tissue

Radia-
tion

Temp.

between

treat, and

exam.", °C

between
treat, and
minimal
mitotic
activity,
hours6

Reference

Grasshopper

Embryonic cells exclusive
of neuroblasts

X

28

3

Creighton, 1941

Neuroblasts

X

26

W

Carlson, 1940, 1942

7

38

1

Carlson et al., 1949

Salamander

Cornea

X

—

6

Alberti and Politzer,
1924

Tadpole

Brain and eye

7

RT

2

Spear and Gliicksmann,
1938

Chick embryo

Neural epithelium,
mesenchyme

X

39

y.

Regaud et al., 1925

Neural tube

X

39

y

Butler, 1932

Fibroblasts in vitro

X

39

m

Strangeways and Oak-
ley, 1923; Strange-
ways and Hopwood,
1926; Lasnitski, 1940

7

39

H-2

Canti and Spear, 1929;
Spear, 1931; Wilson
et al., 1935; Simon-
Reuss and Spear, 1947

7. X

39

i

Kemp and Juul, 1930;
Juul and Kemp, 1933

0

39

m

Lasnitski, 1948

Rat

Retina

7

BT

1-3

Tansley et al., 1937

Fast neu-

BT

1-3

Spear and Tansley,

trons

1944

Carcinoma

7

BT

m-m

Warren, 1937

X

BT

2

Luther, 1943

Mouse

Epidermis, adrenal gland,
lymph node, jejunum

X

BT

1-2

Knowlton et al., 1948;
Knowlton and Hemp-
elmann, 1949

Sarcoma

X

BT

3

Marshak, 1937

Vicia

Root tip

7

—

9

Mottram, 1936

7

25

3

Gray et al., 1940

X

18

Jungling and Langen-
dorff, 1930

RT

19-21

Pekarek, 1927

18

12

Deufel, 1951

Scilla
Vicia

Root tip

X

— ■

>7H

Marquardt, 1938

Pisum

Allium

■

Root tip

X

—

3

Marshak, 1937

Lycopersicum,

a — •. no information on temperature. RT, room temperature. BT, body temperature.
b In most of these studies the minimal mitotic activity was zero or close to zero.

CELL DIVISION, MORPHOLOGY, VIABILITY

777

overlaps between certain of these doses shown in Table 11-2 are obviously
not real but due to the fact that counts were not made frequently enough
to bring out minor differences. After doses up to those barely sufficient
to cause a fall to zero in mitotic activity, the duration of the period of
decreasing mitotic activity of a given tissue is determined by the rate at
which the cells recover and begin to progress mitotically after treatment.
Since, after small doses, recovery begins sooner than after large doses, the

Table

11-2

Time

Organism

Tissue

Radi-
ation

Dose,
r

Temp.

after

treat.",

°C

between

treat, and

minimal

mitotic

Reference

activity,

mm

Chick embryo

Fibroblasts
in vitro

7

50

83

300

1000

39

40

80

80

120

Spear, 1931

Mouse

Epidermis

X

5

15

25

35

325

BT

60
60
90
90
120

Knowlton et al, 1948;
Knowlton and
Hempelmann, 1949

Chortophaga

Neuroblast

7

8

64

128

256

38

66
66
88
88

Carlson, unpublished

Vicia

Root tip

7

Small
Large

25

180''
540

Gray et al., 1940
Mottram, 1936

a BT, body temperature; — , no information on temperature given.
6 See p. 775 and footnote 5.

rise in mitotic activity that marks the end of the period of mitotic fall
occurs somewhat earlier afte? small than large doses.

Prolongation of the period of mitotic decrease after doses larger than
those sufficient barely to reduce the mitotic count to zero is apparently
due to two factors: a radiation-induced retardation in the mitotic rate of
those cells that complete mitosis immediately after treatment and an
increase in the time required by cells to pass through anaphase, if the dose
is large enough to cause stickiness of the chromosomes. Jungling and
Langendorff (1930) found that in the Vicia root tip cessation of mitosis
occurred 18 hours after 420 r but 33 hours after 550 r. Increased doses of
X rays cause increased delay in the breakdown of the nuclear membrane
and increased stickiness of the chromosomes of the Chortophaga neuro-

78

RADIATION BIOLOGY

blast, both of which tend somewhat to delay the completion of mitosis
(Carlson, 1941). The amount of delay is directly related to the nearness
of the neuroblast at the time of treatment to the stage of mitotic stoppage ;
the more recently the treated cell has passed this stage, the greater is the
delay.

The interval of time elapsing between treatment and the virtual dis-
appearance of mid-mitotic stages6 is considerably greater than the average
interval of time required by the untreated cell to complete mitosis from

180

12 3 4 5 6

DURATION OF INCUBATION IN HOURS SUBSEQUENT TO IRRADIATION

Fig. 1 1-5. Mitotic effects of y rays from radium on chick fibroblasts in vitro. Curves
from top to bottom represent effects of 50, 83, 300, 1000, and 2000 r, respectively,
given at 33 r/min (after Cade, 1948, after Canti and Svear, 1929).

the stage just following that at which radiation blockage occurs. The
untreated chick fibroblast requires about 35 minutes on the average to
pass from the beginning of prophase to the formation of the daughter
nuclei (Strangeways, 1922) or about 20 minutes to complete metaphase,
anaphase, and telophase (Simon-Reuss and Spear, 1947), but it is not
until 2 hours after irradiation that mid-mitotic stages have virtually dis-
appeared (Canti and Spear, 1929). The root-tip cells of Vicia require

6 Mid-mitosis is used in this chapter to include prometaphase, metaphase, and
anaphase stages, i.e., the period between breakdown of the nuclear membrane and the
loss of the smooth form of the chromosomes after they have reached the poles.

CELL DIVISION, MORPHOLOGY, VIABILITY 779

about 2>2 hours to progress from early prophase through telophase (Gray
and Scholes, 1951), while mitosis does not fall to zero until 9-12 hours
after treatment (Mottram, 1936; Deufel, 1951). Simon-Reuss and Spear
(1947) found that as small a dose of y rays as 88 r retarded temporarily or
permanently mitotic progression in over one-half the cells treated in
metaphase, anaphase, and telophase. Neuroblast cells of the grasshopper
require on the average about 26 minutes to progress through very late
prophase, prometaphase, metaphase, and anaphase (Carlson and Hol-
laender, 1948), but after irradiation the zero level is not reached until
about 50 minutes after treatment. Recent studies carried out in our
laboratory on neuroblasts of Chortophaga seem to indicate that doses of
250 r and greater delay the completion of mitosis by retarding the dis-
solution of the nuclear membrane, by producing chromosome stickiness
and delayed separation of the chromatids at anaphase, or by retarding the
formation of the cleavage furrow (Carlson and Harrington, unpublished).
Smaller doses do not affect appreciably the rate at which cells past the
critical period complete mitosis (Carlson and Harrington, unpublished;
Wolf son, unpublished). After small doses, e.g., 32 r, up to half of the
late prophase cells may pass the block and thus contribute to this dis-
crepancy, but this does not appear to take place at 128 r. Our present
conclusion, reached from studies of the grasshopper neuroblast in vitro, is
that among both treated and control cells considerable variation exists
in the duration of very late prophase, so that, although most cells require
no more than 26 minutes to progress from the beginning of very late
prophase to the end of anaphase, enough cells require a longer time to
account for the presence of mid-mitoses 44 minutes after treatment.
These data cast doubt on the validity of using the time required for
mitosis to disappear after treatment as a measure of the average time
required for the cells to pass through the stages in question, unless a
correction is made for this variation.

Knowlton and Widner (1950) have recently proposed a method of
using X-ray-induced stoppage of mitosis to calculate the duration of the
intermitotic period. In any given tissue the ratio of the number of
mitotic to intermitotic cells should equal the ratio of the duration of
mitotic to intermitotic stages. In the equation,

No. of mitotic cells duration of mitosis

No. of intermitotic cells duration of intermitotic period

the number of mitotic and intermitotic cells is obtained by direct counts
and the duration of mitosis is determined from rate of fall of the number
of mitotic cells following treatment, The accuracy of this method
depends on the validity of certain assumptions that can be determined
only with difficulty in many kinds of biological materials; therefore,
certain information about the material and the X-ray effect must precede

780 RADIATION BIOLOGY

its application. First, the stage of blockage must not be among the
stages classed as mitotic. If any of the prophase cells that are being
included in the counts of mitotic cells are blocked by the X rays or caused
to revert mitotically (Carlson, 1940, 1941), the rate of fall in numbers of
mitotic cells after treatment will not be a true measure of the normal
mitotic rate of these cells. Second, the stage of blockage must precede
by only a short interval of time the stages included among those desig-
nated as mitotic stages. If the stage of blockage precedes by a long
interval the stage at which counts are being made, cells will enter mitosis
as fast as they leave it until the supply is depleted. By this time a certain
proportion of the blocked cells will have begun to recover and enter
mitosis, thus making it difficult to determine accurately the rate of fall of
mitosis. Third, the dose must be large enough to prevent all but an
inappreciable number of cells from leaking through the stage of blockage
or recovering early enough to swell the mitotic count in the postirradia-
tion period. Fourth, the dose must be small enough that delay is not
induced in the progress of cells through the mitotic stages in which cells
are being counted. A dose of 256 r, for example, as determined by
timing experiments on living Chortophaga neuroblasts in vitro will delay
appreciably the progress of cells through mid-mitosis (Carlson and
Harrington, unpublished). These cells were treated in very late pro-
phase, that is, just past the blockage stage. Such a delay decreases the
rate of fall of mitoses following irradiation. Fifth, the tissue must be
homogeneous with respect to the mitotic activity of its cells. If, for
example, only one-tenth of the cells in a certain tissue divide regularly,
then the intermitotic period of these cells must be one-tenth as long as
that calculated by this method, in order to account for the number of
mitotic cells present. If we were to apply the method of Knowlton and
Widner (1950) to the Chorto-phaga neuroblast, for which we have much
information on the conditions just discussed, we would proceed as follows.
Extensive series of counts of mid-mitotic cells at 22-min intervals after
treatment of neuroblasts in vitro with 64 r of X rays give average counts
of approximately 10, 7, 2, and 0 at 0, 22, 44, and 66 min after treatment,
respectively. The slope of this curve is steepest from 22 to 44 min. A
straight line with the same slope drawn through 10 on the Y axis inter-
sects the X axis at approximately 44. Substituting this value for the
duration of mid-mitosis, 10 for the average number of mid-mitotic cells,
and 190, which has been obtained by averaging direct counts, for the
number of intermitotic7 cells, we have

10 44

190 duration of intermitotic period

7 For the sake of simplifying this analysis, "intermitotic" as used in this paragraph
refers to the period from the end of one to the beginning of the next mid-mitotic period.
This includes telophase, interphase, and prophase.

CELL DIVISION, MORPHOLOGY, VIABILITY

781

The answer arrived at is 836 min, or about 14 hr for the duration of the
intermitotic period; yet we know from direct observations that under
good conditions most neuroblasts in hanging-drop preparations will go
through a complete mitotic cycle in 5-6 hr. Where is the error in this
calculation? In the first place, we know from direct timing experiments
that the average neuroblast passes through prometaphase, metaphase,
and anaphase, in 22 min while the 44 used above is approximately the
time taken by the slowest cells. If we use 22 instead of 44 in the above
equation, we arrive at an intermitotic time of about 7 hr, which is still
considerably higher than we observe. Probably this is not far from a
correct average figure when we take into account the fact that occasional
neuroblasts spend an excessively long time in interphase or early pro-

lOO-i

en
</>
o

<

s

ct
o

z

fesoH

z
u

o

tc.

UJ

a

o4-o 1 A — I 1 1 1

100 200 300 400 500

DOSE, r

Fig. 11-6. Relation of dose of radiation to maximum depression of mitotic activity.

Solid circle, chick fibroblast (Spear and Grimmet, 1933) ; triangle, cell of developing rat

retina (Ta7isley, Spear, and Gliicksmann, 1937); open circle, Chortophaga neuroblast.

phase, and so are often disregarded in the calculations as "abnormal"
cells.

The efficiencies of different doses of radiation in decreasing mitotic
activity — as measured at the time after treatment when the effect is
greatest — are quite different for different kinds of cells. The mitotic
activity of Chortophaga neuroblasts is reduced to a much greater degree
than that of either rat retinal cells or chick fibroblasts by a given dose of
radiation (Fig. 11-6). A comparison of these efficiencies with those
obtained when testing the capacity of radiation to retard the mitotic
progress of the same cells treated in mid-mitosis points to the fallacy of
concluding that the relative radiosensitivities of different kinds of cells as
measured by one effect are necessarily correlated with their radiosensitiv-
ities as determined by another effect. A dose of 88 r, for example, which
reduces the mid-mitotic count of the neuroblast to zero and that of the
fibroblast to 20 per cent of normal, has no detectable effect on the mitotic

782 RADIATION BIOLOGY

progress of neuroblasts treated in metaphase, anaphase, or telophase, but
retards considerably over half the fibroblasts treated in these stages.

After doses of radiation, up to and including those barely sufficient to
reduce the mid-mitotic count to zero, recovery from the radiation effect is
accompanied by a rise of the mid-mitotic count at a rate roughly similar
to the rate of fall in the count immediately after treatment, with no pro-
longed low count intervening (Fig. 11-5) (Canti and Spear, 1929; Kemp
and Juul, 1930; Spear, 1931, 1932, 1935; Wilson et al, 1935; Mottram,
1936; Tansley et al, 1937; Lasnitski, 1940; Carlson, 1942; Simon-Reuss
and Spear, 1947; Knowlton, Hempelmann, and Hoffman, 1948; Knowl-
ton and Hempelmann, 1949; Deufel, 1951). This rise in the mid-mitotic
count does not stop at the normal level but continues above it before
returning to the normal level. Following very small doses, the rise above
the normal after recovery compensates approximately for the fall below
normal that precedes it (Fig. 11-5) (Canti and Spear, 1929; Spear, 1931,
1932, 1935; Wilson et al., 1935; Tansley et al, 1937; Knowlton and
Hempelmann, 1949; Carlson, Snyder, and Hollaender, 1949). From
what we know of the behavior of individual cells after treatment, this is
exactly what we would expect. In the grasshopper neuroblast, for
example, blockage from irradiation affects not only the cells at the most
sensitive stages (namely, middle and late prophase) but also cells entering
these after treatment in earlier stages, so that an excessively large number
of cells accumulates during the period of decreased mitotic activity. A
similar increase has been found by Mallet and Perrot (1951) in the
Allium root tip. With recovery and the resumption of mitotic progres-
sion, these cells will complete mitosis and their numbers should com-
pensate exactly for the previous deficit. With larger doses, however, the
number of mitoses never exceeds the normal and with still larger doses the
return to normal may be greatly prolonged (Alberti and Politzer, 1924;
Pekarek, 1927; Canti and Spear, 1929; Kemp and Juul, 1930; Spear,
1931, 1935; Tansley et al, 1937; Lasnitski, 1946, 1948; Simon-Reuss and
Spear, 1947; Knowlton et al., 1948; Knowlton and Hempelmann, 1949;
Carlson et al., 1949). Failure of the mid-mitotic count to surpass the
normal is apparently due to the extension of the radiation effect to earlier
and earlier stages of the mitotic cycle as the dose is progressively
increased, until cells in all stages of the cycle are retarded.

If the dose is large enough to lower the mid-mitotic count to zero, the
larger the dose, the more prolonged is the period of zero activity. Few
studies have been made of the relation of the length of this period to
dosage, probably because, for most purposes, the investigator has found it
more profitable to work with doses that do not completely inhibit mitosis.
A dose of 2000 r of y rays reduced the mitotic count of chick fibroblasts
in vitro to zero until about 5 hours after treatment (Fig. 11-5). Jungling
and Langendorff (1930) reported that in the Vicia root tip mitosis

CELL DIVISION, MORPHOLOGY, VIABILITY

783

remained at zero for about 16, 18, and 24 hours after doses of 175, 420,
and 550 r, respectively. In the Chortophaga neuroblast the time from
irradiation to the reappearance of the first metaphases after mitotic
cessation approximates the curves shown in Fig. 11-7 for 26 and 38°C.
None of the tissues of Amby stoma larvae exposed to 15,000 r of X rays
exhibited any mitoses one week later (Rugh, 1949).

lO.OOO-i

1,000-

UJ

O
o

100-

10-

1^
10

I
20

30

HOURS BETWEEN BEGINNING OF X IRRADIATION AND REAPPEARANCE

OF MITOSIS

Fig. 11-7. Graph showing the relation between dose and the time after the beginning
of X-ray treatment when mitoses reappear. Postirradiation temperature, 26° and
38°C; material, Chortophaga neuroblast.

In contrast to the foregoing investigations, which point only to depres-
sion of mitotic activity as a result of treatment with ionizing radiations,
a few studies have indicated that certain exposures may actually speed
up the mitotic rate. Richards (1915) reached the conclusion that
exposure of the fertilized egg of the snail, Planorbis, to X rays during the
spindle formation period of the maturation or early cleavage divisions
stimulated the cell to complete its division much more rapidly than if
untreated; one experiment, for example, showed a change from 75 to
3 minutes for the time of the first cleavage division. The next succeeding
division was usually faster than normal, but following divisions showed a
progressive retardation. If the exposure was made, therefore, during the

784 RADIATION BIOLOGY

first cleavage division, development of the treated eggs would usually be
behind that of the untreated eggs at the 24-celled stage. No stimulation
and probably some depression of the division rate was produced by treat-
ment of the resting stage. Packard (1916) reported a 7-ray-induced
acceleration of the first cleavage division of Arbacia that amounted to a
5-15 per cent increase in the speed of division. Richards and Good
(1919) found that a small dose of X rays applied to the fertilized egg of
Cumingia stimulated division at first, and then retarded it. Irradiation
of sperm or eggs or both before fertilization, however, produced either no
effect or a retardation of subsequent cleavage divisions. More recently,
Darlington and La Cour (1945) found increases in the frequencies of
metaphases and anaphases of Vicia root tips 4 hours at 24°C after treat-
ment with 45 r of X rays. Since the numbers of cells in these stages
exhibited decreases at other doses (90 and 135 r) at the same temperature
and at all three doses at 16 and 30°C, they conclude that there is an
optimum temperature and dose at which mitosis may be temporarily
stimulated by X rays. The numbers of metaphases and anaphases in
the controls for their different experiments differ widely, however, and
they give no tests of significance for these numbers. La Cour (1951) also
reports an X-ray-induced acceleration of mitosis by 90 r at 20°C but does
not include the data on which this conclusion is based. In view of the
fact that, except for these studies, no other researchers seem to have
found evidence of a stimulating effect of radiation, confirmation by other
workers, duplicating as exactly as possible the procedures originally
used, would seem worth while. An attempt to confirm the earlier studies
was made by Seide (1925), who used a different biological material, how-
ever. He treated the Ascaris egg in the pronuclear stage with small doses
of X and 7 radiation, but was unable to demonstrate a constant increase
in the mitotic rate as determined by the time required for the treated
cells, as compared with the controls, to complete the first and the second
divisions.

Evidence relating to the means by which ionizing radiations interfere
with mitosis has pointed strongly to a primary effect on the nucleus.
The first cleavage division of the Arbacia zygote is delayed to about the
same extent, whether the sperm or the egg is irradiated (Henshaw and
Francis, 1936; Henshaw, 1940b; Henshaw and Cohen, 1940). Since the
sperm consists almost entirely of nuclear material, it is natural to con-
clude that the X-ray effect must involve changes induced in the nucleus
rather than in the cytoplasm. Further, if nucleated and nonnucleated
egg fragments obtained by centrifugation are irradiated and then
fertilized with untreated sperm, it is found that a delay is produced in
the nucleated fragments comparable to that produced in the whole egg,
while no delay is produced in the nonnucleated fragments (Henshaw,
1938).

CELL DIVISION, MORPHOLOGY, VIABILITY 785

Miwa, Yamashita, and Mori (1939b), on the other hand, obtained
evidence that mitotic delay may be induced by treatment of the cytosome.
Unfertilized eggs of the sea urchin, Pseudocentrotus, were irradiated with a
rays through aluminum filters varying from 0 to 45 n in thickness.
Sensitivity of the eggs as measured by delay in the first cleavage division
decreased markedly as the increased thickness of the filters used pre-
vented the a rays from penetrating to the nucleus, but even with a-ray
ranges that fell short of the nucleus, some eggs showed marked delay.

It has also been demonstrated that the amount of cleavage delay
resulting from X-raying Arbacia sperm is not influenced by the concentra-
tion of the sperm or the composition of the medium in which they are
irradiated (Evans et al, 1942). When Arbacia sperm were mixed with
heavily irradiated water containing X-ray-induced hydrogen peroxide,
and then used to fertilize untreated eggs, delay of the first cleavage divi-
sion resulted (Evans, 1947). The amount of delay was positively cor-
related with the dose. Heilbrunn and Young (1935), however, found
that, while the concentration of unfertilized Arbacia eggs X-rayed in sea
water had no effect on the delay of the first cleavage division, irradiation
in the presence of ovarian tissue produced a considerably greater delay
than irradiation of the eggs in sea water alone or in concentrated suspen-
sion. They suggest that ovarian cells exposed to X rays produce a sub-
stance or substances that act on the eggs to enhance the direct effect
obtained in their absence.

Increased absorption of 2537 A ultraviolet radiation in the cytoplasm
of irradiated neoplastic cells is interpreted by Mitchell (1942a, b, 1943),
to result from radiation-induced inhibition of desoxyribonucleic acid
(DNA) formation and the consequent accumulation of pentose nucleo-
tides in the cytoplasm. He suggests that this might cause inhibition of
mitosis.

That the cytosome, however, may be responsible for recovery from
radiation effects is suggested by the findings of Henshaw (1940a) using
X rays, and Miwa et al. (1939a) and Mori, Miwa, and Yamashita (1939)
using /3 rays, who state that there is no recovery in sea urchin sperm
between treatment and insemination. Since the same investigators have
found that sea urchin eggs undergo recovery between treatment and
insemination, it seems likely that the cytosome of the egg is responsible
for recovery.

Kind of Radiation. Of special importance in the interpretation of the
mechanism of the action by which radiations are able to alter the rate of
so complex and yet so precise a living process as cell division are studies of
the relative capacities of different kinds of radiations to interfere with
mitosis. From the point of view of the biologist, different radiations are
of interest because of their different ion distributions or densities. These
range from the widely separated ions produced by high energy /3 and y

786 RADIATION BIOLOGY

radiation (about 8.5 ions//* in tissue), the denser ionizations of medium
X rays (about 100 ions//z), the still more densely ionizing recoil protons
from fast neutron irradiation (up to 1100 ions//x), to a particles with
densities of 3700-4500 ions/M (Gray, 1946, 1947). Knowledge of the
relative efficiencies of the different radiations in interfering with mitosis
might give us a clue to the means by which radiation energy is trans-
formed into observable effects on cell morphology and behavior. Since
such determinations depend directly on the accuracy with which the dose
absorbed by the cells is measured, conclusions reached from comparative
studies of this type are valid only insofar as the energy measurements are
accurate. As might be expected, only quantitative, not qualitative,
differences in the effects produced by different radiations have been
demonstrated.

Fast neutrons have been found to be more effective than y rays in
reducing the mitotic activity of a number of different kinds of cells.
Zirkle, Aebersold, and Dempster (1937), who compared the efficiencies of
neutrons and X rays by irradiating fern spores, germinating them, and
determining the proportion that had undergone their first cell division at
the end of 10 days (normally this division occurs on the sixth day of
germination), found that neutrons were 2.5 times as effective as X rays in
reducing division by 50 per cent. Small doses of neutrons were found by
Gray et at. (1940) to be 2.1 times as effective as 7 rays in reducing the
number of mitoses in broad bean root tips 3 hours after the mid-point
of the treatment period. Similar experiments carried out with chick
fibroblasts in culture also showed neutrons to be more effective than
7 rays, as determined by the reduction in mitotic activity 80 minutes
after irradiation. When the percentage of normal mitoses was plotted
against dose, the neutron curves were exponential and the 7-ray curves
sigmoid, the two intersecting at some dose higher than any used in the
study. They postulated that the neutron effect may result from the
passage of a single recoil proton within a certain limited volume of radius
2 or 3 (j. in the cell, and that the 7 radiation effect would require some two
hundred secondary /3 particles.

The evidence with regard to the effectiveness of a particles in inhibiting
mitosis, however, is conflicting. Gray and Read (1950) found a rays to
be less effective than either neutrons or 7 rays in lowering the mitotic
count 3 hours after the mid-point of treatment in the broad bean root tip
(Fig. 11-8). The relative efficiencies of 7 rays, neutrons, and a rays were
about 1:2.1:0.6. They point out that these relative efficiencies of neu-
trons and a rays are in good agreement with the ion density ratios of
recoil protons to a rays, if reasonable assumptions be made for the 5-ray
ionization. On the other hand, in the frog tadpole treated with a radia-
tion by immersion of the animal in water containing radon and subse-
quently sectioned and stained at intervals for study, a particles proved to

CELL DIVISION, MORPHOLOGY, VIABILITY

787

be twice as effective as 7 rays (Spear and Glucksmann, 1938; Tansley
et al., 1948). A dose of 134 energy units of a radiation reduced the
numbers of eye and brain cells in mitosis, to approximately the same
level as 268 r of 7 rays (tissue exposed to 1 energy unit has received the
same increment of energy/g as water exposed to 1 r of 7 rays).

Miwa et al. (1939b) observed in Pseudocentrotus that after ft irradiation
the cleavage delay of all the treated eggs was about the same (with a
spread of 30 minutes), while after a irradiation the spread was much
greater, some eggs being delayed hardly at all and others 3 or more

2.0-

X

UJ
Q

o

h-
o

h-

o
o

_)

10

NEUTRON

0 100 200

DOSE, r or eu

Fig. 11-8. Relative efficiencies of a rays (solid circle), 7 rays (open circle), and fast
neutrons (square) in reducing the mitotic index in Vicia root tip meristems 3 hours
after irradiation (after Gray and Reed, 1950).

hours. Because their study did not include careful measurements of the
doses used, it is difficult to relate these results to those of other a-ray
studies.

Lasnitski and Lea (1940) compared the efficiencies of 7 rays with
medium X rays, and hard X rays with soft X rays in reducing the pro-
phase, metaphase, and anaphase-telophase counts in chick fibroblasts
in vitro. For several doses within the range of 20-200 r, the hard and
soft X rays were found to be equally effective, but the 7 rays were found
to be less effective than medium X rays by a factor of about 2:1. These
conclusions are based on the assumption that the low point is reached a'
80 minutes after each kind of radiation, when all counts were made; if it is
not, the different values may be measures of the rapidity at which the
mitotic count falls or at which recovery takes place.

788 RADIATION BIOLOGY

Lasnitski (1948) determined the relative efficiencies of /? and X rays
in lowering the mitotic count in the chick fibroblast culture. She found
that, although the over-all depression of mitotic activity during the
24-hour period following treatment and the maximum reduction of
mitosis was comparable for the two radiations, mitotic activity fell off
more abruptly and during recovery rose more rapidly in the /?- than in the
X-irradiated material (Fig. 11-15). This illustrates very clearly the
danger of interpreting the extent of radiation-induced mitotic depression
on the basis of mitotic counts made at the end of a single time interval
after treatment.

Dosage Rate. In the great majority of studies in the field of radiation
biology treatment administered is expressed in terms of dose, with little
emphasis placed on the time-intensity relationship. The dose, however,
represents only the total quantity of radiant energy dissipated in a tissue.
It tells us nothing of the intensity of the radiation or of the time of
exposure, and these may be quite important in determining the effect
produced. Excellent discussions of these factors in biological research
are presented by Lea (1938a, 1946) and Gray et at. (1944).

The time-intensity factor is important, and at the same time peculiarly
difficult to deal wTith experimentally for several reasons. First, if
recovery of the cell from radiation effects occurs at all, it begins immedi-
ately after the first effects are produced and continues during treatment
(Canti and Donaldson, 1926; Henshaw et al., 1933); therefore the bio-
logical effect as determined at the end of a long treatment period, during
which recovery from the earlier effects produced has occurred, will be
less than that obtained after a corresponding dose delivered at high
intensity in a shorter period. Second, a certain detectable biological
effect may depend on interaction between the products of two or more
primary effects that are subject to recovery during the treatment period—
perhaps at the molecular level or perhaps at the microscopic level, e.g.,
broken chromosomes which by interaction (fusion) give rise to transloca-
tions. If these are subject to recovery during the treatment period,
small doses will produce little or no effect, whether delivered in a short
period at high intensity or in a long period at low intensity, because the
primary changes will be so few and far between on the average that little
or no interaction will take place. At successively larger doses, however,
interaction will have a successively better opportunity of occurring and
the observed effect will increase theoretically as the square of the dose-
actually it is usually less than this because of the recovery that takes
place during treatment. Third, since the living cell is dynamic and not
static and since visible biological effects cannot be detected immediately
after treatment but only at some later phase of a cell's physiological and
morphological state, and then may be manifest maximally for only a brief
period, results will often depend on the time after treatment at which the

CELL DIVISION, MORPHOLOGY, VIABILITY 789

determination is made. As the treatment becomes lower in intensity
and more prolonged in time, the choice of the appropriate time at which
the effect is to be determined becomes more and more difficult. Fourth,
the cell and its parts undergo a whole series of cyclic changes in their
physical and chemical nature during mitosis; treatment extending over a
long period of time will increase the chances of a certain highly sensitive
stage in the mitotic cycle receiving radiation, while a brief treatment will
decrease this possibility. The proportion of a given quantity of radia-
tion that a cell receives in a particularly sensitive or insensitive stage,
therefore, should not be overlooked in the interpretation of dosage-rate
effects. Though recovery processes in the cell usually begin as soon as
the first effect of the radiation is produced, damage to the cell during the
treatment period — unless the dosage rate is very low — will occur at a
faster rate than repair, and the maximum residual effect will be present at
the end of the radiation period. From this time on recovery will take
place gradually until the capacity of the cell to progress mitotically has
been restored to its original state. If, on the other hand, the dosage rate
is extremely low, e.g., about 0.8 r/hour in treatment of the grasshopper
neuroblast with 7 rays, an equilibrium between radiation damage and
tissue repair will be established soon after the start of treatment, the
residual effect remaining relatively constant over a long portion of
the treatment period (Carlson and Harrington, 1953). Not only is it
essential that these factors be taken into account in the interpretation
of radiation results, but they may also be utilized in designing experi-
ments to test certain hypotheses relating to the effects of radiations on
living material.

The interpretation of the results of studies of the effects of different
dosage rates on mitosis in selected cells that were in known stages of
mitosis at the time of treatment and that can be observed at desired
intervals after treatment, e.g., marine invertebrate eggs and hanging-drop
preparations of grasshopper neuroblasts, offers no particular difficulties,
if treatment is not so prolonged that deterioration sets in during the
course of the experiments. The Arbacia sperm and egg offer, in fact, a
unique opportunity for time-intensity studies. As described previously,
Henshaw (1932) found that recovery of eggs occurred between treatment
and cleavage. Further, Henshaw et at. (1933) found that recovery took
place between the beginning and end of irradiation. If Arbacia eggs
were exposed to a given dose of X rays administered at different rates,
the cleavage delay produced by the lower dosage rate was 20-40 per cent
less than a dosage rate eight times as high and therefore given in one-
eighth the time. On the other hand, the sperm gives no evidence of
recovery before insemination (Miwa et at., 1939a; Henshaw, 1940a).
These results would lead us to expect that the Arbacia egg would exhibit a
dosage-rate effect, while the sperm would not; and such, indeed, is the

790 RADIATION BIOLOGY

case. Using dosage rates of 120-960 r/minute, Henshaw and Francis
(1936) found that X irradiation of the sperm was more effective than X
irradiation of the egg in producing mitotic delay in the z}rgote. On the
other hand, a dose given at 7800 r/minute produces greater mitotic delay
when administered to the egg than to the sperm (Henshaw, 1940b; Hen-
shaw and Cohen, 1940). Apparently the initial effect of irradiation is
greater in eggs than in sperms, but, owing to recovery during the treatment
period, the residual effect at the end of treatment will be less in the egg
than in the sperm, if the treatment period is prolonged.

Accurate comparisons of the effects of different dosage rates on tissues,
however, in which the results are based on counts of cells in different
mitotic stages at certain intervals of time following treatment, introduce
special difficulties. We may consider separately the problems presented
by relatively short and relatively long exposures.

If the times of irradiation are short, i.e., occupying only a small part of
the period during which the number of mitoses is falling and therefore a
small portion of the duration of one mitotic cycle, the minimum levels of
mitotic activity reached following treatment can be compared to deter-
mine the effectiveness. In such experiments, where a single count of
certain mitotic stages following irradiation is used as a measure of effec-
tiveness, the results will be different depending on whether the mitotic
count is made a certain number of minutes after the beginning, the mid-
point, or the end of the treatment period. Probably timing from the mid-
point of the treatment period is the most reliable procedure, when com-
parisons of the effects of different exposure times are to be made, unless
the time of the low-intensity treatment is greatly prolonged. The results
of the 7-ray studies on dosage-rate effect by Canti and Spear (1927) and
Spear and Grimmett (1933) on chick fibroblast in vitro are based on
counts of the numbers of cells in mitosis in treated cultures fixed 80 min-
utes after the end of the irradiation period and expressed as the percentage
of the number of mitoses in the controls. Both studies indicated that at
dosage rates of approximately 20 r/minute and higher, the dose required
to produce a given reduction in mitosis — to 50 per cent of that in the
controls in the former study and to 40 per cent in the latter — was inde-
pendent of the dosage rate. At rates below 20 r/minute, however, Canti
and Spear found that the dose increased successively with successively
lower dosage rates, while Spear and Grimmett's more detailed study in
the 4-20 r/minute region shows a fall in dosage from 17.2 to 8.7 r/minute
and a rise from 8.7 to 4.3 r/minute. This apparent demonstration of an
optimum dosage rate below or above which the mitotic depression is less
might be due to the fact that all their results were based on counts made
on cultures fixed 80 minutes after the end of the treatment period, while
the suppression of mitosis by irradiation is initiated at the beginning of
irradiation. At the time the counts were made, therefore, all the primary

CELL DIVISION, MORPHOLOGY, VIABILITY 791

effects produced by the high dosage-rate irradiation would have had less
time for recovery than all except the last effects produced by the low
dosage-rate treatment. An optimum effectiveness would then be
expected for the dosage rate for which a minimum of recovery had
occurred at 80 minutes after the end of treatment.

A more laborious but at the same time more informative procedure is to
make counts at intervals following irradiation. Comparison can then be
made of the rates of fall and rise, as well as the minimums of mitotic
activity at certain times after the beginning, mid-point, or end of the
irradiation period.

Using a dose of 100 r delivered at rates of 9.3, 29.8, and 103 r/minute,
Lasnitski (1946) determined that depression of mitotic activity was
approximately the same 80 minutes after treatment for all three dosage
rates. Recovery occurred more rapidly after 103 r/minute than after
29.8 r/minute and more rapidly after 29.8 r/minute than after 9.3 r/min-
ute. After 2500 r, however, no mitotic cells were present within 24 hours
if the dose was delivered at 101 r/minute, 0.4 per cent of the cells were in
mitosis at 24 hours if the rate was 29.8 r/minute, and about 1 per cent if
the rate was 9.3 r/minute. (The per cent of mitotic cells in the control
cultures at any one time during the same period ranged from 2.6 to 8.0.)
Counts of the mid-mitotic cells in hanging-drop preparations of grass-
hopper neuroblasts give comparable results. Curves based on twenty
successive counts of ten to fourteen embryos made at 22-minute intervals
beginning 20 minutes after the mid-point of the radiation period were not
significantly different for doses of 8 and 64 r at high (32 r/min) and low
(2 r/min) dosage-rate y radiation (Carlson et at., 1949). After doses of
128 and 256 r the rate of fall in mitotic activity and the length of the
cessation period are comparable for both the 2 and 32 r/minute dosage
rates, but the return of the mitotic count to normal is much slower after
the 32 than after 2 r/minute treatment (Fig. 11-9).

Another method of comparing the efficiencies of different dosage rates
in reducing the mitotic activity of a tissue is to compare the totals of the
mitotic counts obtained at uniform intervals over a certain period follow-
ing treatment. The time interval at which counts are made may be
fixed arbitrarily, but, if the interval taken represents the average time
required for the cells to pass through the mitotic stage or stages to be
counted, the total of the average counts will approximate closely the
number of cells passing through the stages counted and therefore through
mitosis during the period under consideration. This method presup-
poses, however, that the stages counted are not prolonged to such an
extent by either of the treatments that the same cell is counted twice;
otherwise a high count and, therefore, a false indication of increased
mitotic activity will result when mitotic retardation actually occurs.
This procedure can be used with confidence, then, only when it is possible

792

RADIATION BIOLOGY

to check the mitotic time schedule by direct observations of living cells.
Carlson et al. (1949), using this method, showed that the 7-ray-induced
reduction in prometaphase-through-anaphase stages during a 7-hour
period following treatment was not appreciably different after dosage
rates of 2 and 32 r/minute at doses of 8, 32, and 64 r, but that the 32
r/minute rate caused a much greater reduction than the 2 r/minute rate
at 128 and 256 r.

The problem becomes quite different if the dosage is spread over a long
period, e.g., hours or days. In such experiments the recovery factor

1 3 5

HOURS AFTER IRRADIATION

Fig. 11-9. Effects of different doses of 7 rays delivered at different dosage rates on
mitosis in the Chortophaga neuroblast: open circle, 2 r/min; solid circle, 32 r/min (after
Carlson, Snyder, arid Hollaender, 1949).

dominates the picture. A brief high-intensity treatment will be com-
pleted before the mitotic activity will have had time to fall off appreciably,
while at the end of a prolonged low-intensity treatment, radiation effect
and recovery processes will have reached equilibrium and the mitotic
activity will be at a minimum ; therefore the shape of the mitotic activity
curves, based on a series of counts immediately following treatment, will
be so different as not to be readily comparable. If the mitotic activities
after a certain dose delivered over a short and over a long period are com-
pared at corresponding times measured from the beginning or mid-point of
the radiation period, all the primary effects produced by the high-inten-

CELL DIVISION, MORPHOLOGY, VIABILITY

793

sity treatment will have had approximately the same long period for
recovery, while only the first of the primary effects produced by low-
intensity treatment will have had a correspondingly long period for
recovery. The cells given a brief, high-intensity treatment may have
recovered completely, while those subjected to the prolonged, low-
intensity treatment, even though they received the same dose, will
exhibit a decrease in the number of cells in mitosis. Probably the most
accurate method of comparing the mitotic depression after short and
very long exposures to the same dose is to obtain a series of mitotic
counts at short time intervals for a few hours immediately following each
treatment. The mitotic picture during the long exposure can be obtained
by running other material at the same dosage rate and making a single
count immediately after any desired length of exposure to determine the

0 2 4 6 8 10 12 14 16

HOURS AFTER IRRADIATION

Fig. 11-10. Relation of mitotic rate to time after y irradiation for different doses given

at 0.25 r/min; Choriophaga neuroblasts at 38°C: open circle, 8 r; solid circle, 32 r;

divided circle, 128 r (from Carlson, 1950).

mitotic activity at that time. A curve illustrating the mitotic picture
from the beginning to several hours after the termination of a prolonged,
low-intensity treatment can thus be obtained. It can be seen, for
example, that the 128-r curve in Fig. 11-10, if projected to the left, would
parallel closely the zero ordinate to a point to the left of the first 32-r
count and then upward through the first 8-r count.

Fractionated Treatment. A given dose of radiation may be delivered
not only in different periods of time and at different dosage rates but also
in two or more portions with intervening nonirradiation intervals.
Fractionated or spaced irradiation resembles low-intensity treatment in
that it reduces the possibility of interaction between primary effects by
giving time for recovery from one group of effects before the next group
is produced. Because mitosis is a cyclical process, however, with some
phases of the cycle more highly radiosensitive than others, the interpreta-
tion of the effects of fractionated treatment must take into account not

794 RADIATION BIOLOGY

only the reduced opportunity for interaction of effects, but also the
mitotic stages of the cells at the time of treatment and at observation and
the relation of these to the radiosensitivity of the stages involved.

Spear (1932) compared the effects of 2 3^- and 5-minute continuous
exposures of fibroblast cultures to a given radium source with two 2}4~
minute exposures, separated in the one case by 80 minutes and in the
other by 160 minutes. In the former, the second dose of y rays was
delivered at the depth of depression of mitotic activity, which was
lowered still further. The mitotic activity was thus lowered to a greater
extent and for a longer time after the two 2^-minute exposures than
after the continuous 5-minute exposure. In the latter experiment the
second fraction was given at the time the mitotic activity had returned
to normal, and the curve representing the resulting change in mitotic
count closely resembled the first one. These experiments suggest that
there is an optimum dose that is more effective per r than other doses in
reducing mitotic activity and that the 2^-minute exposure represents a
dose nearer to the optimum than the 5-minute exposure. This is sub-
stantiated by a comparison of the 2)4- and 5-minute continuous exposure
curves. The former shows a minimum mitotic count only about 7 per
cent lower and a return of mitotic activity to normal only about 20
minutes later than the latter.

Temperature. It appears that the temperature at which cells are
irradiated has little or no effect on their subsequent mitotic activity
(Gorman S. Hill, unpublished). Grasshopper embryos were given 8 r of
X rays while at temperatures of 2, 18, and 38°C, maintained subse-
quently at 38°C, and fixed at 44, 88, 132, 176, and 220 minutes. There
were no significant differences in the number of neuroblasts in prometa-
phase, metaphase, and anaphase at any of these time intervals for the
three temperatures tested.

The times after treatment at which the mitotic activity will reach a
minimum and then will begin to rise during recovery are functions of the
temperature at which the cells are maintained after irradiation. Main-
tenance of the cells at a low temperature after irradiation prolongs the
period of mitotic decrease and delays the return of mitotic activity. In
the Allium root tip, the time of minimum mitotic activity after 150 r is
about 10 hours at 24° and about 24 hours at 16°C (Darlington and
La Cour, 1945). In the grasshopper neuroblast, the low point of mitotic
activity is reached at about 66 minutes after 32 r of y rays at 38°C
(Carlson et al., 1949) and at about 100 minutes after 31 r of X rays at
26°C (Carlson, 1942) ; the time intervals between treatment and resump-
tion of mitotic activity are about 165 and 195 minutes at 38 and 26°C,
respectively.

Low temperature following X irradiation also delays both the fall in
the mitotic count immediately after treatment and its rise during recov-
ery. Walter G. Adams (unpublished) has studied the effect of low

CELL DIVISION, MORPHOLOGY, VIABILITY

795

t.ol (°>

4 0-

20

*

*

i

i

\

44

88

132

176

286 330

MINUTES AFTER IRRADIATION

Fig. 11-11. Per cent of Chortophaga neuroblasts in metaphase at different times after
32 r of X rays. These cells were kept at 38°C except for different intervals after
irradiation. The periods during which the cells were at 1°C are not included in the
graph. A, Control, B, 1°C from 57 to 132 minutes after X irradiation; C, 1°C from 1
to 66 minutes after X irradiation; D, 1°C from 1 to 220 minutes after X irradiation
(after Adams, unpublished).

temperature after treatment on X-ray-induced changes in mitotic activity
in the grasshopper neuroblast. He found that exposure to 3°C for 220
minutes immediately after irradiation (32 r), followed by incubation at
38°C, delayed the fall to zero in the metaphase count and the subsequent
return of metaphases, as compared with cells kept at 38°C from treatment
on, by only slightly less than the time of the cold treatment (Fig. 11-11).
If the embryos were changed from 38 to 3°C at the beginning of the period

796 RADIATION BIOLOGY

when the metaphase count reached zero, i.e., at 66 minutes after X
irradiation, and returned to 38°C after 66 minutes, the reappearance of
metaphases was delayed by almost the time of the cold treatment, as
compared with cells kept at 38°C throughout. Exposure to 3°C, there-
fore, brings almost to a standstill both mitotic progress and recovery.
Henshaw (1940c) found that Arbacia eggs, when kept at 0°C from treat-
ment to insemination, showed less recovery and, consequently, greater
cleavage delay than those kept at 24°C during this period.

Chemical Agents. Immersion of unfertilized Arbacia eggs in 0.35 M
potassium citrate to remove most of the calcium from the cell during
X-raying was found by Wilbur and Recknagel (1943) to decrease the
cleavage delay normally caused by irradiation. The effect, however, was
slight and occurred mainly after large doses (30,400 and 53,200 r).
Increased calcium and magnesium content of the sea water during X
irradiation had no effect on cleavage delay.

Using chick fibroblast cultures, Paterson and Thompson (1949) found
that urethane, which in a concentration of 0.2 per cent inhibits mitosis,
when added to the culture medium in a concentration of 0.1 per cent just
before or up to 40 minutes after X-raying, reduced the mitosis-inhibiting
effect of the X rays. They draw the conclusion that the capacity of
urethane to reduce the X-ray effect when added after irradiation indicates
that X-ray action is incomplete at the end of irradiation. The method
used in determining the degree of metotic inhibition induced and the
data on which the conclusion is based are omitted from this preliminary
report, so it is not possible to evaluate their interpretation.

pH. Zirkle (1936) found that the degree of acidity or alkalinity of
the medium in which fern spores were X-rayed affected their sensitivity,
as determined by the proportion in which the first cell division had
occurred by the tenth day after treatment — it normally occurs on the
sixth day. In 0.006 M carbon dioxide in culture medium the radio-
sensitivity was greatest, being less in stronger or weaker concentrations of
carbon dioxide. Minimum radiosensitivity was exhibited in 0.003 M
ammonia; it was greater in either stronger or weaker concentrations of
ammonia.

Oxygen Concentration. A series of studies has recently been made by
Gaulden, Nix, and Moshman (1953) on the mitotic effects of exposing the
grasshopper egg during X irradiation to different concentrations of
oxygen. In concentrations of 0, 2, 5, 10, 21, and 100 per cent oxygen,
64 r of X rays reduced the number of prometaphase, metaphase, and
anaphase cells to zero about 1 hour after treatment. The greater the
per cent of oxygen, however, the longer these stages remained at zero and
the longer the time interval between irradiation and the peak of mitotic
activity following recovery. Treatment with 8 r of X rays, however,
which does not reduce the mitotic activity to zero, resulted in no sig-

CELL DIVISION, MORPHOLOGY, VIABILITY

797

nificant difference in the extent to which mitosis was depressed, whether
irradiation was carried out in vacuum or in 100 per cent oxygen. It
seems, therefore, that the concentration of oxygen at the time of treat-
ment has no effect on the degree to which mitotic activity is reduced but
does determine the rate of mitotic recovery. The results were the same
whether 100 per cent carbon dioxide, 100 per cent nitrogen, or a vacuum
was used ; therefore it was clear that the oxygen concentration of the gas
mixtures determined the effectiveness of the X rays in these experiments.
Type of Preparation and Kind of Tissues. The effects on mitosis of y
irradiation of pre- and postcirculatory chick embryos have been com-

HOURS AFTER IRRADIATION

Fig. 11-12. Comparison of effects of approximately 130 roof y rays from radium on
mitosis in chick embryo tissues differing in age. Precirculatory chick designated by
broken line; postcirculatory chick by solid line (reconstructed from Spear, 1935; Wilson
el al., 1935).

pared with the effects produced by comparable doses in the chick fibro-
blast culture preparation (Wilson et al., 1935; Spear, 1935). The mitotic
count of the embryo with blood circulation established fell to a minimum
in about the same length of time as the culture preparation, but recovery
was earlier and more abrupt (Fig. 11-12). In the precirculatory embryo,
mitosis reached a minimum sooner and recovery was greatly delayed. A
comparison of the effects of y radiation on malignant cells of the mouse
in vitro and in vivo showed no difference in the time required for the
mitotic activity to reach a minimum (Lasnitski, 1945). Mitotic activity
recovered, however, more rapidly in vivo than in vitro.

The postirradiation picture of mitotic activity in a number of different
tissues in the mouse has been determined by Knowlton and Hempelmann

798 RADIATION BIOLOGY

(1949). No striking differences in the relation of dose to rate of decrease
of mitotic activity after X radiation are evident. Only the adrenal gland
shows a typical compensatory effect a few hours after treatment; epi-
dermis, jejunum, and lymph nodes give little or no indication of a rise of
mitotic activity above normal. A greatly delayed overshooting does
occur 3 and 7 days after treatment in the epidermis and lymph nodes,
respectively, but, as the authors point out, this may be due to secondary
physiological factors.

Chromosome Number. Marshak and Bradley (1944) found that the
inhibition of mitotic activity in root tips of three species of Triticum and
two species of Bromus with diploid chromosome numbers of 14, 28, 42, 56,
and 84, as determined by mitotic counts made 3 hours after X irradiation,
was neither directly nor inversely proportional to chromosome length but
did vary inversely as the chromosome number. Evidence from other
studies on plant root tips suggests that mitotic activity has not reached a
minimum value at 3 hours. If that is true in these species these results
might be a measure of the rate of fall of mitotic activity rather than of
minimum activity, and rate of fall is mainly determined by the normal
mitotic rate (see p. 775). The inverse relation to chromosome number
could be due, therefore, as well to a lower mitotic rate in cells with more
chromosomes as to increased resistance to radiation-induced mitotic
delay in cells with the larger chromosome number.

MORPHOLOGICAL EFFECTS

Nuclear Components. Effects of ionizing radiations on chromosomes
fall into two main classes: those referred to as "primary," "physio-
logical, " or "stickiness" effects, and those termed "secondary" or "aber-
ration" effects. Primary effects may be described as changes of a
general, or not highly localized, character in the physical, and possibly
also the chemical, nature of the chromosome, which are evidenced during
the ensuing metaphase and anaphase in a tendency of sister chromatids
or of different chromosomes to adhere. Sister chromatid fusion results in
bridge formation at anaphase. Secondary effects, or chromosome abera-
tions, are highly localized alterations of the chromatid or chromosome that
are evident as breaks, exchanges, or inversions in metaphase and anaphase
cells subsequent to irradiation. For obvious reasons the terms "pri-
mary," "physiological," "stickiness," and "secondary," as applied to
effects on the chromosome are objectionable. The terms " nonlocalized "
and "localized" are used here as substitutes for "primary" and "second-
ary," respectively.

Localized effects are of interest in this chapter only insofar as the time
of their appearance is related to that of nonlocalized effects and the
period of mitotic inhibition. It seems apparent from evidence now avail-
able that nonlocalized effects are limited to cells that complete mitosis

CELL DIVISION, MORPHOLOGY, VIABILITY 799

immediately after irradiation during the period of diminishing mitotic
activity, while localized effects occur most frequently in those cells that
enter metaphase and anaphase after the period of irradiation-induced
minimum mitotic activity. That localized effects are not restricted to
the postinhibition period is evidenced by the recording of acentric frag-
ments 7^2 hours after treatment in Scilla root tips (Marquardt, 1938),
shortly after irradiation in Tradescantia microspores (Sax and Swanson,
1941), at 4 hours in Trillium, Allium, and Vicia root tips (Darlington and
La Cour. 1945), in anaphases of the first meiotic division in Habrobracon
eggs treated in late prophase or metaphase (Whiting, 1945), at 1 hour in
root tips of Vicia (Deufel, 1951), and 22 minutes after treatment in
neuroblasts of the grasshopper (Gaulden, unpublished). The explana-
tion offered by Sax and Swanson that such fragments may be due to
breaks resulting from stresses imposed by stickiness of sister chromatids
at anaphase may also apply to the Habrobracon egg treated at prophase I,
for anaphases of the latter showed fusions between separating chromo-
somes. It is unlikely that this would explain their presence at anaphase
I in the Habrobracon egg treated in metaphase I or at anaphase in the
grasshopper neuroblast treated in very late prophase, for in these no
fusions between separating chromosomes were evident. It will certainly
not account for their presence in metaphases of Scilla, Vicia, Allium, and
Trillium root tips. The frequency of fragments in irradiated cells before
the period of minimum mitotic activity is very low in most cells compared
to the number found in cells after the resumption of mitotic activity.
The high percentage of cells with fragments found by Deufel (1951) in
root tips of Vicia as early as 1 hour after 150 r may not be inconsistent
with other results because he includes in his tabulation "fragments"
connected by constrictions with the remainder of the chromosome as well
as those completely separated from it, and no breakdown of the two types
is given. It may be that the ion densities of X rays of the wave lengths
usually used is not high enough to produce many breaks of the heavily
condensed late prophase-through-anaphase chromosomes; perhaps neu-
trons or a particles would produce a greater frequency of breaks during
this period. It is of interest to note that, though localized and nonlocal-
ized effects in the form of breaks and fusions, respectively, may occur
during the period of decreasing mitotic activity, fusions apparently do
not occur at the breakage loci to give translocations, as Marquardt
(1938) has pointed out.

Nonlocalized effects in the form of clumping of metaphase chromo-
somes or the sticking together of sister chromatids to give anaphase
bridges have been described by numerous investigators (Alberti and
Politzer, 1923, 1924; Strangeways and Oakley, 1923; Kemp and Juul,
1930; Lewitsky and Araratian, 1931; Crow, 1933; White, 1937; Mar-
quardt, 1938, 1950; Sax and Swanson, 1941; Carlson, 1941; Roller, 1943;

800

RADIATION BIOLOGY

Darlington and La Cour, 1945; Rugh, 1950; Deufel, 1951). For the most
part these effects have been seen in fixed and stained material soon after
treatment, and the mitotic stage at which they were induced is not known
with certainty. Direct observations of the postirradiation history of
selected living grasshopper neuroblasts in stages of mitosis identified
before treatment has provided us with some exact information regarding
the stages at which these nonlocalized effects are produced and the rela-
tion of this to dosage (Carlson and Harrington, unpublished). With a
series of doses ranging from 64 to 4096 r it is found that successively
increasing doses give rise to successively increasing degrees of response as
follows: (1) delayed anaphase separation of a few of the sister chromatids

Fig. 11-13. Chromosomes of Chortophaga neuroblast, showing X-ray-induced sticki-
ness at anaphase. 512 r; 30 minutes after treatment: A, early anaphase; B, late
anaphase.

in cells that were in very late prophase at the time of treatment and,
therefore, the last cells to pass through mitosis before the interval of
minimal mitotic activity, (2) a similar effect but with many sister
chromatids involved (Fig. 11-13), (3) stickiness involving different
chromosomes, induced at very late prophase and evident in prometa-
phase, and anaphase, (4) fusion of different chromosomes by treatment at
the same stage in which it is observed, viz., prometaphase, metaphase, or
anaphase, and (5) fusion of all the chromosomes into a single, irregular
mass at prometaphase, metaphase, or anaphase as a result of treatment at
the same stage. Subsequently, this chromosomal mass elongates in the
direction of the poles and appears to be divided by a pressing inward of
the cleavage furrow (Carlson, 1941).

It has been postulated that the stickiness induced in chromosomes by
irradiation is due to depolymerization of the thymonucleic acid of the
chromosomes (Darlington, 1942) and to an excess of nucleic acid charge

CELL DIVISION, MORPHOLOGY, VIABILITY 801

(Darlington and La Cour, 1945). The in vitro nucleic acid studies of
Sparrow and Rosenfeld (1946) and Taylor, Greenstein, and Hollaender
(1947, 1948) showed a viscosity fall that indicated at least a partial
depolymerization of the nucleic acid, and the viscosity continued to fall
for several hours after the cessation of X irradiation. This parallels the
results obtained by Harrington and Koza (1951), who found that the
methyl green staining reaction of X-rayed grasshopper chromosomes
reached a minimum as late as 10 hours after X-raying. It is probable
that many of the abnormal anaphases studied by Marshak (1938) in
Vicia and Allium root tips 3 hours after X irradiation were the result of
nonlocalized rather than localized effects. Though depolymerization of
nucleic acids is interfered with at high pH, it seems doubtful whether the
capacity he found for increasing concentrations of ammonium hydroxide
to decrease the percentage of X-ray-induced abnormal anaphases could
be due to a raising of the intracellular pH to the level necessary for such
an effect.

Changes may also be induced by X rays in the intranuclear chromo-
somes. The studies of Duryee (1939, 1947, 1949, 1950) were made on the
later stages of amphibian oocytes, when the "lampbrush" chromosomes
of the large germinal vesicle consist of chromonemata with chromomeres
spaced at intervals along them and numerous lateral loops attached at
both ends to the central chromonemata. Doses of 5000-10,000 r and
more produced breakage in the lateral loops. Chromonemata were occa-
sionally broken by 10,000 r; multiple fragmentation was produced by
30,000 r and more of X rays (Fig. 11-14). Such changes, which were
visible 15 minutes after treatment, appear to be prophase manifestations
of chromosome breakage that are not discernible in most cells until the
succeeding metaphase and anaphase, when they would be classed as
chromosome aberrations.

The chromosomes of the Chortophaga neuroblast, after 250 r of X rays,
undergo changes suggestive of reversion to an earlier stage of mitosis
(Carlson, 1940). The diameter of the chromosome thread resembles,
successively, that of the middle prophase and then of early prophase
cells. Instead of being uniform in diameter from end to end, however, it
has a beaded appearance, consisting of granules separated by narrower
regions. It finally acquires the coarsely granular character of the inter-
phase nucleus. This is a reversible, and not a degenerative, change,
however, for at the time of recovery these cells pass through mitosis in an
apparently normal fashion.

Marquardt (1938) described excessive relational coiling of chromatids
in Bellevalia microspores 2-3 hours and abnormally short metaphase
chromosomes 3-4 hours after X-raying. Defective internal and rela-
tional coiling of the chromatids, according to Darlington and La Cour
(1945), results from X-ray-induced depolymerization of the chromosome

802

RADIATION BIOLOGY

Fig. 11-14. X-ray-induced fragmentation of intranuclear chromosomes of ovarian
eggs. Rana catesbiana. A, control; B, 2000 r, some fragmentation of chromosomes;
C and D, 50,000 r, much chromosome fragmentation (Duryee, 1949).

nucleic acid. Elongation of the chromonemata of Trillium chromosomes
between diakinesis and the first meiotic anaphase is reduced to consider-
ably less than half of normal by X-raying cells in the first meiotic pro-
phase (Sparrow, 1946). It is suggested by Sparrow that this may result
from X-ray-induced DNA deficiency.

CELL DIVISION, MORPHOLOGY, VIABILITY 803

Irradiation of isolated amphibian eggs with as little as 1000 r of X rays
is sufficient to affect the appearance of the nucleoli (Duryee, 1949).
They are changed from small, rounded bodies somewhat irregular in
shape and with small internal vacuoles to much larger bodies more nearly
spherical in form and with large internal vacuoles. Cattley (1909)
noted an increase in the number of nucleoli per cell in plant root tips soon
after X irradiation. Grasshopper neuroblasts subjected to a dose of
10,000 r of X rays at telophase exhibit shortly afterward several spherical
instead of the usual two nucleoli (Carlson and McMaster, 1951). Treat-
ment of neuroblasts at other stages of the mitotic cycle or at interphase
fails to alter the nucleoli.

With regard to the means by which high-energy radiations act on the
nucleus, Duryee (1939, 1947, 1949) has made a strong case for an indirect
effect through the cytosome. Fragmentation of chromosomes and their
lateral loops and vacuolation and enlargement of nucleoli were used as
criteria of radiation damage. X irradiation of nuclei in situ or micro-
injection of the cytosome of nonirradiated eggs with cytoplasm withdrawn
from irradiated ones produced these changes. Immersion of isolated,
untreated nuclei in irradiated cytoplasmic brei led to loss of chromosomal
loops and heavy nucleolar damage. On the other hand, nuclei irradiated
with comparable doses after removal from the egg cytosome exhibited no
detectable injury. Duryee has concluded, on the basis of these results,
that the primary physical or radiochemical changes are produced in the
cytosome, that substances toxic to the nucleus accumulate in the cyto-
some, and that the subsequent movement of these toxins into the nucleus
effect the morphological changes seen after irradiation.

Achromatic Figure and Cleavage. It has generally been found that only
very large doses of radiation, i.e., many thousands of r, affect the achro-
matic figure of the dividing cell. Henshaw (1940d, 1941) demonstrated
that 62,400 r administered to either gamete of Arbacia produced multi-
polar cleavage in almost 100 per cent of the zygotes. Polyspermy was
ruled out as a cause of this, for cytological examination showed that the
percentage of polyspermy was no greater in treated than control speci-
mens. He concluded, because multipolar cleavage was present after
treatment of the sperm, which contains almost no cytoplasmic material,
as well as the egg, which contributes no aster in fertilization, that the
supernumerary asters must result from an effect on nuclear rather than
cytoplasmic material. Unless one is willing to assume that the effective-
ness of the treatment depends on the quantity of cytoplasm irradiated or
that the sperm is devoid of cytoplasm exclusive of centrioles, the possi-
bility of an indirect effect through the cytoplasm is not eliminated com-
pletely. As much as 8000 r applied to the dividing grasshopper neuro-
blast has no demonstrable effect on the structure or functioning of the
spindle.

804 RADIATION BIOLOGY

In the amphibian egg, X-ray doses of 50,000 r and more have been shown
by Duryee (1949) to produce enough solvation of the karyoplasm, or
presumptive spindle substance, to disorient certain of the chromosome
pairs within the nucleus. In the light of present information the 7-irradi-
ated Chaetopterus "chromosomes" that failed to move to the poles at
anaphase (Packard, 1918) were apparently acentric fragments instead of
chromosomes with radiation-induced destruction of the capacity to
develop spindle fibers, as postulated by Packard. Large doses applied
to dividing animal cells lead to an immediate fusion of all the metaphase
or anaphase chromosomes to form a single mass from which chromatin
material seems to flow toward opposite poles, but there appears to be no
change in spindle appearance or behavior (Alberti and Politzer, 1923,
1924; Carlson, 1941; Rugh, 1950).

On the other hand, Marquardt (1938) and Roller (1943), whose
observations were made on Scilla root tips and Tradescantia microspores,
respectively, have concluded that small doses of X rays (360 r in Trad-
escantia) can lead to abnormal orientation or complete suppression of
spindle formation soon after treatment. Roller used delay of the chromo-
somes in attaining metaphase after breakdown of the nuclear membrane
and clumping of chromosomes without any orientation as evidence of the
absence of the metaphase spindle in the Tradescantia microspore. In
Scilla, absence of the spindle led to a certain disorientation of the meta-
phase and anaphase chromosomes, but the latter exhibited repulsion of
sister chromatids toward the opposite sides of the cell and a certain degree
of stickiness.

CELL VIABILITY EFFECTS

Nature of Effect. The presence of pyknotic and degenerating cells in
tissues after irradiation, the destruction of malignant growths, and the
failure of animals to hatch or seeds to germinate may all be the result of
irradiation-induced cell-lethal effects. We are concerned in this chapter
only with those cell-lethal effects that are evident very soon after
irradiation.

Degenerative changes in irradiated cells of the tadpole brain and eye
are described as follows by Spear and Glucksmann (1938). At first there
is separation of chromatic from nonchromatic nuclear material, the
former gradually accumulating in the peripheral region of the nucleus,
while the latter forms a large central vacuole. Following this, the
nucleus breaks up into a number of parts, some of which contain deeply
staining chromatin scattered through the cytosome. Eventually the
cell undergoes fragmentation and dissolution. The chromatic elements
of the nucleus, which change from Feulgen positive to eosin positive,
gradually become smaller as they dissolve. A similar description is given
by Lasnitski (1943b, 1946) for the degeneration occurring in resting

CELL DIVISION, MORPHOLOGY, VIABILITY 805

cells following large doses of radiation (2500 r or more). She recognizes
also "mitotic degeneration" resulting from the unsuccessful attempt of
irradiated cells to undergo mitosis. In this type "the chromatic material
(chromatopycnosis) is assembled in bands (hyperchromatosis) which
later tend to accumulate at the nuclear membrane and finally (chromato-
lysis), undergo shrinkage and lysis. The cytoplasm of these cells under-
goes fatty or colloquative degeneration at an early stage, shrinks, and
disappears."

The formation of large intranuclear vacuoles surrounded by irregular
layers of desoxy pentose nucleic acid within a few hours of the injection of
P32 is described for the intestinal cells of rats by Warren, Holt, and
Sommers (1951). Such changes lead to eventual degeneration.

On the basis of direct observations of living chick fibroblasts in culture,
Strangeways and Oakley (1923) described the "breaking down" of cells
after X irradiation and concluded that this resulted from damage to cells
about to divide. Some cells were not affected until telophase, when one
or both of the daughter cells broke down. Their observations have been
confirmed and extended by more recent investigations based mainly on
material fixed and stained at intervals following treatment. Degenerate
cells in irradiated tissues seem to belong to one of three classes:

(1) After doses of ionizing radiation sufficiently small for mitotic
recovery to take place within a few hours after treatment, the maximum
number of degenerating cells coincides more or less closely in time with
the high point of mitotic activity that marks recovery from the mitosis-
depressing effect of radiation (Strangeways and Fell, 1927; Tansley et al.,
1937; Spear and Glucksmann, 1938; Glucksmann and Spear, 1939;
Lasnitski, 1940, 1943a; Tansley et al., 1948). The close correspondence
between rise in mitotic activity and the degenerate cell count suggests
that the effect of the irradiation has been to render cells about to enter
division incapable of completing it, so that they degenerate during or
after division. If the radiation dose and other conditions are such that
a large proportion of the dividing cells complete mitosis instead of or
before degenerating, the degenerate cell high will follow the mitotic high ;
if they are such that few of the cells that undertake division complete it,
undergoing degeneration instead, the degenerate cell high may precede
the mitotic high (Glucksmann and Spear, 1939; Lasnitski, 1943a).
Glucksmann and Spear (1939), who altered the mitotic activity of the
cells of the tadpole eye by fasting and by exposure of the animals to
low temperatures, were able to correlate the amount of cell degeneration
following 7 irradiation with the amount of mitotic activity of the tissues.
Since the degenerate cell counts in many of their experiments greatly
exceeded the mitotic counts, they reached the conclusion that it was net
the cells in division at the time of treatment, but those approaching
division, that subsequently constituted the bulk of the degenerating cells.

806 RADIATION BIOLOGY

Feeding one week after irradiation, which occurred on the seventh day
of fasting, led to the appearance of degenerate cells, presumably because
feeding stimulated the cells to undertake division.

(2) After the exposure of chick fibroblasts in vitro to such large doses of
X radiation that mitosis did not reappear within at least 24 hours of
treatment, Lasnitski (1943b) found that there nevertheless occurred an
appreciable rise in the number of degenerate cells. This reached a
maximum 3 hours after the end of treatment. Doses of 2500, 5000, and
10,000 r were followed by 53, 58, and 75 per cent, respectively, of degener-
ate cells. She has postulated that the lethal radiosensitivity of these
cells may be positively correlated with their proximity to mitosis, so that
at successively higher doses, cells successively farther removed from
mitosis may be killed, even though they make no attempt at cell division.

(3) Whether the radiation dose is large or small as judged by its effect
on mitotic activity, degenerate cells are generally present at least in
small numbers, very soon after treatment, when the mitotic activity is at
a minimum or completely absent (Strangeways and Fell, 1927; Tansley
et al., 1937; Spear and Glucksmann, 1938; Glucksmann and Spear, 1939;
Lasnitski, 1943a, b, 1945). Their interpretation that this is an effect on
cells that were in division during treatment is confirmed by the work of
Simon-Reuss and Spear (1947), who observed the breaking up of living
fibroblasts that were in metaphase, anaphase, or telophase at the time of
treatment. Such degenerating cells represent only a small proportion of
the total number of cells present, regardless of the dosage, because in the
tissues studied (tadpole brain and eye, avian fibroblasts, rat retinal cells,
mouse malignant cells) , the number of dividing cells is small in relation to
the number of resting cells. It is interesting to note that in contrast to
the results obtained with chick fibroblast preparations, in which cells
may break up before completing mitosis after as small a dose as 88 r
(Simon-Reuss and Spear, 1947), neuroblasts of Chortophaga complete
mitosis almost without exception after doses at least as large as 4000 r,
yet the latter cells are much more radiosensitive than the former when the
criterion is decreased mitotic activity (Fig. 11-6).

By comparing the lethal effects of X radiation on malignant cells of the
mouse in vitro and in vivo Lasnitski (1947) concluded that the immediate
effect of the treatment was mainly a direct one, for the amounts of cell
degeneration in the two were similar. A much higher percentage of cell
degeneration in vivo than in vitro occurred on the second day after
treatment, however, which suggested an indirect effect through damaged
blood circulation.

The nuclear changes leading to pyknosis and cell disintegration in the
amphibian egg are described by Duryee (1949) as follows:

(1) Prophase chromosomes normally transparent and invisible become shorter,
thicker, darker, and beaded. (2) In young ovocytes the nascent chromomere

CELL DIVISION, MORPHOLOGY, VIABILITY 807

lateral loops swell and disintegrate. (3) Nucleoli along the inner surface of the
nuclear membrane evert their contents into the cytoplasm, and their residual
shells swell and disintegrate. (4) The central nuclear ground substance . . .
changes from a gel to a sol, thereby allowing the chromosomes to tangle, mat, and
clump. (5) Later radiation damage with advanced pyknosis consists of forma-
tion of a clumped central nuclear body.

Nuclear damage is induced in ovarian eggs of Triturus within 2 days at
22-24°C by doses of 2000 and more r of X rays applied to the whole body
of the female (Duryee, 1949). Experiments in which the ovaries were
shielded during irradiation showed that this was the result of direct
effects on the eggs and was not secondarily induced through the body of
the female. If the animals were kept at 4°C from the end of treatment
on, the pyknotic changes were temporarily arrested, but not for more than
about two weeks. After warming to 22°C, pyknosis appeared. Main-
tenance of animals at 27°C, however, after irradiation increased the rate
of nuclear disintegration.

Lea (1946) has discussed at some length radiation-induced lethal effects
and their causes in higher organisms. From evidence based largely on
radiation of the Drosophila sperm, the Tradescantia microspore, and the
bean root tip cell, he has concluded that chromosome losses resulting from
asymmetrical interchanges and simple breaks are responsible for at least
some of the cell deaths that occur at or following division. It should be
pointed out, however, that the evidence from these organisms, in which
the analysis of chromosome changes is possible, cannot justifiably be
applied to the elucidation of evidence from cells that have been used in
the study of immediate lethal effects, which are manifest in the treated
cells before, during, or immediately following the first postirradiation
division of the cell. The test of a lethal effect induced in the Drosophila
sperm is failure of the egg fertilized by that sperm to hatch. Chromo-
some losses might be expected to lead to faulty differentiation and death
of the embryo, but this appears to me to be a quite different cause of
death from the immediate effect that would result in the death of the
zygote or one or both of its daughter cells. The same is true of the
bean root-tip cell where the test of lethal effect is the death of the root
many cell generations and many days (about 14) after treatment. The
one example cited by Lea of an immediate lethal effect is the failure of the
irradiated Tradescantia microspore to differentiate or failure of the pollen
tube to develop. The microspore, however, is a haploid cell and any
chromosome loss might be expected to produce an immediate lethal effect.
The chick fibroblast, the cells of the brain and eye of the tadpole, malig-
nant cells of the mouse, and rat retinal cells are not haploid. Unless both
members of a chromosome pair suffered the loss of corresponding regions,
and this would be a rare event except after large doses, there is no reason
to think that the loss would cause the death of the cell. It seems to me,

808 RADIATION BIOLOGY

therefore, that we should look to some effect other than chromosome loss
for the cause of immediate lethal effects on cells.

Hevesy (1945) believes that the basic effect of X rays on cells is the
inhibition of DNA formation. A comparison of the amounts of P32
incorporated in the nucleic acid molecules of irradiated and unirradiated
tissues of the rat showed a significantly higher amount in the latter than
in the former. The percentage of DNA formed was not appreciably dif-
ferent in dividing and nondividing cells. In order to account for the
greater lethal effect in dividing than in nondividing cells — in both the
percentage of DNA formed is about the same — he concludes that inhibi-
tion of DNA formation in a fully developed, rarely dividing group of cells
is not critical, because on the average most of these cells will have time
to eliminate this disturbance before it is their time to divide. The more
frequently the cells of a tissue are dividing, the greater will be the number
of degenerate cells formed, because the cell will attempt to divide before
recovery has had a chance to take place.

The diverse radiosensitivities of different kinds of cells to immediate
lethal effects as compared with mitotic effects are very striking. Tansley
et al. (1937), for example, found that 72 r of y rays, which reduced the
mitotic count in cells of the developing rat retina to a minimum of about
10 per cent of normal, produced a lethal effect on 11 per cent of the total
cells as determined 6 hours after treatment. In the Chortophaga neuro-
blast, however, the mitotic count is reduced to the same extent by as
little as a ninth of that dose, or 8 r of X rays, while 1250 times that dose,
or 10,000 r, causes virtually no neuroblast deaths within at least 8 hours
after treatment (Gaulden, unpublished). This is unrelated to the kinds
of radiation used; for no comparable difference has been detected so far in
the efficiencies of y rays and medium X rays in reducing mitotic activity
or killing cells. Apparently, some basic biological difference in the cells
themselves is responsible. This also demonstrates the fallacy in com-
paring radiosensitivities of different biological materials, when these
sensitivities are based on different criteria.

Under the heading of immediate lethal effects we might also include
chromosome destruction or inactivation produced in Habrobracon eggs by
large doses of X rays (Whiting, 1948). If eggs treated with doses of
14,420-36,050 r in the first meiotic prophase or metaphase are laid by
females mated with untreated males, a small percentage of haploid
males will develop, which contain only the chromosome set of the male
parent. The maternal chromosomes are so adversely affected, presum-
ably by chromatin bridges which interfere with their anaphase move-
ment and with the subsequent movement and internal changes of the
female pronucleus, that they take no part in cleavage and are eventually
eliminated.

CELL DIVISION, MORPHOLOGY, VIABILITY 809

Kind of Radiation. Lasnitski (1948) discovered no striking differences
in the proportion of degenerate avian fibroblasts in vitro exposed to either
1000 r of X or j3 rays (Fig. 11-15). Degenerate cells made their appear-
ance slightly earlier, however, after /3-raying than after X-raying.

A comparison of the efficiencies of fast neutrons and y rays in producing
cell degeneration of the developing rat retina led Spear and Tansley
(1944) to conclude that the former were 6.5 times as effective as the latter
per n unit. The degenerate cell count rises more rapidly after neutron
treatment than after y irradiation. Also after neutron treatment the

IOO-, (0)

0 5 10 15 20

TlME.hr
Fig. 11-15. Effects of 1000 r of X rays (a) and /3 rays (b) on mitosis (heavy, solid
line), abnormal mitoses (broken line), and degeneration of cells (light, solid line) of
avian fibroblasts in vitro (after Lasnitski, 1948).

time of appearance of the degenerate cells does not vary with the dose
as it does after y irradiation, when the larger the dose, the greater is the
delay in their appearance. This is interpreted to indicate that, while
after each irradiation the degenerate cells are made up of those killed
outright during mitosis and those that die later when they attempt, but
fail, to complete mitosis, in the case of neutrons a larger proportion of
the degenerating cells represent direct kills. The much greater effective-
ness of fast neutrons than y rays in killing mice by whole-body irradiation
is attributed by Mitchell (1947), at least in part, to degeneration of radio-
sensitive cells, such as bone marrow. He also suggests that neutrons

810 RADIATION BIOLOGY

can probably kill cells in interphase as well as early prophase, while 7 rays
have their main lethal effects on the latter alone.

Tansley et al. (1948) compared the cell degeneration induced in the
germinative zones of the eye and brain of the frog tadpole by a rays with
that induced by 7 rays. Small doses of each were found to produce
comparable amounts of degeneration, but larger doses of a rays produced
a degree of cell degeneration out of all proportion to that following similar
doses of 7 rays. They suggest that the greater effectiveness of a rays
may be caused either by an effect on cells farther removed from mitosis
than those killed by 7 rays or by an abnormally great accumulation of
injured cells that break up on attempting mitosis.

Dosage Rate. In order to test the efficiencies of different dosage rates
in causing cell degeneration in the brain and eye of the tadpole, Glucks-
mann, Tansley, and Wilson (1945) administered a dose of 336 r of 7 rays
at rates of 5.05, 8.37, 12.6, 15.0, and 20.1 r/minute. They found that the
dosage rate is positively correlated with the maximal degeneration count
and with the length of the interval of time between irradiation and the
maximal count, but that the total amount of degeneration is greatest at
15.0 r/minute. Lasnitzki (1946) determined the relative efficiencies of
dosage rates of 9.7, 29, and 101 r/minute in producing cell degeneration
in chick fibroblasts in vitro. After a dose of 100 r the number of degenera-
tions expressed as percentage of resting cells was negatively correlated
with the dosage rate, but the differences were very slight. After 2500 r,
however, there was a positive correlation of dosage rate with degeneration.
At the lower dose the degenerations were mainly mitotic ones, but at the
higher dose they were mainly resting cell degenerations. Her interpreta-
tion of these apparently contradictory dosage-rate efficiencies at small
and large doses is based on the relation of degeneration to the lowering of
mitotic activity after the different dosage rates.

Temperature. The experiments of Strangeways and Fell (1927) indi-
cate that the amount of cell degeneration that would normally result after
X raying, if chick embryos are incubated at 38°C, is greatly reduced if
they are kept at 0 or 5°C for 5 to 24 hours after irradiation. This is indi-
cated not only by the lowered frequency of degenerating cells seen in
fixed and stained preparations after cold treatment, but also in the
greater proportion of successful in vitro cultivations obtained from cells
that had been cold treated as contrasted with those incubated only at
38° C after treatment.

ULTRAVIOLET RADIATIONS

Unlike X-ray studies, ultraviolet radiation investigations have of
necessity been limited to single, isolated cell types or to cells that are
situated or can be grown in cultures on the surface of tissue masses

CELL DIVISION, MORPHOLOGY, VIABILITY 811

and in which, therefore, penetration of the radiation is not a serious
problem.

MITOTIC EFFECTS

Nature of Effect. Reduction of mitotic activity immediately following
treatment with mixed wave lengths of ultraviolet radiation has been
described for chick fibroblasts in vitro (Kemp and Juul, 1932; Juul and
Kemp, 1933; Mollendorff and Laqueur, 1938), marine invertebrate eggs
(Nebel, Harvey, and Hollaender, 1937; Chase, 1938; Blum et al., 1949;
Blum et al., 1950; Blum and Price, 1950), salamander corneal eipthelium
(Politzer and Alberti, 1924) and rat corneal epithelium (Buschke,
Friedenwald, and Moses, 1945). After monochromatic radiation, divi-
sion of the marine invertebrate egg is retarded by various wave lengths of
2260-3130 A inclusive (Hertel, 1905; Nebel et al., 1937; Giese, 1938a, b,
1939a, b, c, 1946; Marshak, 1949; Wells and Giese, 1950). Mitotic
activity is reduced in the rat corneal epithelium by wave lengths 2480
through 3160 A (Friedenwald et al., 1948). In the grasshopper neuro-
blasts, only 2250 and 2537 A have been studied in detail and found to
retard division (Carlson and Hollaender, 1944, 1945, 1948) but pre-
liminary experiments indicate that longer wave lengths, up to and
including 3130 A, are probably also effective.

Comparative studies of the effectiveness of different wave lengths
point to the 2480-2804 A region inclusive as producing the greatest
retardation of mitosis (Mayer and Schreiber, 1934; Giese, 1938b, 1939a,
b; Friedenwald et al., 1948). Giese (1946) found that treatment of cer-
tain echinoderm sperm gave an action spectrum for delay through the
third cleavage that resembles the absorption spectrum of nucleoprotein
(high at 2600 A), while treatment of the unfertilized egg gave an action
spectrum resembling protein (high at 2800 A) . He suggested that cleav-
age may be slowed by different types of induced changes in the sperm and
egg. Friedenwald et al. (1948), however, found that the action spectrum
for mitotic activity in the rat corneal epithelium was high at 2480 and
2804 and lower at 2650 A.

Of the tissues so far examined only periblem cells of the root tip of
Vicia have given evidence of an increase in the percentage of cells in
mitosis immediately after treatment with ultraviolet radiation. Taka-
mine (1935) found that after exposure of one side of root tips to the radia-
tions from a quartz mercury vapor lamp, 2-3 hours after an exposure of
3^-3 hours, the percentage of cells in mitosis (prophase, metaphase,
anaphase, and telophase) on the exposed side of the root tip was about
1 per cent higher than on the opposite side. Subsequently, the percent-
age on the treated side fell until at 15 hours it was about 1 per cent less
than the control side; and the percentage was still below normal at 48
hours. The effects of monochromatic radiation of 2537 and 3650 A were

812 RADIATION BIOLOGY

also studied. The latter gave a slight increase in the number of dividing
cells during the first 3 hours after treatment, while the former reduced the
number of mitoses to slightly below normal during 24 hours after treat-
ment. Since the numbers of cells on which these percentages are based
are not given, it is not apparent how significant they are. It is note-
worthy, however, that exposures of 0.5, 1.0, 1.5, and 3.0 hours all showed
a consistently similar effect on mitosis. It is a question also whether
the ultraviolet — especially the shorter wave lengths — would penetrate
the root tip to the periblem, let alone to its deeper layers. A differential
penetration by the wave lengths used is indicated by the different effects
of 2537 and 3650 A. Seide (1925) treated Ascaris eggs in various stages
of the first cleavage division with small doses of ultraviolet radiation of
2804 A and of mixed wave lengths from the mercury quartz lamp, but was
unable to demonstrate a positive shortening of the time required by the
eggs to reach the two- and four-celled stages. In 1945 Buschke, Frieden-
wald, and Moses described what appeared to be increased mitotic activity
following weak doses of nonmonochromatic ultraviolet radiation of the
rat cornea, but it was demonstrated in a later study (Friedenwald et al,
1948) that the effective agent was a gas — probably ozone — generated by
the ultraviolet lamp used in the treatments.

Of the tissues examined for mitotic activity after exposure to ultra-
violet, only the corneal epithelium of the rat exhibits a compensatory
effect at recovery, i.e., a temporary increase in the number of mitoses in
excess of normal immediately after recovery (Friedenwald et al, 1948).
This peak is reached approximately 12 hours after treatment.

It appears from present evidence that interphase and early prophase
are the stages most sensitive to ultraviolet radiation. Investigating the
radiosensitivity of different parts of the interval between the first and
second cleavages of the Arbacia egg, Blum and Price (1950) found that a
dose of radiation applied in the early part of this interval caused the
greatest mitotic delay, the radiosensitivity then diminished to a minimum
at 12 minutes; and from this time to the completion of the second cleavage
division, irradiation did not induce delay. In studying the effects on the
grasshopper neuroblast of 2250 and 2537 A radiation, Carlson and Hol-
laender (1944, 1948) found that, if the radiosensitivity was measured by
the time required for a cell treated in one stage to reach the next stage,
early prophase was the most sensitive; for cells treated in either inter-
phase or early prophase remained in early prophase an excessively long
time. If, on the other hand, the increase in the time required by the cell
to progress from the stage in which it was treated to anaphase was the
criterion of sensitivity used, at both wave lengths interphase was found
to be more sensitive than early prophase (Carlson and Hollaender, 1948).
A detailed analysis of the radiosensitivities of different stages of mitosis to
2250 A, as indicated by their delay in reaching anaphase, showed middle

CELL DIVISION, MORPHOLOGY, VIABILITY

813

and late prophase to be only slightly less affected than early prophase,
and metaphase to be much less sensitive. Cells in middle prophase were
caused to undergo temporary regression by a dose of 34,650 ergs/cm2
and late prophase by one-fourth as large a dose. Recovery and progres-
sion through anaphase followed. Delay induced in prometaphase and
metaphase cells is probably the result of damage to the spindle-forming
substance and spindle, respectively (see p. 814).

Intensity. In time-intensity studies the problems encountered in the
choice of materials and methods and in the interpretation of results are
essentially the same for ultraviolet as for high-energy radiations (see p.

t.oo

0.80
0.60

0.40
Q

I-
<

cr
o

p 0.20

0.10
0.08

0.06

0

20

24

4 8 12 16

xlO3 ERGS PER CM2
O LOW INTENSITY • HIGH INTENSITY

Fic;. 11-16. Effects of different intensities of 2537 A ultraviolet radiation on the
mitotic ratio of treated to control cells (Chortophaga neuroblasts) (after Carlson and
Hollaender, 1945).

788). In addition, the relatively slight penetration of many ultraviolet
wave lengths into organic materials limits drastically the kinds of cells
amenable to mitotic studies. Carlson and Hollaender (1945) compared
the effectiveness of selected doses of 2537 A radiation delivered in 3.5-13
seconds and in 3.75-4 hours. It was found that the number of cells pass-
ing through mitosis in a 2-hour period beginning approximately 4 hours
after the mid-point of the irradiation period was decreased to approxi-
mately the same extent by doses of 5760 or 11,520 ergs/cm2, but that at
doses of 17,280 or 23,040 ergs/cm2 the brief, high-intensity exposure was
considerably more effective in reducing the mitotic activity than the pro-
longed, low-intensity one (Fig. 11-16).

Chemical Agents. Marshak (1949) found that none of the following
chemical agents were effective in altering 2537 A ultraviolet-induced

814 RADIATION BIOLOGY

cleavage delay in Arbacia: streptomycin, adenosine, folic acid, 2,4-
diamino-5-p-chlorphenoxypyrimidine, or riboflavin.

CELL MORPHOLOGY EFFECTS

Chromosomes. After generative nuclei of Tradescantia pollen tubes
growing on artificial sugar-agar-gelatin medium had been treated in
prophase with a sublethal dose (3000 ergs/mm2) of 2537 A radiation,
the metaphase chromosomes were much shorter and thicker than normal
(Swanson, 1942). This was apparently due to partial despiralization of
the chromonemata, for the number of coils in these chromosomes was
reduced from the normal twenty to about seven to ten per chromosome
and the gyres were increased in width. The chromosome matrix, which
was rarely seen under normal conditions, appeared as a transparent
hyaline mass surrounding and holding together the two chromatids.
Frequently the matrixes of two or more chromosomes were fused,
a change suggestive of the nonlocalized or "stickiness" effect found
immediately after large doses of ionizing radiations. None of these
changes were seen after treatment with a mixture of 2967 and 3022 A
radiation.

If neuroblasts of the grasshopper are exposed to large doses of 2537 or
2650 A radiation and examined subsequently in the living, unstained
state, the chromosomes of cells in prometaphase, metaphase, and ana-
phase appear blurred and indistinct. Fusions between chromosomes may
lead to defective anaphase separation and the hourglass-shaped telophase
chromatin masses seen soon after large doses of ionizing radiations
(Carlson, 1941). A change in the prophase chromosomes from threads of
uniform diameter to a moniliform shape accompanies ultraviolet-induced
mitotic reversion at this stage.

Achromatic Figure and Cleavage. One of the striking effects of certain
wave lengths of ultraviolet is their inhibitory action on spindle develop-
ment (Nebel et al., 1937; Carlson and Hollaender, 1948) and their destruc-
tive action on the fully formed spindle (Carlson and Hollaender, 1948).
Ultraviolet of 2250 A acts on the spindle and its precursor, the karyo-
lymph, in the grasshopper neuroblast very much as colchicine does
(Gaulden and Carlson, 1951). If the spindle is not well formed, the
karyolymph of the nucleus accumulates in one or more hyaline globules
and the spindle remains small, the centromeres of the chromosomes being
held in a compact group by the small spindle. Later, certain of these
centromeres may move poleward along the spindle, but anaphase is not
initiated until all have returned to the equatorial plate. Wave length
2804 A is equally effective in altering the spindle; wave lengths 2399,
2537, 2650, 2967, and 3022 A are less effective (Carlson and McMaster,
unpublished). The action spectrum resembles, therefore, a protein
absorption curve. Destruction of the spindle probably accounts for the

CELL DIVISION, MORPHOLOGY, VIABILITY

815

binucleate cells observed by Takamine (1923) in Allium root tips 1 hour
after treatment with 2500 A radiation.

Normally, in the grasshopper neuroblast, the cleavage furrow forms
near one end of the elongated anaphase cell and the resulting daughter
cells are unequal in size; the daughter neuroblast is large, while the

Q

Fig. 11-17. Diagram showing ultraviolet-induced abnormalities in mitosis of the
Chortophaga neuroblast. A-G, treatment at late metaphase or early anaphase;
spindle abnormally short (A), cleavage furrow appears at cell equator (B, C) to give
an equal division (D, E) or a secondarily produced unequal division (F, G). H-L,
treatment at prometaphase or early metaphase with a dose sufficiently large to prevent
centromere division and chromosome separation. Spindle greatly reduced, its sub-
stance mainly in the form of a hyaline globule (hg); note pseudopodia and distal
separation of chromatids (I-K) {from Carlson and Hollaender, 1948).

daughter ganglion cell is small. The reduction in spindle size induced by
2250 A radiation is apparently correlated with a tendency of the cleavage
furrow to form at the cell equator (Fig. 11-17) (Carlson and Hollaender,
1948). In many cells, as the cleavage furrow deepens, the bulk of the
cytoplasm flows toward one of the poles, so that two more or less normal
daughter cells of unequal size are formed. In some cells, however, no

816 RADIATION BIOLOGY

shift occurs and two cells approximately equal in size result. That this is
caused by the reduced spindle size is evidenced by a similar phenomenon
in colchicine-treated neuroblasts, in which the spindle is also reduced in
size (Gaulden and Carlson, 1951). If the dose of radiation is large enough
to damage the spindle to such an extent that the chromosome halves do
not separate, even at the time when pseudopodia-like outpushings of the
cell indicate the onset of telophase, the chromosomes may all be incorpo-
rated in a single nucleus. One of the pseudopodia is eventually cut off by
a furrow from the nucleated portion of the cell.

Nucleolus. The form of the neuroblast can be altered drastically by
all ultraviolet wave lengths from 2250 to 3130 A (Carlson and McMaster,
1951). This effect is studied most readily in hanging-drop preparations
of the living cell. Normally the nucleolus of the neuroblast appears as an
irregular mass of low refractility during interphase and most of the pro-
phase. Within 25-35 minutes after irradiation the nucleolar mass
becomes transformed into about ten nucleolar fragments, which gradually
separate to form a cluster of highly refractile spherules. If the dose is
large, these subsequently fuse to form fewer and larger spheres, until,
ultimately, a single, clearly defined, spherical body results. If the dose is
less than that sufficient to produce this series of changes, recovery by a
return to the original condition may take place at any stage. The
2399-2804 A region of the spectrum is most effective in producing this
change, which suggests that absorption by both protein and nucleic acid
may be involved in its production.

VIABILITY EFFECTS

Few studies have been made on the capacity of ultraviolet radiations to
kill animal cells soon after treatment; these are all based on mixed wave
lengths and contain no exact information on the doses used. Politzer and
Alberti (1924) described necrosis of the cells of the upper two layers of the
cornea of salamander larvae 1 hour after ultraviolet irradiation. Mollen-
dorff and Laqueur (1938) found that, after moderate doses, fibroblasts in
culture recover and progress through mitosis normally, but after large
doses they break down on attempting division at the end of the radiation-
induced, mitosis-free period. They suggest that ultraviolet irradiation
produces a toxic substance, which acts at the time of division to prevent
the successful completion of mitosis. Buschke' et al. (1945) recorded
aggregation of nuclear chromatin in clumps, followed by fragmentation
of nuclei within 6 hours of treatment, and finally complete destruction of
the cells of the two uppermost layers of the rat cornea. Both low tem-
perature and anaerobiosis, produced either by immersion of the enucle-
ated eye in buffer solution without stirring or by maintaining it in a
vacuum, caused a lag in the appearance of nuclear fragmentation. From
this they concluded that cell disintegration resulting from ultraviolet

CELL DIVISION, MORPHOLOGY, VIABILITY 817

irradiation depends on secondary intracellular processes that follow
the treatment and that, at least in part, can be identified as oxidative
ones.

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818 RADIATION BIOLOGY

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820 RADIATION BIOLOGY

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Habrobracon eggs treated in first meiotic prophase and metaphase. Am. Natural-
ist, 79: 193-227.
(1948) Incidence and origin of androgenetic males in X-rayed Habrobracon

eggs. Biol. Bull., 95: 354-360.

824 RADIATION BIOLOGY

Wilbur, K. M., and R. O. Recknagel (1943) The radiosensitivity of eggs of Arbacia

punctulata in various salt solutions. Biol. Bull., 85: 193-200.
Wilson, C. W., A. F. Hughes, A. Gliicksmann, and F. G. Spear (1935) Bestrahlungs-

versuche an Huhnerembryonen in vitro und in vivo mit Radiumgammastrahlen.

Strahlentherapie, 52: 519-524.
Wright, G. P. (1925) The relative duration of the various phases of mitosis in chick

fibroblasts cultivated in vitro. J. Roy. Microscop. Soc, 45: 414-417.
Yamashita, H., K. Mori, and M. Miwa (1939) The action of ionizing rays on sea

urchin. II. The effects of roentgen, gamma, and beta rays upon the fertilized

eggs. Gann, 33:117-121.
Zirkle, R. E. (1936) Modification of radiosensitivity by means of readily penetrating

acids and bases. Am. J. Roentgenol. Radium Therapy, 35: 230-237.
, P. C. Aebersold, and E. R. Dempster (1937) The relative effectiveness of

fast neutrons and X-rays upon different organisms. Am. J. Cancer, 29: 556 562.

Manuscript received by the editor Nov. 23, 1951

CHAPTER 12

Genetic Effects of Radiation in Mammals1

W. L. Russell

Biology Division, Oak Ridge National Laboratory

Introduction: Effect of radiation on reproduction. Dominant lethals, semilethals, and
subvitals: Offspring of presterile-period matings of irradiated males — Offspring of irra-
diated females — Offspring of poststerile-period matings of irradiated males — Sex ratio.
Dominant sterility. Dominant partial sterility. Dominant visibles. Recessive lethals,
semilethals, and viables. Human hazards. References.

INTRODUCTION

Most of the information on the genetic effects of radiation in mammals
has come from work on the mouse. The small amount of data obtained
from rats, rabbits, and guinea pigs is, in general, confirmatory of the
results on mice. The only form of ionizing radiation that has been used
extensively is X radiation. There is a little information from limited
investigations with neutrons and 7 rays.

Short general review articles have been presented by Snell (1941b) and
Russell (1952). The paper by Lea (1947) includes a review of the work
on dominant lethals and hereditary partial sterility. Methods for the
detection of mutations in mammals have been discussed by Hertwig
(1932), Snell (1935, 1945), Catcheside (1947), Falconer (1949), and
Russell (1951, 1952).

Mutations are commonly divided into two categories, chromosomal and
point mutations, according to whether or not a structural change in the
chromosomes can be detected. Whether radiation-induced point muta-
tions include gene mutations, or represent only certain types of chromo-
somal change, is still debated. Regardless of its exact nature, the dis-
tinction between chromosomal and point mutations has proved to be
descriptively useful in Drosophila. In this organism, where observation
of the fine details of chromosome structure is possible, and where the
chromosomes are so thoroughly marked genetically, even minute struc-
tural changes in chromosomes can be detected, and the level below which
they would no longer be apparent is sharply definable. In mammals, the

1 Work at Oak Ridge and preparation of manuscript under Contract No. W-7405-
eng-26 for the Atomic Energy Commission.

825

826 RADIATION BIOLOGY

available cytological and genetic tests for structural chromosomal changes
are far less critical. The category "point mutations," or mutations in
which structural change has not been detected, is not, therefore, a par-
ticularly useful one in mammals at the present time. In this chapter,
mutations are classified according to their dominance relations and
phenotypic effects. Wherever there is proof, or evidence, that a chromo-
somal change is involved, this is, of course, discussed.

Effect of Radiation on Reproduction. In investigating the genetic
results of radiation in mammals the experimental procedures are in some
respects limited, and in others aided, by the effects of the radiation on the
reproduction of the exposed animals. It is desirable, therefore, to give
an outline of these effects before considering the results of genetic studies.
Furthermore, the two subjects are not unrelated: some of the effects on
reproduction are themselves the result of damage to the genetic material
of the germ cells.

The procedure used in most genetic studies has been the exposure of
males to a single, high intensity dose of X rays. Considering, first, the
effects on reproduction of this treatment, it is now well established by the
results of many investigators that within the range of approximately
400 1000 r, the limits depending on the species and on other factors, a
period of fertility immediately following irradiation is succeeded by an
interval of sterility after which fertility returns. A sample of data on the
lengths of the initial fertile period and the temporary sterile period is
given in Table 12-1. The length of the initial fertile period is probably
underestimated in some of the experiments because of its dependence on
number of copulations (Snell, 1933b; Brenneke, 1937; Hertwig, 1938a), or
because of insufficient testing of males. The data are erratic and there is
no clear-cut dependence on dose. The length of the sterile period, how-
ever, is apparently affected by dose. This conclusion is confirmed by
the findings of Strandskov (1932) who showed that the length of time
during which no motile sperm were present in electrically induced ejacula-
tions of guinea pigs was related to the dose received by the males. Litter
size in the first fertile period is reduced and is related to dose. After the
sterile period the litter size is probably slightly below normal, but so close
to it that a difference is hard to establish. Details on litter size are
presented later.

For doses below about 400 r there is no clear-cut period of complete
sterility. As the dose increases above 1000 r the litter size in the period
immediately following irradiation reduces toward zero, most matings
producing no young at all. At very high doses it appears that some
permanent sterility is induced (Table 12-2), but whether this results from
direct damage to the testis or from the systemic effects, which are notice-
able even with the partial-body irradiation, is not known. In either case,
there appear to be differences in response attributable to the strains of

GENETIC EFFECTS IN MAMMALS

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828

RADIATION BIOLOGY

mice used, or to other variables in experimental conditions, for it can be
seen from Table 12-2 that the data of Hertwig show appreciable incidences

Table 12-2. Incidence of Permanent Sterility in Male Mice Exposed to

Various Doses of X Radiation

Whole-

Permanently sterile

Reference

Dose, r

or part-
body

No. cfcf
tested

irrad.

Number

Per cent

Hertwig (1938a)

400

P

10

0

0.0

500

P

5

• 0

0.0

600

P

10

1

10.0

800

P

36

5

13.9

1000

P

36

11

30.6

1200

P

6

2

33.3

1500

P

21

17

81.0

3000-4000

P

5

5

100.0

Russell et al.a

500

W

4

0

0.0

600
L Control

W

1717

2 + 2?6

0.12-0.23

1134

3 + l?6

0.26-0.35

800

W

6

0

0.0

-

("1000
L Control

P

414

3?c

0.00-0.72

392

2 + 2?<"

0.51-1.02

a Unpublished data of W. L. Russell, Josephine S. Gower, Gloria J. Jasny, and J. C.
Kile.

6 Died less than twelve weeks after mating and tested against only one female each.
c Tested against only one female each.

of permanent sterility with doses of 1000 r and below, while those of
Russell et at. show no difference from the controls even with 1000 r.

Up to the dose level above which breeding experiments become imprac-
ticable, no effects on the viability or motility of mature sperm have been
observed. In electrically induced ejaculations from guinea pigs, Strand-
skov (1932) found motile sperm up to four weeks following exposure to
2592 r of X rays. Snell (1933b) performed a bilateral vasa efferentia
ligation on six mice and irradiated three of them with 800 r of X rays.
There was no difference between the irradiated and nonirradiated animals
in length of survival of motile sperm in the epididymides.

The testis was one of the first organs studied histologically for the
effects of X rays and has since been the object of numerous investigations.
The references cited, particularly Schinz and Slotopolski (1925) and
Glucksmann (1947), may be used as an introduction to the voluminous
literature which can be only briefly summarized here. Hertwig (1938b)
and Schaefer (1939) have studied the subject with particular regard to
its bearing on genetic problems. There is now almost unanimous agree-
ment that spermatocytes, spermatids, and sperm, along with Sertoli cells

GENETIC EFFECTS IN MAMMALS

829

and interstitial tissue, are resistant to radiation, and that spermatogonia
are quite sensitive.2 Reduction in numbers is found first in the spermato-
gonia. The later germ cell stages disappear in the order in which they
are formed in spermatogenesis and at approximately the same rate as
that of the spermatogonial disappearance (Hertwig, 1938b; Eschen-
brenner and Miller, 1950; Fogg and Cowing, 1951a, b). As there is
evidence that the spermatocytes and spermatids present at the time of
irradiation complete their development (Schaefer, 1939; Eschenbrenner
and Miller, 1950), the disappearance of these classes is attributable to
failure of replacement by the depleted spermatogonia. Since Fogg and
CoAving (1951a, b, 1952) nowhere mention this commonly accepted view,
and even wrongly imply that Eschenbrenner and Miller adopted another
explanation, it must be concluded that their own suggestion, that the
progressive disappearance of later stages reflects a delay in response to
irradiation correlated with cell specialization, was made in unawareness
of the consequence of loss of spermatogonia. The number of spermato-
gonia rapidly reaches a minimum. The reduction in number was
measured, in mice, by Hertwig (1938b) for a dose of 800 r, by Eschen-
brenner and Miller (1950) for 400 r and, less quantitatively, by Fogg and
Cowing (1951a, b) for other doses (Table 12-3). The number of sperma-

Table 12-3. Reduction in Number of Spermatogonia in Mice Exposed to

Various Doses of X Rays

Reference

Dose,
r

Days after
irradiation

when minimum
number of

spermatogonia
observed

Number

of cross

sections

of tubules

examined

Per cent of

cross sections

showing any

spermatogonia6

Per cent

of normal

number of

spermatogonia

Fogg and Cowing (1951a)

Eschenbrenner and Miller (1950)
Hertwig (1938b)

300

400

800

1440

7

7°
4-6
10

1000

59
200

10

14
1

1.5«

0 76d

Fogg and Cowing (1951b)

° No observation made at earlier time.

6 Fogg and Cowing state that normal testis shows spermatogonia in 80 per cent of cross sections of
tubules.

c Based on reduction of resting spermatogonia to 3.3 per cent and of mitotic spermatogonia to 0
per cent. Eschenbrenner, Miller, and Lorenz (1948) give the ratio of these two types in testes of unirra-
diated mice of similar age as 8.9: 11.1.

d Hertwig found a total of 9 spermatogonia in the 59 cross sections examined and states that a cross
section of a tubule of a normal testis contains 20 spermatogonia.

togonia then increases, and eventually the later germ cell stages reappear
in the order in which they disappeared (Hertwig, 1938b; Eschenbrenner
and Miller, 1950; Fogg and Cowing, 1951a, b). The time taken, from the

2 Unpublished data, obtained by E. F. Oakberg at this laboratory since this manu-
script was submitted, show that primary spermatocytes, as well as spermatogonia,
show appreciable sensitivity to radiation.

830 RADIATION BIOLOGY

first reappearance of mitotic spermatogonia to the production of sperm, is
approximately four weeks in the mouse. The interesting results of
Howard and Pelc (1950), who used autoradiographs, following P32 injec-
tion, to measure the times taken by various stages of spermatogenesis in
the mouse, show that the interval from spermatogonial metaphase to
immature sperm is more than 10 days.

The surviving cells have been referred to as spermatogonia. This is the
view taken by most authors, including Hertwig (1938b) who identifies the
cells as spermatogonia with "dusty," pale, oval nuclei. Other interpreta-
tions are mentioned by Hertwig. Among the spermatogonia there is
great variety in size and appearance of nuclei. The frequencies of these
types at various intervals after irradiation, together with interpretations
of what stages in spermatogonial divisions the types represent, are also
presented by Hertwig.

From the histological observations it appears that matings made in the
initial period of fertility following irradiation utilize sperm that were
mature at the time of irradiation, and that copulations occurring in the
latter part of the period may use some sperm that matured from cells that
were spermatids or spermatocytes when irradiated. The period of
temporary sterility corresponds to the interval in which replacement of
later germ cell stages has not yet been completed owing to the depletion
of spermatogonia. Matings made after the sterile period use sperm that
were in the spermatogonial stage at the time of irradiation. The sterile
period is thus a useful marker for separating genetic changes induced in
spermatogonia from those induced in later germ cell stages. The reduc-
tion in litter size in the initial fertile period is not accounted for by the
histological findings described above, but is explained by the results
described in the next section.

The breeding results and histological changes in the gonads following
exposure to neutrons or y rays, and following the use of X rays on
females, will be described only briefly since these treatments have had
only limited use in genetic studies.

Snell and Aebersold (1937) have shown that the effects on reproduc-
tion, following a single exposure of male mice to cyclotron neutrons, are
similar to those produced by X rays. However, measured on the
roentgen scale of a Victoreen condenser-type dosimeter, neutrons are
from five to six times as potent as X rays in reducing the litter size in pre-
sterile-period matings.

The effects on the gonads and on fertility of long exposure to low
intensity 7 radiation have been described by Lorenz et al. (1947) and
Eschenbrenner et al. (1948). Vigorous hybrid male mice exposed to a
total dose of 1100 r at 8.8 r per day from a radium source and then
removed from the radiation field gave litters of reduced size in early
matings and normal litters later. Those exposed to 1760 r at the same

GENETIC EFFECTS IN MAMMALS 831

rate were sterile at first, but recovered fertility after two months. Expo-
sure to 1100 r at 4.4 r per day did not affect fertility. Hybrid females
exposed to total doses of from 770 to 880 r, regardless of rate, and then
mated, were either sterile or became so after the production of one, or
rarely two, litters of reduced size. When inbred strains were used
instead of hybrids, the same effects in males and females were produced
at lower doses. Histological observations on the testis showed reduction
in spermatogenic elements to stable levels which were dependent on dose
rate. The relative proportions of the cell types were, however, normal
except in males exposed to 8.8 r per day. In these animals, after ten
months, the sperm were relatively few in number and multinucleated
spermatids were common. Eschenbrenner et al. suggest that these effects
resulted from the degeneration of Sertoli cells which was observed only in
this material.

Acute X-ray exposure of female mice with a dose of 150 r, or perhaps
lower, results in permanent sterility. One, and occasionally two, litters
can be obtained, even at much higher doses, before the sterility sets in.
The figures, 800 to 1500 r, given by Glucksmann (1947) as the permanent
sterilizing dose for the female mouse and rat are too high by a factor of
about 10, and are perhaps either a misprint or refer to the dose at which
even the temporary fertility following irradiation is suppressed. The
litter size in this temporary fertile period is reduced, and Snell and Ames
(1939) report that it falls off more rapidly with increasing dose than does
the litter size from irradiated males.

DOMINANT LETHALS, SEMILETHALS, AND SUBVITALS

For purposes of discussion here, lethals are defined as mutations that
cause death usually before birth, semilethals as those which cause death
usually between birth and reproductive age, and subvitals as mutations
that sometimes cause death.

Offspring of Presterile-period Matings of Irradiated Males. Radiation-
induced dominant lethal effects in mammals were reported as early as
1908 by Regaud and Dubreuil who observed a high proportion of abnor-
mal embryos in rabbits sired by males that had been exposed to X rays.
It was not until many years later, however, that it became probable, and
was finally demonstrated, that induced chromosomal aberrations are a
cause of this class of abnormalities. Strandskov (1932) found a mark-
edly reduced litter size in the immediate progeny of X-rayed male guinea
pigs. Since electrically induced ejaculations showed numerous motile
sperm during the times the litters were produced, Strandskov assumed
that the normal number of eggs was fertilized, but that some were later
resorbed. He cited the results of Regaud and Dubreuil, and similar
findings in amphibia, in support of the view that some embryos died.

S32

RADIATION BIOLOGY

He concluded: "The most plausible interpretation seems to be the
induction of dominant lethal mutations." Snell (1932, 1933b), working
on mice, also found a reduction in litter size in early matings of X-irradi-
ated males and no effect on motility of sperm. In the same material he
found that many embryos died usually at, or shortly after, implantation.
He also demonstrated the presence of both male and female pronuclei in
all of fourteen eggs fixed and sectioned 16 hours after impregnation.
Snell concluded that the reduction in litter size, caused by the death of
embryos, is most plausibly explained by the induction of chromosome
abnormalities. Final proof of the correctness of the interpretations of
Strandskov and Snell was provided by Hertwig (1935), Hertwig and
Brenneke (1937) and Brenneke (1937). In the first place, it was demon-
strated that fertilizing capacity of mouse and rat sperm is unaltered even
by high doses. Table 12-4 shows the results obtained for doses up to

Table 12-4. Lack of Effect of Radiation on the Percentage of Unfertilized
Eggs in Presterile-period Matings of X-irradiated Male Mice and Rats

(Data from Brenneke, 1937)

Species

Dose,
r

No. of eggs
examined

Unfertilized

Prob.Dift.

Number

Per cent"

Mouse

800

447

24

5.37

1400

134

8

5.97

1800

171

14

8.19

2200

95

3

3.16

Total irrad.

847

49

5.791

8.86/

0.14

Control

158

14

Rat

800
1400

288
186

14
10

4.86
5.38

1800

54

7

12.96

Total irrad.

528

31

5.871
4.72/

0.64

Control

106

5

a Percentages given in Brenneke's table have been changed to agree with the num-
bers given by her.

2200 r in the mouse and 1800 r in the rat. Hertwig and Brenneke (1937)
state that even with 4000 r, the highest dose used, there was still no effect
on fertilizing ability of mouse sperm. Second, the authors showed that
as soon as the eggs cleave, differences between experimentals and controls
become apparent. Supernumerary nuclei were found sometimes in the
blastomeres of the two-cell stage and more often in the cells of later
stages. In metaphase of cleavage mitoses, portions of chromatin lying
away from the equatorial plate could be found, and, in anaphase, lagging
chromosomes were observed. Amoroso and Parkes (1947), using arti-

GENETIC EFFECTS IN MAMMALS

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834 RADIATION BIOLOGY

ficial insemination of rabbit spermatozoa irradiated in vitro, have con-
firmed both the lack of effect of high doses on fertilizing capacity of sperm
and the occurrence of blastomeres with supernumerary nuclei.

Turning to some of the details of the experimental findings, data for
effect on litter size in presterile-period matings of irradiated males are
given in Table 12-5. The litter sizes are based on the numbers of litters
actually born. This underestimates the effect of radiation, particularly
at higher doses, because some pregnancies, the proportion increasing with
dose, fail to produce any young at term. Even without this correction,
the data in Table 12-5 clearly show that litter size is dependent on dose.
An effect of dose on litter size is also indicated in the few litters allowed to
go to term in the work of Amoroso and Parkes (19-47) on artificial insemi-
nation of rabbit spermatozoa irradiated in vitro. In the data of both
Snell and Hertwig, the reduction in litter size is greater for the later
matings within the presterile period. The proportion of pregnancies that
fail to reach term is, according to Hertwig (1938a), also greater in the
later matings. Although the results of Russell et at. do not show a greater
drop in litter size in the later matings, it should be pointed out that they
were obtained from an experiment in which each male was mated to only
one female in each of the two weeks and, therefore, possibly expended his
store of sperm at a slower rate than that occurring in the experiments of
Snell and Hertwig. The data of Brenneke (1937) show that the propor-
tion of eggs having abnormal cleavages may increase both with number
of matings and time. These data also provide the most direct evidence
that the increased effect in later matings is attributable to a higher inci-
dence of dominant lethals.

Information on the time of death of the embryos carrying dominant
lethals is given in Table 12-6. Additional information is provided by the
studies of Brenneke (1937) on the proportion of abnormal cleavages found
at various times after fertilization. Brenneke 's results for 800 r showed
abnormality in 23.2 per cent of 186 two-cell stages, 44.4 per cent of 90
three- to six-cell stages and 41.9 per cent of 105 seven- to twelve-cell
stages. Because of the possibility of further cleavage of some of the
abnormal embryos found at any one stage, the total percentage of
abnormality cannot be accurately estimated from these data. However,
it is clearly higher than the percentage of preimplantation death esti-
mated from the 800 r data of Snell and of Russell et at. If the difference
proves to be real, it would indicate that some of the embryos showing
abnormal cleavages actually survive to implantation. Most of the
dominant lethal types that survive to implantation were found by Snell
to die shortly thereafter. This appeared to be true in the material of
Russell et al, but, with the females opened at a later stage in pregnancy,
the time of death could not be accurately determined. Henson found
degeneration occurring shortly after implantation in six of eleven rat

GENETIC EFFECTS IN MAMMALS

835

embryos examined on the eighth or ninth day of pregnancy of females
mated to males exposed to 1000 r. There is some evidence, from a com-
parison of implantation data for 1200 r with the two sets of data for
800 r (Table 12-6), that the effect of raising the dose is to increase the
ratio of preimplantation to postimplantation death; for, although no
controls are given for the 1200 r data, the low mean number of implanta-
tions indicates a high incidence of preimplantation death. This effect of
an increase in dose, as Lea (1947) pointed out, could be attributed to
more of the sperm carrying more than one lethal chromosome change and
to this resulting in a relatively greater death in early stages. The data
of Amoroso and Parkes (1947) on irradiation of rabbit spermatozoa in

Table 12-6. Time of Death of Embryos Carrying X-ray-induced Dominant

Lethals

Mean

no. implants

Species

Dose

to dV,
r

No.
preg-
nant

99

Day after
fertiliza-
tion when

99
dissected

per pregnancy

Estimated

per cent of

induced death

occurring

before

implantation

Reference

Living

embryos

Dead

embryos
and re-
sorption

Total

sites

Snell (1933b)

Mouse

Control

9

5-11

7.22

0.33

7 . 56"

Mouse

800

8

5-11

2.63

3.88

6.50"

23

Brenneke (1937)

Mouse

1200

19

9-13

0.74

3.05

3.79

Henson (1942)

Rat

Control

5

8-14

9.00

Rat

1000

8

8-14

7.13

Russell et al. (1951).

Mouse

Control

47

16-18

8.19

0.98

9.17

Mouse

800

50

16-18

3.92

3.64

7.56

38

" Excludes an insignificant proportion of unclassified embryos.

vitro show that, when very high doses are given to the sperm, death of
embryos takes place still earlier in preimplantation development.

The exact nature of the chromosomal aberrations that cause dominant
lethality in mammals is not known. Snell (1933b) pointed out that his
data, Table 12-5, fit the interpretation that litter size falls off logarith-
mically with dose. This would be expected if the dominant lethals were
the result of single-hit effects. However, the data are not extensive
enough to warrant definite conclusions on the relation of lethality to dose.
It is difficult to obtain accurate information on this relation because with
increasing dose there is an increasing proportion of fertile matings that
produce no young at term and possibly an increasing proportion of still-
births, some of which may be missed. The cytological observations of
Brenneke (1937) and Amoroso and Parkes (1947), as far as they go,
strengthen the view that dominant lethals in the mouse and rat are
analogous to those of Drosophila. The much higher rate of induction in

836 RADIATION BIOLOGY

mammals than in Drosophila would be expected from the larger number
of chromosomes. Lea (1947) has speculated on the frequency of chromo-
some breakage that would account for the observed results. The higher
frequency of dominant lethals in offspring of later matings within the
presterile period was attributed by Hertwig (1938a) to fertilizations by
germ cells that had been irradiated in prespermatozoal stages which
were assumed to be more sensitive. Although the observed frequency of
major chromosomal disturbances is adequate to account for most of the
radiation-induced reduction in litter size, it is, however, possible that
dominant lethal, or subvital, point mutations also occur and contribute a
small portion of the cause of death of embryos.

The percentage of stillborn offspring from presterile-period matings of
irradiated males is higher than in the controls in the data of both Snell
(1933b) and Hertwig (1938a). However, both authors believe that much
of this effect is not due to genetic damage in the stillborns, but to diffi-
culties of parturition when litter size is small and the young consequently
bigger at birth. Snell provides evidence for this by showing that the per-
centage of stillbirths is higher for the smaller litter sizes within one dose
group. In guinea pigs, Strandskov (1932) found no significant difference
between presterile-period matings and controls in percentage of still-
births. In the guinea pig, the normal litter size is much lower (2.79 in
Strandskov's material) than in the mouse, so there is presumably less
likelihood of parturition difficulties resulting from a reduction in litter
size. Among the stillborns found by Hertwig and Snell there were, how-
ever, a few pathological cases that could hardly be accounted for by
difficulties in parturition. These indicate that, as would be expected, at
least a portion of the stillbirths are the result of chromosomal aberrations
or dominant point mutations in the affected individuals. It is clear, how-
ever, that, because of the difficulty of distinguishing between internal and
external causes of death, stillbirths are particularly unsuitable material
for the estimation of mutation rates.

Hertwig (1938a) found a significantly higher percentage of death
between birth and 75 days of age in the offspring of presterile-period
matings than in the controls. The percentage of death was greater for
later than for earlier matings within the presterile period. Autopsies did
not reveal the causes of death, but retarded growth was a frequent
characteristic of the animals that died. Strandskov (1932) found a
slight, but insignificant, increase in death in the 0- to 30-day age interval
in progeny of presterile-period matings of irradiated male guinea pigs.
The mean 30-day weight of the survivors was lower than that of the
controls, especially after correction for litter size. This is all the more
striking in view of the fact that the mean 0-day weight of these same
animals, i.e., of those raised to 30 days, was actually, though not sig-
nificantly, higher than the controls in both the uncorrected and cor-

GENETIC EFFECTS IN MAMMALS 837

rected figures. In 102 offspring of matings made within three weeks after
exposure of male rats to doses of from 100 to 1000 r, Henson (1942) was
able to raise only 46 per cent to weaning as against 79 per cent of 96
controls. An effect of radiation on postnatal viability is also indicated in
the few litters allowed to come to term in the work of Amoroso and
Parkes (1947) on irradiation of rabbit spermatozoa in vitro. Thus the
data so far obtained indicate an appreciable manifestation of dominant,
deleterious, postnatal effects in offspring of presterile-period matings of
irradiated males. The relative importance of chromosomal aberrations
and of semilethal and subvital point mutations as causes of the postnatal
mortality is not known.

Offspring of Irradiated Females. Information on the radiation induc-
tion of dominant lethals in females does not go beyond the observation of
a reduction in litter size. Snell and Ames (1£39) state that, as the dose is
increased, litter size in the mouse falls off more rapidly for females than
for males, and they attribute the greater effect to a radiation damage of
the mother which interferes with development of the young. Another
factor that should be kept in mind is that, as the second maturation
division of oogenesis in the mouse is not completed until after fertiliza-
tion, the germ cells in the female are irradiated in oocyte stage. The
results obtained may, on this account alone, prove to be different from
those observed for irradiated sperm.

Offspring of Poststerile-period Matings of Irradiated Males. In contrast
to the marked effect of irradiation of the male on litter size in presterile-
period matings, there is little reduction in litter size in matings made in
the post sterile period (Table 12-7). The germ cells utilized in poststerile-
period matings received the radiation in spermatogonial stages. There is
no reason to doubt that dominant lethals are induced in spermatogonia,
but, as suggested by Strandskov (1932), it is quite likely that many of
them would fail to pass through the cell divisions between spermatogonia
and sperm. Chromosome aberrations of the types which, when induced
in sperm, cause breakdown in early cleavages would seem to be especially
subject to elimination by germinal selection if they are induced in
spermatogonia. That such elimination is occurring is suggested by the
observed reduction in number of spermatogonia following irradiation.
Schinz and Slotopolski (1925), Hertwig (1938b) and Fogg and Cowing
(1952) have reported finding degenerating cells at short intervals after
irradiation. The fact that Eschenbrenner and Miller (1950) found little
evidence of degenerating spermatogonia at later times (one week or more)
after irradiation of mice with 400 r hardly justifies the authors' conclusion
that the effect on the spermatogonia "is not one of cell death but is one
of inhibition of division." There may well be some inhibition of mitosis,
but it is difficult to see how this could result in a reduction in number of
spermatogonia to 1.5 per cent of normal. The results of Eschenbrenner

838

RADIATION BIOLOGY

and Miller perhaps indicate not the absence of killing, but rather the
rapid disappearance of degenerating cells. This would also explain the
fact that the number of degenerating cells found by those investigators
who have reported seeing them is not high enough, at any one time, to
account for the magnitude of the reduction in number of spermatogonia.

Table 12-7. Litter Size in Poststerile-period Matings of Irradiated Males

Reference

Strandskov(1932).

Snell (1933b)

Hertwig (1938a).

Species

Guinea pig
Guinea pig

Mouse
Mouse

Snell and Aebersold
(1937)

Mouse
Mouse
Mouse
Mouse
Mouse
Mouse
Mouse
Mouse
Mouse
Mouse

Radiation

Control
•X rays

Control
X rays

Mouse
Mouse

Russell et ah' ".

Mouse
Mouse

Control
X rays
X rays
X rays
X rays
X rays
X rays
X rays
X rays
X rays

Dose, r

173-2592

600-800

Control
Neutrons

Control
X rays

200

400

500

600

800

1000

1200-1400

1500-1600

Total irrad.

No. of
litters

140
62

24

4

Mean

litter

size

2.69
2.16

7.71
8.00

120-140 "r"6

600

472

5.54

13

5.24

78

5.42

30

5.67

36

4.86

136

5.06

110

5.36

30

5.70

14

4.64

447

5 . 26"

Per cent

of

control

litter

size

80

104

9
4

9,710
12,986

8.44
5.75

5.75

5.58

95
98

102
88
91
97

103
84
95

68

97

" Not given by Hertwig. Calculated as weighted mean of mean litter sizes for
each dose.

b Cyclotron neutrons measured on r scale of Victoreen condenser-type dosimeter.

c Unpublished data collected by W. L. Russell, Josephine S. Gower, Gloria J. Jasny,
Elizabeth M. Kelly, Mary H. Major, and Patricia A. Sarvella. Litter size recorded
three weeks after birth.

The view that spermatogonia are killed is supported by extensive data
showing that cells of other rapidly dividing animal tissues are destroyed
by moderate doses of radiation and that the degeneration occurs at cell
division. It, therefore, seems plausible to attribute at least the major
part of the radiation-induced reduction in number of spermatogonia,
and the consequent sterile period, to dominant cell lethals. If this
interpretation is correct, then dominant lethals are apparently induced

GENETIC EFFECTS IN MAMMALS 839

with greater frequency in spermatogonia than in sperm, for the loss of
spermatogonia for given doses (Table 12-3) is much greater than would
be expected from the reduction in litter size produced by the same doses
given to sperm. Several factors could be involved. In the first place,
the spermatogonia are diploid and the sperm haploid. Furthermore, the
frequency of chromosome breakage, and the proportion of breaks that
result in lethal aberrations may be influenced by the state of the chromo-
somes, the phases of the mitotic cycle in the spermatogonia, and the
oxygen tension or other metabolic characteristics. The total effect of
these possible factors cannot be accurately estimated. All that can be
said is that there is still plenty of latitude for the assumption that more
dominant lethals are induced in spermatogonia than in sperm.

If the reduction in litter size shown in Table 12-7 is real, and certainly
some effect of mutation would be expected, it could be attributed to
dominant lethal chromosomal or point mutations that are not eliminated
by germinal selection, to dominant subvital mutations, or, perhaps, to a
combination of these. The effect is apparently too slight, however, for
easy analysis of the time and nature of death of the missing individuals.

The percentages of stillbirths and of postnatal deaths before 75 days of
age in progeny of poststerile-period matings are slightly, but not sig-
nificantly, higher than in the controls in Hertwig's (1938a) data on mice.
Strandskov's (1932) data on the guinea pig show no difference between
experimentals and controls in postnatal death before 30 days of age, but
the experimentals had a higher percentage of stillbirths and lower 0-day
and 30-day corrected weights. The differences are on the borderline of
significance and conditions that could have affected the 30-day weights
were not exactly matched in experimentals and controls. Some effect,
through the induction of dominant mutations, would be expected, but, as
with litter size, it is apparently too small for easy detection.

Sex Ratio. Death of .embryos from dominant lethals could, through
differential mortality of males and females, result in a disturbed sex
ratio at birth. Certain types of chromosomal aberration, such as loss of
part or perhaps all of the X chromosome, could conceivably upset the
sex ratio without affecting survival. The available data do not present a
consistent picture. Parkes (1925), who exposed male mice to an X-ray
dose below that which produces temporary sterility, reported a barely
significant increase in proportion of males born from matings made from
0 to 4 days after irradiation (59.4 per cent of 133 animals as against 51.6
per cent of 735 controls) and a significant drop in proportion of males
born from 5- to 18-day matings (33.6 per cent of 143 animals). In the
offspring of still later matings (19 57 days), the sex ratio was normal
(54.4 per cent of 217 animals). Hertwig (1938a), however, found, for
male mice exposed to 400-1400 r, no effect on sex ratio at birth in the
progeny of presterile-period matings as a whole and no difference between

840 RADIATION BIOLOGY

the 1- to 8-day and 8- to 1-i-day mating groups. Unpublished data on
the sex ratio at birth from our experiments, tabulated according to the
division into time intervals made by Parkes, show, for male mice exposed
to 600 r, 54.1 per cent males in 218 offspring of 0- to 4-day matings and
54.6 per cent males in 370 offspring of 5- to 14-day matings. Thus
Parkes' results are not confirmed by later work at higher doses. Further-
more, the low proportion of males in the 5- to 18-day mating group in
Parkes' data is hard to explain. An increased death rate of male embryos
can scarcely be the cause, because the litter size is only slightly depressed.
The question, therefore, arises as to whether the disturbance in sex ratio
may have been the result of causes other than irradiation. It is also
possible that the fluctuations were random ones. The probability of
this is not necessarily as remote as the tests of significance indicate,
because the time sequences were apparently chosen on the basis of the
differences shown by them and not on a priori biological grounds.

The sex ratio at birth in the offspring of poststerile-period matings of
mice shows, in the data of Hertwig (1938a), a larger proportion of males
than in the controls in all except the lowest of seven dose groups ranging
from 400 to 1600 r. The difference from the controls is significant for
doses of 1200 r and above. The total number of animals in the irradiated
group was 2240, in the controls 2595. On the other hand, the sex ratio,
recorded at three weeks of age, in unpublished data from our extensive
experiment with exposure of male mice to 600 r shows a slight decrease in
proportion of males in progeny of poststerile-period matings (50.35 per
cent males in 72,472 offspring as against 51.00 per cent in 55,828 controls).

DOMINANT STERILITY

The sterility described in this section is, like the lethality already dis-
cussed, called "dominant" solely because it appears in the Fi of an
irradiated parent. Its dominance precludes transmission to later genera-
tions and consequently makes analysis of its nature difficult. For
dominant lethals, there is, as has already been mentioned, cytological
evidence that chromosomal aberration is at least the major cause. The
cytology of the chromosomes of dominant steriles has not yet been
described. The available information on dominant sterility is limited to
the incidence and to the morphology and histology of the defect.

Data from various experiments with mice on the incidence of dominant
sterility in the offspring of irradiated male or female parents is given in
Table 12-8. Although far more attention has been paid by all investigators
to the partial sterility described in the next section, it is apparent that
the rate of induction of sterility in the offspring of presterile-period
matings of irradiated males is far from negligible. In the male offspring
it may be taken as about 10 per cent for a 700-r X-ray exposure of the

GENETIC EFFECTS IN MAMMALS

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842 RADIATION BIOLOGY

sire. This is nearly one-half the rate of induction of partial sterility at
the same dose level.

The data for presterile-period matings in Table 12-8 show a higher
incidence of sterility in males than in females. The material is too
heterogeneous for an accurate test of significance, but it is noteworthy
that the sex difference is in the same direction in all investigations.

The only data tabulated to show the proportion of steriles produced by
early and late matings within the presterile period of irradiated males are
those of Hertwig (1938a). Of the 10 male steriles listed in that publica-
tion, 8 occurred in 88 offspring of matings made from 1 to 8 days after
irradiation and 2 in 24 offspring of matings made from 8 to 14 days after
exposure.

As shown in Table 12-8, Hertwig's results indicate that radiation-
induced dominant sterility does not appear in the progeny of poststerile-
period matings.

Only scattered information is available on the nature of the sterility.
Hertwig (1935) reports that in one sterile male, spermatogonia and
spermatocytes were present in the testes, but all later stages of spermato-
genesis were lacking and no sperm could be found in the epididymis or
vas deferens. The histological information on the three sterile males
obtained by Snell (1939) in his neutron experiment is as follows: One male
proved to have spermatogonia and spermatocytes, but no spermatids ; one
had a few spermatids, some in a rather advanced stage, but no spermato-
zoa; and the remaining male showed a very few motile sperm in the left
epididymis, a few immotile sperm in the left vas, and no sperm in the
right reproductive tract. Of the two sterile males found by Snell and
Ames (1939) in the offspring of X-irradiated females, one had spermato-
gonia and spermatocytes, but no spermatids or spermatozoa, and the
other lacked testes, although epididymides and vasa deferentia were
present. The sterile female obtained in this experiment was small and
died at seven months of age. The ovaries, which were invaded by a
lymphoblastoma, showed primordial follicles, but no mature ones.

The mutational changes involved in the production of dominant
sterility in mice are not known. Snell (1935) suggested that fragmenta-
tion or deletion of the Y chromosome in irradiated Y-bearing sperm could,
as in Drosophila, account for sterile males. This hypothesis could prob-
ably be tested by cytological examination. Sterility might also result
from damage to the X chromosome in eggs or X-bearing sperm, and this,
too, might be checked cytologically. Some of the animals classified as
sterile may have had chromosomal aberrations, such as multiple trans-
locations, that gave a high proportion of aneuploidy in the gametes.
However, the animals examined histologically were clearly sterile from
causes other than aneuploidy of mature gametes. More information is
needed on the anatomical and histological nature of the sterility in both

GENETIC EFFECTS IN MAMMALS 843

males and females, and on the incidence in the two sexes. Is absence of
testes a common cause and does this occur in offspring of irradiated
males? Whatever the exact nature of the mutational changes, it seems
likely, from the high incidence, that most of the sterility will turn out to
be the result of chromosomal aberrations rather than gene mutations.
The failure to complete the later stages of spermatogenesis, which appears
to be the commonest cause of sterility in males, suggests chromosome
damage of a type that interferes with the maturation divisions. This, in
turn, provides a plausible explanation for the lack of sterility in the
progeny of poststerile-period matings of irradiated males. A mutation
that had such an effect would, if induced in spermatogonia, fail to pass
through to mature sperm.

DOMINANT PARTIAL STERILITY

The radiation induction of hereditary partial sterility in mammals was
reported first by Snell (1933a, 1934, 1935) and shortly afterward by
Hertwig (1935). In Snell's work, male mice were exposed to doses of
from 200 to 1200 r of X rays and mated to nonirradiated females. The
fertility of each of several offspring sired in the presterile period was
tested by repeated matings to nonirradiated animals. The distribution
of the mean sizes of the litters produced proved to be bimodal, whereas
that of the controls was unimodal. The upper mode in the experimental
group corresponded approximately to the mode in the controls, and the
lower mode was at about one-half the litter size of the upper mode.
Snell referred to the animals that consistently produced small litters,
that is, the animals grouped around the lower mode, as "semi-sterile."
The term "partially sterile," adopted by some authors, is used in prefer-
ence here because it avoids any implication as to the extent of reduction
in fertility. Snell showed that the reduction in litter size could be
accounted for by the death of embryos. He found that partial sterility
was transmitted, like a dominant, to one-half of the surviving progeny
of outcrosses of partially sterile animals. He pointed out that the
results fitted the interpretation that the partially sterile animals were
heterozygous for a reciprocal translocation. After laborious linkage
tests, Snell (1941a, 1946) finally achieved genetic proof of this in one
partially sterile line. In this line, the normally unlinked genes a and b
were found to be linked. Cytological confirmation of translocation was
provided by Roller and Auerbach (1941) and Roller (1944) who demon-
strated the association of four chromosomes at synapsis in each of three
partially sterile lines examined. Hertwig (1935, 1938a) confirmed Snell's
early findings and, in addition, showed that there was little or no induced
partial sterility in the offspring of poststerile-period matings of irradiated
males.

844 RADIATION BIOLOGY

The above historical account summarizes the major findings. Detailed
aspects, and some additional results, follow. It should be kept in mind
that most of the partially sterile animals obtained in the experiments to
be discussed were not actually proved cases of translocation and that
some of them were not even tested for transmission of partial sterility to
descendants. Furthermore, if any translocations in heterozygous condi-
tion show considerably more than 50 per cent fertility, these would usually
not have been detected. It is also possible that some of the partially
sterile animals carried more than one translocation. In spite of these
qualifications, however, it seems safe to assume that the frequency of
induction of translocations is represented, in at least a rough way, by
tabulation of partial sterility.

The frequency of occurrence of partial sterility in the offspring of pre-
sterile-period matings of male mice exposed to various doses of X rays is
shown in Table 12-9. Since Snell found no clear-cut cases of transloca-
tion in his controls, and Charles (1950) only two in 2755 animals, it is
likely that in Hertwig's controls, which were not tested for transmission of
partial sterility to their descendants, most, or all, of the observed partial
sterility was not due to translocations, but was simply an expression of
the variation, apparently large in Hertwig's material, of other factors
affecting fertility. Although these factors presumably did not contribute
greatly to the results in Hertwig's irradiated material, some of which, in
contrast to the controls, was tested for transmission, it is possible that
they accounted for some of the recorded partial sterility and perhaps for
more in females than in males. Making allowance for this, the data in
Table 12-9, for all investigations combined, indicate that induced partial
sterility occurs with approximately equal frequencies in the male and
female offspring of presterile-period matings of irradiated mice. The
data are still not sufficient for an accurate analysis of the relation between
frequency and dose. They do, however, show that the rate is much
higher than that for translocations in Drosophila. The yield from a dose
of about 700 r in the mouse is comparable to that from 4000 r in Dro-
sophila melanogaster. A difference of about this magnitude would be
expected from the larger number of chromosomes in the mouse, but
whether or not other factors are involved is not known.

Since, as will be discussed later, the yield of partially sterile offspring
from irradiated oocytes may be lower than that from irradiated sperm, it
might be expected that the yield from irradiated spermatocytes would
also be low. If this assumption is correct and if, as Hertwig has main-
tained, later matings within the presterile period of irradiated males
utilize some sperm that was irradiated in spermatocyte stage, then these
later matings should give a lower percentage of partial sterility than the
earlier matings. The difference between spermatocytes and sperm would
thus be in the opposite direction to that obtained for dominant lethals.

GENETIC EFFECTS IN MAMMALS

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846 RADIATION BIOLOGY

The only data that can be analyzed with regard to this factor are those of
Hertwig (1938a) and she tabulates only the results for partially sterile
male progeny. In these she found 12 partially steriles in 88 offspring
from matings made 1-8 days after irradiation and 2 partially steriles in
24 offspring from the 8- to 14-day matings. As far as these data go, they
are in agreement with the above assumption, but they are obviously not
extensive enough to be considered a satisfactory test.

The information obtained by Snell (1939) on the incidence of partial
sterility in the offspring of presterile-period matings of male mice irradi-
ated with fast neutrons is also given in Table 12-9. As would be expected
from their effect on chromosome breakage in other organisms, neutrons
appear to be more effective than X rays in producing partial sterility, but
it is clear that data obtained with modern methods of irradiation and
dosimetry are needed to establish the ratio of effectiveness.

The incidence of partial sterility in offspring of X-irradiated female
mice was determined by Snell and Ames (1939), as shown in Table 12-9.
Comparison of the result with incidence in offspring of irradiated males is
difficult because there are no data from males exposed at this dose level,
and the data from higher doses are, as has already been pointed out, not
adequate for establishing the frequency-dose curve. However, the inci-
dence of partial sterility in the offspring of females appears to be lower
than what would be expected from males irradiated with the same dose.
Since, in Drosophila, the incidence of translocations in irradiated female
germ cells is much below that in irradiated sperm (Glass, 1940), some
consideration should be given to the possibility that the frequency in
progeny of irradiated female mice may be even lower than the results of
Snell and Ames indicate. The fact that two of the three partially sterile
animals found in the 151 offspring came from the same female suggests
the possibility that this female was herself carrying dominant partial
sterility. As she had only one other offspring that was tested, no definite
conclusion can be reached. This uncertainty, coupled with the limited
total of only three partially sterile animals obtained, still leaves some
latitude for assuming that in mice, as in Drosophila, translocation induc-
tion in females may be much less frequent than in males.

As Hertwig (1935, 1938a) first clearly showed, Table 12-9, there is no
evidence for the occurrence of radiation-induced partial sterility in the
offspring of poststerile-period matings of X-irradiated male mice. This
indicates either that translocations are not induced in spermatogonia or
that they occur and are eliminated by germinal selection. However,
elimination seems unlikely in view of the fact that translocations are
easily transmitted through descendant generations. The hypothesis of
lack of occurrence, therefore, seems to be the more plausible one to
adopt. Since there is evidence, from the great reduction in number of
spermatogonia following irradiation, that chromosome breakage resulting

GENETIC EFFECTS IN MAMMALS 847

in dominant lethals does occur, the absence of translocations is probably
not attributable to lack of breaks. Thus, it appears that it is the process
of segmental interchange which fails to occur.

Using 7 rays from a radium source, Lorenz et al. (1947) found, by
fertility tests, no evidence of translocations in the offspring of male mice
exposed to 8.8 r given in 8 hours per day to a total dose of 1100 r, or in
the offspring of female mice exposed to 8.8 r given 24 hours per day to a
total dose of 770 r. The details of this experiment were presented in an
earlier report by Deringer et al. (1946). In all, forty-two offspring of
irradiated males and thirty-one offspring of irradiated females were tested.
The offspring of the irradiated males came partly from matings made
immediately after removal of the males from the radium field and partly
from later matings, but the proportions and time intervals are not given.
The absence of partial sterility in the offspring of the irradiated males in
this experiment is just what would have been expected from the earlier
work of Snell and Hertwig. For, firstly, even if all the offspring had been
obtained from matings made immediately after removal from the field,
the total dose received in spermatozoal stages, and perhaps in all post-
spermatogonial stages, would hardly have been high enough, even if it
had been given as an acute dose, to produce any translocations in the
number of animals tested. Secondly, although the average dose received
in spermatogonial stages may have been close to the total dose of 1100 r
given, Hertwig had already found no translocations in a slightly larger
sample of offspring of poststerile-period matings of males exposed to
acute doses of from 1200 to 1600 r (Table 12-9). Since the dose of radia-
tion to which the sperm were exposed in the experiment of Lorenz et al.
was not sufficient, even if it had been given as acute radiation, to have
produced any translocations in the sample tested, the data do not answer
the question of whether chronic irradiation of sperm is less effective in
translocation production than acute irradiation, or equally effective, as
would be expected if the chromosome breaks in mouse sperm are like
those in Drosophila sperm and remain open until after fertilization.
Taking SnelPs data on the incidence of partial sterility in the offspring
of females exposed to acute irradiation at face value and comparing them
with the results of Lorenz et al. for irradiated females, it appears that, if
translocations are induced in oocytes, chronic irradiation is less effective.

The only information on the incidence of partial sterility in the descend-
ants of mammals exposed to repeated small doses of high intensity radia-
tion is that provided by Charles (1950). He exposed male mice to 0.1,
0.5, 1.0, or 10.0 r of X rays daily, for 6 days a week, mated them to
unexposed females throughout the weeks of irradiation and tested the
fertility of the offspring. The average accumulated exposures at time of
mating for the different dose levels were 13, 69, 134, and 238 r, respectively,
and 60 r for all dose levels combined, but there was a wide range of

848 RADIATION BIOLOGY

accumulated exposure within each group. Seven partially sterile ani-
mals were found in 3072 tested offspring. This rate does not differ sig-
nificantly from that of two in 2755 found in the controls. It is not
possible in Charles' data to separate accurately the dose received by
spermatozoal stages from that received by prespermatozoal stages, a dis-
tinction which, as has already been shown, is all important. It is prob-
able that any real excess over the controls was due solely to that portion
of the total irradiation which was received by the sperm but, even if this
is assumed, the data are not suitable, or extensive enough, for answering
the question as to whether or not fractionated doses to the sperm are less
effective than a single dose.

It may be concluded from Charles' data and from the work of Snell, who
found no translocations in 196 control mice in all his experiments com-
bined, that the rate of occurrence of spontaneous translocations is quite
low in mice. Charles has not yet presented any detailed information on
the two spontaneous cases reported by him. The only spontaneous
mammalian translocation thoroughly analyzed is one in the rat investi-
gated by Waletzky and Owen (1942), Tyler and Chapman (1948), and
Bouricius (1948). The pattern of fertility reduction, abnormal embryos,
and cytological aberrations described falls well within the range of
variability found for X-ray-induced translocations in the mouse.

Turning to a consideration of the characteristics of the translocations,
the degree of effect on the fertility of the animals that are heterozygous
for a translocation is of first importance. Hertwig (1940) found that the
fertility, expressed as percentage of control litter size at birth, of trans-
location heterozygotes in eleven of her X-ray-induced partially sterile
lines of mice ranged from 42 to 59 per cent. The mean of these lines was
45 per cent. Uterine dissections in the same eleven lines and a few others
showed similar reductions in fertility as measured by number of living
embryos. Hertwig concluded that the differences between lines could
be attributed to chance. The mean fertility of heterozygotes in three
translocation lines of mice studied by Roller (1944) was 30-39 per cent in
one line, 46 per cent in another, and 44-46 per cent in the third. Cyto-
logical analysis showed correlation between reduction in fertility and fre-
quency of nondisjunctional coorientation of chromosomes in the ring-of-
four in first meiotic division. In the six translocation stocks investigated
in detail by Snell (1946), the fertility of heterozygotes ranged from 38 to
62 per cent for females and from 41 to 69 per cent for males. Differences
between lines were significantly greater than could be expected by chance.
In five of the six lines, the percentage fertility of males was higher than
that of females and the difference was significant in two of these lines.
The assumption that the percentage fertility corresponds to the per-
centage of orthoploid gametes is supported by the observed proportion of
normal embryos in one of the lines, and is tenable as part of the explana-

GENETIC EFFECTS IN MAMMALS 849

tion of the results obtained from the matings of partially sterile to par-
tially sterile made within two of the lines. Snell and Picken (1935)
report a presumed translocation in which the fertility of heterozygotes
was almost normal. This case was discovered through the occurrence
of a few abnormal offspring. It should be kept in mind that, as has
already been mentioned, translocations showing considerably more than
50 per cent fertility in the heterozygote would usually not be detected by
fertility tests. Present determinations of the average fertility of trans-
location heterozygotes may, therefore, be biased.

The time of death and nature of the abnormal embryos produced by
the aneuploid gametes in translocation stocks obtained from irradiated
mice have been studied by Snell, Bodemann, and Hollander (1934), Snell
and Picken (1935), Hertwig (1938a, 1940), and Otis (1949). These
investigations show that the commonest time of death is at implantation
or shortly after. There is evidence of some death before this time and
also of some survival even to birth. The proportions falling into these
classes vary considerably among the translocation lines. Snell and his
associates found that failure of the neural groove to close at its anterior
end was a common type of defect in the small proportion of abnormal
embryos that survive to later stages.

The expected proportion of partially sterile animals in the viable off-
spring of a mating of a translocation heterozygote with a normal is one-
half. It would, however, be lower than this if the heterozygotes were
less viable than the normals. On the other hand the proportion of
normals would be depressed if any aneuploid zygotes were viable.
Hertwig (1940), using the mean-litter-size test for partial sterility, found
that the percentage of partially steriles in the offspring of such crosses fell
below 50 per cent in ten out of eleven translocation lines. The mean for
the eleven lines was 45 per cent. In contrast, Snell (1946), who made use
of genetic markers to establish the proportion of partial sterility, found
no significant departure from a 1 : 1 ratio. In all of five sets of his data,
involving three marker genes for one translocation and two marker genes
for another, there was actually a slight, though not significant, excess of
the classes marked by the gene that entered the cross with partial sterility.

Translocations have been obtained in homozygous condition in two, or
possibly three, of six lines tested by Hertwig (1940) and in both of two
lines tested by Snell (1946). The failure to obtain the homozygotes in
Hertwig's other lines could be attributed to limitations of the test rather
than to lethality of the homozygotes. The bearers of the homozygous
translocations proved to be phenotypically normal with the exception of
one of Snell's which showed "a suggestion of a reduced viability." Thus,
as far as they go, the data for the mouse indicate that position effect may
be less important than it is in Drosophila.

The translocations produced by irradiation in mice are proving increas-

850 RADIATION BIOLOGY

ingly useful as tools for genetic and cytological research. Snell (1946)
has attempted a genetic analysis of the frequencies of occurrence of the
nondisjunction classes of gametes found in two translocation lines and
has also obtained some information on the positions of the centromeres on
the two chromosomes involved in one of these lines. Slizynski (1952) has
begun the task of determining, in the mouse, which genetically deter-
mined linkage group corresponds to which cytologically identifiable
chromosome.

DOMINANT VISIBLES

No dominant mutations with externally visible effects were observed by
Snell (1935) in 178 offspring of male mice given a mean dose of 681 r of
X rays (numbers obtained from Snell, 1933b). However, autopsies in
the third generation revealed what proved to be a dominant mutation
affecting spleen shape. Penetrance was incomplete. Affected individ-
uals had markedly reduced vigor and somewhat subnormal fertility.
The autopsied F3 mice were descended from ninety-one offspring of
treated males, but, as the descendant lines were not always large enough
to give near certainty of recovering even dominant mutations, the
mutation rate must be taken as one mutation in something less than
ninety-one sperm for a dose of approximately 700 r.

Charles (1950) found seven dominant visible mutations, including
those detected only by autopsy, in 3072 offspring of male mice exposed to
a mean dose of 60 r. The difference from the controls, in which there
were no mutations in 2755 animals, is significant. Two of the mutant
phenotypes were characterized by abnormal connections of minor tribu-
taries of the vena cava, one had extra nodules of adrenal cortical tissue,
one showed incomplete suture of the parietal bones, one exhibited "con-
fused" behavior occasionally associated with cataract and deafness and
two had altered hair color, one of these also having reduced eye size and
lowered fertility.

Five dominant visible mutations in approximately 30,000 offspring of
poststerile-period matings of male mice exposed to 600 r of X rays are
recorded by Russell (1951) in a preliminary account of an investigation
designed primarily for the detection of recessive mutations at seven
specific loci. The loci were chosen so that recessives could be detected
by rapid examination of the coat color and ears. Dominant mutations,
anywhere in the genome, that had effects on these characters were, there-
fore, automatically detected. In order not to interfere appreciably with
the rapidity of observation necessitated by the large number of animals
required for the determination of mutation rates at the specific loci, the
only additional character routinely observed for dominants was the tail.
For this reason, the mutation rate must be far below the over-all rate to
dominant visibles and is, therefore, of only limited value as an absolute

GENETIC EFFECTS IN MAMMALS 851

rate. It is, however, interesting to compare the results obtained for
dominants with the data on mutations at the specific loci. Two of the
dominants affected the tail, one affected the ears and possibly the coat
color, one produced white spotting and one mottling. Thus, only three
of the dominants affected the characters associated with the specific loci.
In the population in which these now established dominants were found,
thirty-two mutations were observed at the specific loci. It can be con-
cluded that, at least for the coat color and ears, the mutation rate to
dominant visibles at all loci is lower than the mutation rate to recessive
visibles at a total of only seven selected loci.

It was mentioned by Russell (1951) that about one-half of the twenty-
eight induced mutations at the S locus showed, in addition to the recessive
spotting effect, a dominant reduction in body size. With the accumula-
tion of more data on body weights in descendant generations it now
appears that most, or perhaps all, of the *S-locus mutations will show this
effect to some extent. There is also increasing evidence for an associated
reduction in viability which would place these mutations under the
classification of dominant subvitals. In any case, it is apparent that the
mutation rate to dominants detected by a change at the S locus is much
higher than would have been anticipated from the over-all rate to
dominants affecting coat color, ears, and tail. It is possible that dominant
mutations affecting size are far more frequent than those causing coat
color changes or gross morphological abnormalities. It is also possible
that the >S-locus mutations fall in a special class.

RECESSIVE LETHALS, SEMILETHALS, AND VIABLES

The only data on the possible occurrence of radiation-induced sex-
linked recessives in mammals is that presented by Charles (1950) who
briefly reports two sex-linked lethals in 3072 offspring of male mice
exposed to a mean dose of 60 r of X rays and one in 2755 control animals.
The rest of this section is concerned with information obtained on auto-
somal recessives.

Hertwig (1939, 1941) reports two recessive visibles, both causing
retarded growth, and four recessive lethals in the descendants of thirty-six
male offspring of presterile-period matings of male mice exposed to from
600 to 1200 r of X rays. Three visibles, one causing anemia, one oligo-
dactyly (both described in detail in Hertwig, 1942a, b), and one causing
dwarfism, were found in the descendants of eighty-two male offspring of
poststerile-period matings of irradiated males. The dose range, 800-
1600 r, is given only for the first fifty-eight (sixty ?) male offspring tested.
Two other possible lethals are reported as incompletely tested, but it is
not stated whether these came from presterile- or poststerile-period
matings. Two lethals were found in the descendants of seventy-two

852 RADIATION BIOLOGY

control males. In later publications Hertwig (1942a, b, 1944, 1949, 1951)
describes two recessive visibles, shaker-syndactyly and "Kreisler," that
were recovered from presterile-period matings. These are first described
as having come from males exposed to 1000 r and would, therefore, appear
to be additional to the visibles from presterile-period matings reported in
1939 which were recorded as coming from males treated with 800 r.
There seems to be some confusion over doses, however, because shaker-
syndactyly is later described as having come from a male exposed to
1500 r, and the dose given to the male that produced anemia in descend-
ants of a poststerile-period mating is reported as 1500 r in one place and
1000 r in another. Combining offspring of presterile- and poststerile-
period matings, taking the total number of visibles as from five to seven,
assuming that the total number of offspring tested was the 118 reported in
the 1941 publication, and using the weighted mean dose of that portion of
the data for which the doses are recorded, the induced mutation rate to
recessive visibles is from 4.1 to 5.7 X 10~5 per r. All of the four muta-
tions that have been described in detail fall on the borderline between
semilethals and subvitals, and only one has shown any fertility. Taking
the number of lethals as from four to six in 118 tested offspring, assuming
the mean dose used above, and subtracting the control rate, the induced
mutation rate to lethals is from 0.59 to 2.2 X 10-5 per r. Since visibles
with not easily recognizable effects were probably not detected, and since
lethals are difficult to detect with the methods available in mammals, the
above rates are probably much below the over-all mutation rates to
visibles and lethals.

In the investigation by Russell (1951), which has already been men-
tioned in the section on dominant visible mutations, wild-type male mice
were exposed to a single whole-body dose of 600 r of X rays and, after
their period of temporary sterility, mated to a stock homozygous for
seven autosomal recessive visible genes. Control, unexposed, males were
also mated to the test stock. The offspring were observed for mutations
at the seven specific loci. Presumed mutants were saved and breeding
tests made to determine the allelism and the effect of the mutations in
homozygous condition. Thus, the method used detects mutations to
recessives, including those with lethal effect when homozygous, at a
limited number of specific loci. The mutations obtained at each locus
are listed in Table 12-10. Table 12-10 includes the results from a pilot
experiment conducted mainly to determine the relation of dose to survival
and productivity of the exposed animals. Various doses were used in
the pilot experiment and detailed tabulation with regard to this and other
variables was postponed until completion of that experiment.

Taking the total number of mutations in the irradiated group in the
main experiment as fifty-three and subtracting the control from the
experimental rate, the mean induced mutation rate, in spermatogonia, for

GENETIC EFFECTS IN MAMMALS

853

the seven loci is (25.0 + 3.7) X 10-8 per roentgen, per locus. The
observed variation in induced rates at the different loci is significantly
greater than chance variation.

The characteristics of the mutations are of interest from several points
of view. It has already been mentioned that most, or all, of the $-locus
mutations have, in addition to the recessive spotting effect, a dominant
effect causing reduction in body size and possibly lowered viability. No
marked dominant effects have been observed for the other mutations, but

Table 12-10. X-ray-induced Mutations at Specific Loci in the Mouse

(Russell, 1951)

Dose

No. of

animals

examined

Number of mutations at locus

A

B

C

D

P

8

Se

Total

Pilot expe
600 r
Control

^riment"
48,007
37.868

11

1
3-4h

1
6

1

2

8

(3-4") c
25
1

1

8-9

53-54

2

a See text for explanation.

6 Two of the four mutations may be a cluster from a single mutation.
c In the pilot experiment, mutations at the S locus were scored on a smaller sample
of animals than that on which mutations at the other loci were determined.

Table 12-11. Viability of homozygotes of X-ray-induced Mutations at Specific

Loci in the Mouse

Viability of

Tested number of mutations at locus

homozygote

B
2

1

C

D

P

S

Se

Total

Viable

Semilethal

Lethal

2
2

5

3
2

1

2 + °

1

8

7
6 +

a Other mutations at this locus that have been partially tested indicate a consider-
able addition to the lethal category.

the possibility of a slight reduction in viability has not yet been excluded.
Three of the induced mutations, one at the B locus, one at the C locus and
one at the P locus, were to intermediate alleles. The effect of the muta-
tions in homozygous condition is still being tested. The available
information, which includes some not published before, is given in
Table 12-11. The mutations listed do not constitute a random sample
because their inclusion depended on the speed with which the test could
be completed and this, in turn, was affected by the nature of the mutation
and the locus involved. When all the mutations have been tested, the
proportion of viables in the total may turn out to be lower than that in

854 RADIATION BIOLOGY

Table 12-11. The results already indicate that the relative frequencies
of viable, semilethal, and lethal types differ according to the locus. Most
striking is the uniformity of D-locus mutations. Not only are all of the
five tested mutations at this locus semilethal; they also exhibit the same
phenotypic effect: a coat color which is like that produced by the d
allele, but, combined with this, a curious behavior defect characterized by
convulsive fits exhibiting clonus of the limbs and opisthotonos of the body.
Furthermore, this phenotype is indistinguishable from that of the spon-
taneous Z)-locus mutation obtained in this investigation and of the spon-
taneous mutation found by Searle (1951).

The question arises as to how many of the induced mutations scored by
the specific loci test are deficiencies. The following evidence bears on
this question, although it does not provide a definite answer. The fre-
quency of the homozygous lethal effect suggests that at least some of the
mutations may be deficiencies. The occurrence of three mutations that
appear to be intermediate alleles is presumptive, though not conclusive,
evidence of gene mutation in these cases. The most favorable loci for the
detection of deficiencies by phenotypic effect are the D and Se loci which
are closely linked (average crossover percentage 0.16). Of the total of
eight mutations obtained at these loci in the experimental group, none
shows the phenotypic effect that would be expected from a deficiency
involving the linked locus.

HUMAN HAZARDS

The genetic hazards of radiation in man have been discussed by
Haldane (1947), Muller (1950a, b, c; also see Chap. 7, this volume),
Wright (1950), and others. No attempt will be made here to review the
concepts of population genetics used or the resulting estimates of the
hazard, which have been calculated mainly on the basis of mutation
rates in Drosophila. The present section is limited to a brief discussion of
the application of the experimental results that have been presented in
this chapter.

Considering, first, the dominant defects, such as lethality, sterility, and
partial sterility, that appear in the offspring following irradiation of the
later stages in spermatogenesis, it is well established that the rate of
induction of these is much higher in mice than in Drosophila. This would
be expected if most of them are due to major chromosomal aberrations,
for the haploid number of chromosomes in the mouse is twenty, while in
Drosophila melanogaster it is four. It is reasonable to suppose that, with
twenty-four chromosomes in man, the rate in man might be somewhat
higher than that in the mouse. Some of the effects, for example, death of
early embryos caused by dominant lethals or by aneuploid gametes from
translocation heterozygotes, might bring little or no distress, and might
pass unnoticed as individual occurrences, but the rate of induction of those

GENETIC EFFECTS IN MAMMALS 855

dominant defects, such as complete sterility and lowered viability in
postnatal or late prenatal life, which are important at the individual
level, is far from negligible. Fortunately, there is ample evidence to show
that the high incidence in the offspring is found only after irradiation of
later germ-cell stages. It is clear that if the gonads of a man are exposed
to a considerable dose of radiation at one time, or within the few weeks
required for a spermatogenic cycle, the chance of transmission of the
mutational changes responsible for these defects can be greatly reduced
by abstention from fertile matings for a period of a few weeks following
exposure.

Turning to a consideration of point mutations and small deficiencies or
other minor chromosomal changes, it must be remembered that the ratio
of induced mutation rates in spermatogonia and spermatozoa for these
types of mutation has not been measured in mammals. However, from
results on Drosophila, it seems likely that the practice recommended above
for avoiding the transmission of major chromosomal changes would
result in much less of a reduction in the probability of transmitting point
mutations and minor chromosomal changes. If this is true, then, for
these mutations, it is more important to know the induced mutation rate
in spermatogonia than that in spermatozoa, because most of the total
dose received in the ontogeny of a spermatozoon in man will usually have
been accumulated in the spermatogonial stage.

The induced mutation rate in spermatogonia for mutations with clear-
cut dominant effects has not been adequately determined in mammals.
The results of Charles (1950) indicate that the rate to dominant visibles
may be appreciable, but, in his data, the effects of irradiation on early
and late germ-cell stages cannot be separated. In the investigation by
Russell (1951), irradiation of spermatogonia gave a low over-all mutation
rate to dominants affecting coat color, tail, and ears, but a high rate at
one locus to dominants causing reduction in body size and possibly
lowered viability.

The reduction in litter size in litters from poststerile-period matings
of irradiated males (Table 12-7) may prove to be useful in estimating the
total effect from the induction in spermatogonia of dominants that cause
mortality in early development. This effect is apparently not large
enough for easy measurement by a small sample, because of the presence
of variation in the many biological factors that influence litter size, but
the extensive data now being collected may prove informative when
fully analyzed.

The specific-loci mutations induced in spermatogonia in the mouse
(Russell, 1951) have not yet been thoroughly investigated for deleterious
effects. However, the majority of those tested have proved to be lethal
or semilethal in homozygous condition (Table 12-11). Work on Dro-
sophila has shown that most recessive lethals have a deleterious effect on

856 RADIATION BIOLOGY

viability even in heterozygous condition (see Chap. 7, this volume). It,
therefore, appears likely that mutations induced in spermatogonia in
mammals may have appreciable dominant deleterious effects which, in
man, will prove to be more of a hazard than the recessive effects.

The mean induced mutation rate for irradiated spermatogonia in the
experiment of Russell on specific loci in the mouse is (25.0 + 3.7) X 10-8
per roentgen, per locus. This is considerably higher than the rates found
in similar experiments on Drosophila and indicates that estimates of human
hazards based on Drosophila mutation rates may have to be revised.

In the same investigation, the observed rate of mutation was not
appreciably dependent on the length of interval between irradiation and
fertilization. It, therefore, appears that, as far as mutations induced in
spermatogonia are concerned, postponement of procreation would be
ineffective in reducing the probability of transmission.

The discoveries (reviewed by Hollaender, Baker, and Anderson, 1951)
that hypoxia and other treatments protect against the induction by
radiation of chromosomal damage, and perhaps other genetic changes,
has increased the hope that protective agents will be found that will
reduce the genetic hazard of radiation in man. The first test of the effect
of hypoxia on radiation induction of genetic damage in mammals has,
however, proved discouraging. No protection against the X-ray induc-
tion of dominant lethals in sperm in mice was found (Russell et at., 1951).
It is possible, of course, that hypoxia will prove to afford protection
against genetic damage in earlier germ-cell stages, or even against some
types of genetic damage in sperm.

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Eschenbrenner, A. B., E. Miller, and E. Lorenz (1948) Quantitative histologic
analysis of the effect of chronic whole-body irradiation with gamma rays on the
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Falconer, D. S. (1949) The estimation of mutation rates from incompletely tested
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Fogg, L. C, and R. F. Cowing (1951a) The changes in cell morphology and histo-
chemistry of the testis following irradiation and their relation to other induced
testicular changes. I. Quantitative random sampling of germinal cells at inter-
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and (1951b) The changes in cell morphology and histochemistry of

the testis following irradiation and their relation to other induced testicular
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Cancer Research, 11: 81-86.

and (1952) Post-irradiation studies on mammalian testes. Effect at

hourly intervals for first 24 hours. Proc. Soc. Exptl. Biol. Med., 79: 88-92.
Glass, H. B. (1940) Differential susceptibility of the sexes of Drosophila to the

effect of X-rays in producing chromosome aberrations. Genetics, 25: 117.
Gliicksmann, A. (1947) The effects of radiation on reproductive organs. Brit. J.

Radiology, Suppl. 1, pp. 101-109.
Haldane, J. B. S. (1947) The dysgenic effect of induced recessive mutations. Ann.

Eugenics, 14: 35-43.
Henson, M. (1942) The effect of roentgen irradiation of sperm upon the embryonic

development of the albino rat (Mus norvegicus albinus). J. Exptl. Zool., 91:

405-434.
Hertwig, P. (1932) Wie muss man ziichten, um bei Saugetieren die natiirliche oder

experimentelle Mutationsrate festzustellen? Arch. Rassen- u. Gesellschaftsbiol.,

27: 1-12.

(1935) Sterilitatserscheinungen bei rontgenbestrahlten Mausen. Z. indukt.

Abstammungs- u. Vererbungslehre, 70: 517-523.

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Rontgen-bestrahlung von Spermatogonien, fertigen und unfertigen Sperma-
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bestrahlung, unter besonderer Beriicksichtigung der Spermatogonien. Arch,
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- (1939) Zwei subletale rezessive Mutationen in der Xachkommenschaft von
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(1940) Vererbare Semisterilitat bei Mausen nach Rontgenbestrahlung,

verursacht durch reziproke Chromosomentranslokationen. Z. indukt. Abstam-
mungs- u. Vererbungslehre, 79: 1-27.

(1941) Erbanderungen bei Mausen nach Rontgenbestrahlung. Proc. Intern.

Genetic Congr. 7th Congr. Edinburgh, 1939, J. Genetics Suppl., pp. 145-146.

(1942a) Neue Mutationen und Koppelungsgruppen bei der Hausmaus.

Z. indukt. Abstammungs- u. Vererbungslehre, 80: 220-246.

(1942b) Sechs neue Mutationen bei der Hausmaus in ihrer Bedeutung fur

allgemeine Vererbungsfragen. Z. menschl. Vererbungs- u. Konstitutionslehre,
26: 1-21.

— (1944) Die Genese der Hirn- und Gehororganmissbildungen bei rontgen-
mutierten Kreisler-Mausen. Z. menschl. Vererbungs- u. Konstitutionslehre,
28: 327-354.

858 RADIATION BIOLOGY

(1949) Untersuchungen liber die Taubheit bei einem Stamm von rontgen-

mutierten Mausen, den "syndactylen Schiittlern." Proc. 8th Intern. Congr.
Genetics, Stockholm, 1948, Hereditas Suppl., pp. 592-593.

(1951) Entwicklungsgeschichtliche Untersuchungen iiber Bewegungs-

storungen bei Mausen. Verhandl. anat. Ges., 49: 97-107.

and H. Brenneke (1937) Die Ursachen der herabgesetzten Wurfgrosse bei

Mausen nach Rontgenbestrahlung des Spermas. Z. indukt. Abstammungs- u.

Vererbungslehre, 72: 483-487.
Hollaender, A., W. K. Baker, and E. H. Anderson (1951) Effect of oxygen tension

and certain chemicals on the X-ray sensitivity of mutation production and

survival. Cold Spring Harbor Symposia Quant. Biol., 16: 315-326.
Howard, A., and S. R. Pelc (1950) P32 autoradiographs of mouse testis. Preliminary

observations of the timing of spermatogenic stages. Brit. J. Radiology, 23:

634-641.
Roller, P. C. (1944) Segmental interchange in mice. Genetics, 29: 247-263.

and Charlotte A. Auerbach (1941) Chromosome breakage and sterility in

the mouse. Nature, 148: 501-502.

Lea, D. E. (1947) Effects of radiation on germ cells: dominant lethals and hereditary
partial sterility. Brit. J. Radiology, Suppl. 1., pp. 120-141.

Lorenz, E., W. E. Heston, A. B. Eschenbrenner, and Margaret K. Deringer (1947)
Biological studies in the tolerance range. Radiology, 49: 274-285.

Muller, H. J. (1950a) Radiation damage to the genetic material. Am. Scientist,
38: 33-59, 126, 399-425.

(1950b) Some present problems in the genetic effects of radiation. J.

Cellular Comp. Physiol., 35 (Suppl. 1): 9-70.

(1950c) Our load of mutations. Am. J. Human Genetics, 2: 111-176.

Otis, E. M. (1949) Intra-uterine death time in semi-sterile mice. Anat. Record,
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Parkes, A. S. (1925) The effects on fertility and the sex-ratio of sub-sterility expo-
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Regaud, C, and G. Dubreuil (1908) Perturbations dans le developpement des oeufs
fecondes par des spermatozoides Roentgenises chez le lapin. Compt. rend. soc.
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Russell, W. L. (1950) The incidence of sterility and partial sterility in the descend-
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(1951) X-ray-induced mutations in mice. Cold Spring Harbor Symposia

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(1933a) Genetic changes in mice induced by X-rays. Am. Naturalist,

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■ (1933b) X-ray sterility in the male house mouse. J. Exptl. Zool., 65:
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— (1934) The production of translocations and mutations in mice by means of
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— (1939) The induction by irradiation with neutrons of hereditary changes in
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— and F. B. Ames (1939) Hereditary changes in the descendants of female mice
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Manuscript received by the editor June 6, 1952

CHAPTER 13

The Effects of Radiation on Mammalian
Prenatal Development1

Liane Brauch Russell
Biology Division, Oak Ridge National Laboratory

Introduction. Experimental findings: The ^reimplantation period — The period of
major organogenesis; Mortality, Body size, Sex ratio, Morphology — The period of the
fetus; Mortality, Morphology, Time of parturition. Isotope studies. Mechanisms of
radiation effect on the embryo: Influence of the maternal organism — Nature of the primary
damage and intermediate effects — Dosage relations; variability — Comparison with other
agents affecting development. Clinical literature on the effects of radiation on embryo
and fetus; human implications of experimental work. Summary. References.

I. INTRODUCTION

Studies on the effects of radiation on mammalian prenatal development
are of special interest not only because of their human and medical
implications but by virtue of their contributions to the fields of mam-
malian experimental embryology and of developmental mechanics in
general.

Certain inherent peculiarities in the biological material set it apart
from the more conveniently studied invertebrates and lower vertebrates.
Since the mammalian embryo develops within the body of its mother and
cannot, in general, be irradiated without irradiating some maternal tis-
sues, the question of how much of the damage to the conceptus is direct
and how much is caused indirectly through damage to the mother must
always be in the background of an investigation and was, in fact, almost
the sole subject of much of the early work. The dependent nature of
mammalian development is, however, of some advantage for embryological
studies since it probably permits abnormal development to proceed to
more advanced stages than it would in the unprotected embryos of other
forms. Since the embryo is usually not observed at the time of irradia-
tion, its developmental stage can be designated only by its chronological
age, which — because of variations in rate between litters (especially of
different genetic backgrounds) and even within litters — provides only an

1 Work at Oak Ridge and preparation of manuscript were performed under Con-
tract No. W-7405-eng-26 for the Atomic Energy Commission.

861

862 RADIATION BIOLOGY

approximate measure. Unfortunately, the importance of controlling
even the postconceptional interval has eluded some investigators
altogether.

For a few reasons, then, the mammal is not the most suitable material
for investigating the effects of radiation on development. Conversely,
however, radiation is an excellent tool for approaching mammalian
embryology from an experimental point of view. As opposed to other
deleterious agents (e.g., injected poisons), radiation does not have to
"pass the placental barrier" and it reaches the embryo in calculable quan-
tities. Timing of treatment is not complicated by the unknowns existing
for other agents (e.g., how soon effective concentrations are built up in
the circulation or how rapidly the poison is destroyed or eliminated by
the organism). Finally, since the action of radiation has a general dis-
tribution throughout the organism, selective response of structures may
be expected to indicate patterns of sensitivity intrinsic to the embryo.

Work on the effects of radiation on mammalian prenatal development
will thus be reviewed from the points of view of (1) the contributions to
mammalian experimental embryology and developmental mechanics,
and (2) the human implications of the existing experimental material
supported by pertinent clinical findings.

The early investigations in the field were, unfortunately, not well con-
trolled, not only with respect to radiation factors, as might be expected,
but especially as regards timing of developmental stages. They, therefore,
do not contribute substantially to point (1) except for scattered incidental
information. For this reason, they will not be discussed in detail but are,
for the sake of completeness, included in the summarizing tabular mate-
rial (Tables 13-1, 2, 7).

II. EXPERIMENTAL FINDINGS

Although it would be most profitable to discuss separately the results of
irradiating each of the conveniently landmarked stages from conception
to birth, only a very small number of past investigations is suitable for this
type of analysis, and rougher groupings must, therefore, be made. Con-
venient points of division can be derived from a survey of the entire
gestation period of the mouse (Russell, 1950) in which irradiated stages,
differing by 24-hour intervals, ranged from day }4 postfertilization to
near term. The three broad phases which emerged from this study were
as follows: (1) irradiation during the preimplantation period gave a high
incidence of prenatal death but almost no abnormalities in survivors to
term; (2) irradiation with the same dose during the period of major
organogenesis yielded a high incidence of abnormalities at birth but much
less prenatal mortality; and (3) irradiation during the fetal period
(growth, minor organogenesis) did not cause prenatal death, and no gross

RADIATION IN PRENATAL DEVELOPMENT 863

abnormalities at birth. It should be pointed out that the division points
do not represent sharp demarkations but merely separate average
response for entire periods. Appropriate groupings of the work of other
authors support, in general, these broad divisions of prenatal development
with respect to response to radiation. Each division will, therefore, be
taken up in turn.

An attempt at complete coverage of the literature has been made. The
experimental material consists almost entirely of mice, rats, guinea pigs,
and rabbits. In the publications here reviewed, the number of investi-
gators, or groups of investigators, concerned with each of these is 11, 9, 8,
and 15, respectively. Incomplete data are available on two cats (Tous-
sey, 1905; Schinz, 1923) and one dog (Regaud et al., 1912). Only the
mouse and rat have been used in recent experiments, all work with other
species having been published before 1935. In order to determine broad
developmental divisions, similar to those described in the preceding
paragraph, for the species to be discussed, landmarks in development
(e.g., blastocyst, first mesoderm, neural plate, first somite, etc.) were
entered against coordinates representing days of gestation of mouse and of
the animal to be compared with it. It is thus possible to arrive at the
following approximate divisions for preimplantation period, period of
major organogenesis, and period of the fetus, respectively: mouse — 0-5,
6-13, 14 to term (usually days 19 or 20) ; rat — 0-7, 8-15, 16 to term (usu-
ally days 21 or 22) ; rabbit — 0-5, 6-15, 16 to term (usually days 31 or 32) ;
guinea pig — 0-8, 9-25, 26 to term (approximately 9 weeks; but it must
be remembered that birth here does not occur at a comparable stage, a
newborn mouse corresponding, approximately, to a 34-day guinea pig
fetus) .

Whenever possible, work to be reviewed has been placed in a category
defined by the animal with which it deals and the time in gestation at
which irradiation was applied (see Tables 13-1, 2, 7). A few publica-
tions cover two or several categories and are, therefore, listed more than
once. In many cases the stage is only vaguely stated (e.g., "second half
of pregnancy"), in others, although not stated at all, it can be approxi-
mately calculated from incidental information (e.g., by counting back
from term). In still other cases, where a treatment interval is reported
(either because the irradiation was chronic or because the author is not
specific about the date of his acute exposure), there may be overlap into
an adjacent broad developmental phase. For these and other reasons,
Tables 13-1, 2, 7 may be considered only as organized listings of litera-
ture and must not be evaluated as a summary of results. It was felt
that this was the most profitable way of presenting the extremely hetero-
geneous group of investigations, many of which are not definitive by
themselves but may add to the picture as a whole. Not included in the
tables are experiments in which no clue can be obtained of the stage

864 RADIATION BIOLOGY

irradiated (Murphy and de Renyi, Levine, Finkel, von Klot, Forsterling,
Linser and Helber).

In almost all of the experiments, irradiation was administered from an
external source. The whole pregnant female was usually exposed,
although several investigators attempted to direct the beam to the abdom-
inal region. Shielding of extra-abdominal regions was part of the tech-
nique in experiments by Warkany and Schraffenberger (1947), Job et al.
(1935), Kaven (1938a, b), Murphy and de Renyi (1930), and Regaud
et al. (1912). Wilson (1949), and Wilson and Karr (1950, 1951) went so
far as to expose only selected embryos. Finally, Raynaud and Frilley
(1943a, b, 1947a, b, c, 1949a, b) used a beam of 0.5-3.0 mm diameter to
irradiate only selected parts of certain embryos. Radioactive isotopes,
Sr89 and P32, were injected into the pregnant female by Finkel (1947) and
by Burstone (1951), respectively. Bagg (1922) and Gudernatsch and
Bagg (1920) injected a solution of sodium chloride which had been
exposed to 500 mc of radium emanation to obtain an "active deposit."

A. THE PREIM PLANTATION PERIOD

The rather scanty work on the preimplantation period is summarized
in Table 13-1. It may be noted, first of all, that none of the publications
report abnormalities while all report prenatal death. This result is par-
ticularly meaningful in the case of investigators whose work has also
extended into other periods. Thus Job et al. (1935), who obtained
abnormalities from the irradiation of later stages, report only resorption
as a result of 0.8 skin erythema dose (hereafter to be abbreviated to
SED)2 delivered during the preimplantation period. The survivors were
normal. The similar contrast found by Russell (1950) in a survey of the
gestation period has already been mentioned. Kosaka (1928c) states
that % SED delivered during the preimplantation period to guinea pigs
has either no effect at all or kills the embryo. The normality of survivors
from very early irradiation (mice or rats?) extends even to normality of
postnatal development (Kosaka, 1928e). Parkes obtained litters from all
of ten females irradiated in the second and third weeks of gestation, but
only two out of thirteen irradiated during the first week carried to term.

2 The SED is defined (Ellinger, 1941) as the quantity of radiation administered
at 23 cm focal skin distance, 180 kv, 0.5 mm Zn + 3 mm Al filter, and a field of
6X8 cm. This is equivalent to 600 r in air. However, the radiation factors in
most of the experiments using SED dosimetry differ from the standard conditions
enumerated and, since the r equivalent to 1 SED varies considerably depending on
hardness of radiation and other factors, the practice throughout this review has been
to cite the dose exactly as stated by the author. This applies also where other
systems of measurement have been used, such as the Holzknecht unit, or the pastille
tint method. It has been left to the reader to make attempts at approximate conver-
sion to r units in individual cases where it might be of interest and where the original
publication may be consulted for details on radiation factors.

RADIATION IN PRENATAL DEVELOPMENT

865

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866 RADIATION BIOLOGY

In view of recent results to be discussed in more detail, it is of interest
to extract from the literature what information is available as to the stage
in gestation when death occurs. In five investigations (Burckhard,
Parkes, Russell, de Nobele and Lams, Driessen), in which the uterus was
examined within two weeks after irradiation, there is mention of finding
no signs of pregnancy in several of the females. This means either that
death occurred before implantation or that fertilization failed to occur.
Radiation was not administered early enough in any of the experiments
to have interfered with fertilization, although Burckhard (who began his
treatments earlier than most investigators) suggests that killing of sperm
in the female is responsible for nonpregnancy. Since a certain proportion
of matings is apparently always nonfertile, the early experiments, which
lack adequate controls for the determination of this proportion, provide
only suggestive evidence of preimplantation death due to early irradia-
tion. Somewhat more direct evidence comes from Kosaka (1928c) who
opened, within 72 hours after irradiation, three of fourteen guinea pig
females irradiated during the first week after conception and found dis-
turbances in implantation of embryos that had died earlier. Burckhard
(1905) reports retarded cleavage, which may be an indication of disturb-
ances leading to later death, some of it conceivably before implantation.
Finally, there are reports (Trillmich, 1910; Parkes, 1927) of finding no
sign of pregnancy externally, although here there also exists the possibility
of early postimplantation death.

Ample evidence for death after implantation is presented by all investi-
gators except Burckhard, whose observations do not extend far enough
into that period. The usual finding consists of resorbing bodies or
decidual rests in the uterus, but two authors (Trillmich, 1910; Parkes,
1927) also mention abortion in later stages of pregnancy. A few reports
of arrested development (de Nobele and Lams, 1925; Driessen, 1924), are
probably to be classed with more obvious cases of resorption, since there
are indications that the embryos may have died shortly before
observation.

Following up the indications of the survey experiment (Russell, 1950)
in which irradiation during preimplantation stages had reduced the
number of litters at term (see Table 13-3), as well as the litter size in
surviving litters (see Table 13-4), Russell and Russell (1950a) irradiated
a separate extensive series of females }4, ^XA, ^hz, 3H, or 4^ days after
mating. A control was handled simultaneously with each irradiated
female. In an attempt to determine at what stages preimplantation
death occurred and whether any abnormalities were expressed in animals
thus weeded out before term, all uteri were examined either 10^2 or 13^2
days after mating, i.e., 6-13 days after irradiation. It was found that
radiation-induced death was considerable and that, in over-all effect, the

RADIATION IN PRENATAL DEVELOPMENT

867

earliest stages were most sensitive, the average number of living embryos
per treated female being only about 20 per cent of the controls in groups
irradiated on days Y2, \\i, or 2l/2, but 31 and 57 per cent following treat-
ment on days %y2 and 4y2, respectively (Fig. 13-la). Mortality between
days 103^ and 13>£ and between days 13>2 and term was no higher than
in controls and radiation-induced death, therefore, occurred entirely
before day 10>2, m fact, so long before that stage that abnormalities, if

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YIELD 0FL1VE EMBRYOS PER
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A A IMPLANTS PER IRRADIATED FEMALE

•-- -• LIVING EMBRYOS PER CONTROL
A- — A LIVING EMBRYOS PER IRRADIATED

FEMALE
SPACE BETWEEN SOLID LINES REPRESENTS PRE-
IMPLANTATION DEATH OF INDIVIDUALS

Fig. 13-1. Effects of irradiation with 200 r during the preimplantation stages of the
mouse embryo. All observations are made on days 10}-^ or 13} 4 postcopulation, and
points are based on a total of 521 implants in the controls, 310 in the irradiated groups.
Fig. (b) based on pregnant females only. (Russell and Russell, 1950a.)

any were produced, could no longer be recognized at the time of observa-
tion. More specifically, three groups of deaths can be recognized: (1)
Preimplantation death of entire litters, i.e., very early termination of
pregnancy, is indicated by the excess of nonpregnant females at the time
of dissection (see Fig. 13-la). The percentage of copulations resulting in
implants observable on days 10^ and 13>^ is 79 in controls and 44-67 in
different irradiated groups (average, 56 per cent). As a whole, the reduc-
tion is highly significant but fluctuations among the different irradiated
groups are probably random. Therefore, on the average, about 29 per
cent of the embryos irradiated in preimplantation stages are lost in whole-

868 RADIATION BIOLOGY

litter death prior to implantation. (2) The balance of the mortality is
due to death of individuals within surviving pregnancies (see Fig. 13-la).
This accounts for about two-thirds of the total death in groups irradiated
on days x/i, l}4, and 23^, but by day 4^, when over-all mortality has
greatly decreased, death of individuals has become only a minor portion
of the total loss. Further analysis of death of individuals (see Fig.
13-16, based on pregnant females only) indicates that it may occur in
one of two periods: (a) Animals may die before implantation, as shown by
deficiency in total implants in irradiated pregnant females. This type of
death occurs mostly in embryos irradiated on days % and 1%, eliminating
respectively 3.3 and 1.9 implants per surviving pregnancy. By day
43^, irradiation no longer causes preimplantation death of individuals.
(b) Embryos may die after implantation, as shown by the deficiency in
living implants. Further work is needed to test the present indications
of survival being lowest following irradiation on day 23^.

It can be argued that, since preimplantation death involving whole
litters is of approximately equal importance in all groups, the majority of
it may not be caused by direct radiation injury to the early embryo but,
instead, by some effect on the implantation processes of the mother.
Individual preimplantation death, on the other hand, is high only as a
result of irradiating precleavage or very early cleavage stages. Post-
implantation death is greatly increased following irradiation before
implantation but its exact relation to developmental stage irradiated can
be elucidated only by further work.

Assuming that the probability of killing a blastomere with a given dose
of radiation is equal in the one-cell stage and later cleavage stages, it can
be calculated from the incidence of death of embryos that the group of
those surviving radiation on days 13^-43^ probably includes some in
which one to several blastomeres were killed. Since virtually all sur-
vivors from irradiation of cleavage stages are normal, the results, as they
stand, indicate a considerable degree of totipotency in the blastomeres of
the young mammalian embryo.

B. THE PERIOD OF MAJOR ORGANOGENESIS

The bulk of the work on the effects of radiation on the mammalian
embryo deals with the period of major organogenesis and most of the
more careful and extensive studies fall into this group. Table 13-2, which
presents a condensed summary of the literature, immediately reveals the
almost universal discovery of abnormalities following irradiation during
that period. The interest in mortality is, in general, only secondary.
Only in the rabbit is there a dearth of reported abnormalities, but this
may be due to the fact that all the rabbit work was done early in the
century and much of it was not critical with respect to experimental
procedure.

RADIATION IN PRENATAL DEVELOPMENT

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RADIATION BIOLOGY

1. MORTALITY

a. Prenatal. Mortality is reported by almost all investigators, at least
for the higher dose series. For doses comparable to those which were
used during the preimplantation period by investigators whose work
spans both periods, prenatal death, however, is of decreased importance
(Job et al., 1935; Russell, 1950; Bagg, 1922; Parkes, 1927). Russell
(1950) showed that complete interruption of pregnancy, which often
follows irradiation during the preimplantation stages, occurs only rarely
as a result of later irradiation. This is obvious from the almost 100 per

Table 13-3. Percentage of Copulations Resulting in Pregnancy in Controls

and Following Irradiation with Different Doses at Different Stages in

the Prenatal Development of the Mouse

Dose, r

0
100
200
300
400

0
200

Observation

Term"
Term"
Term"
Term"
Term"

10^, lSy2 PC"

ioy2, 13^ PCb

Yield of litters following treatment on postcopulation day

Mr^i

5H-8H

Pregnancy not diagnosable

No. 9 9

Per cent

No. 9 9

treated

w. litter

treated

18

72.2

18

15

46.7

1

20

45.0

29
9

77

79.2

77

55.8

Per cent
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72.2
100
69.0
0.0

w-mi

Pregnancy
diagnosable

No. 9 9
treated

17

19

4

Per cent
\v. litter

94.1

94.7

100.0

"Russell, 1950.

6 Russell and Russell, 1950a.

PC = postcopulation.

cent yield (see Table 13-3) from females treated with 200, 300, or 400 r
in stages when pregnancy can already be diagnosed externally (days 9^2
and 10^2 usually; days 11^ through 13^2 invariably) and no less appar-
ent from comparison with controls of those females irradiated with
200 r at a time when it is unknown whether copulation was fertile (days
5}^-S}4)- The question of whether there is prenatal death of individuals
within litters, as judged by decrease of average litter size at term, is more
difficult to answer since, with the necessarily limited numbers available
after subdivision into stage and dose groups, standard errors are large.
To diminish this difficulty, data accumulated at this laboratory from
several morphological investigations, in which litter size was only an
incidental result, have been pooled in Table 13-4 to give a total popula-

RADIATION IN PRENATAL DEVELOPMENT

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872 RADIATION BIOLOGY

tion of 419 litters (2667 young). (It must be borne in mind that the
table includes several genetic strains — distributed unevenly among groups
— with probable differences in radiation response; see a future section.
It is, therefore, suitable only for showing broad trends.) For irradiation
of postimplantation stages, doses of 100 r and below have apparently no
effect on prenatal viability. A dose of 200 r, which, it may be remem-
bered, caused 80 per cent prenatal death when applied days x/i, 1}4,, or 2^
after fertilization, permits an over-all average litter size of 5.94 ± 0.27
when administered between days 5^ and 12^ inclusive, i.e., causes a
decrease of only 11 per cent from the control mean of 6.69 ± 0.18
(t = 2.43). This over-all slight depression is due primarily to the sig-
nificantly reduced litter sizes of groups irradiated near the beginning of
the period, namely, on days 63^, 7^2> and 8^2 (reduction for these three is
27 per cent; t = 4.46). The 300-r data substantiate the 200-r series in
again showing sensitivity to prenatal killing of individuals only in the
early part of the period of organogenesis.

The findings of other investigators who have worked on the mouse are
in general agreement with the above results. Kaven (1938b) obtained
litter sizes of 4.3 and 4.7 following irradiation with 178 r to the uterus on
days 7 and 8, respectively. The control litter size was 6.8 (see Table
13-6). Litter sizes for irradiation of later stages cannot be accurately
calculated from his data (1938a), but where estimates are possible these
are usually considerably higher than the results from the 7- and 8-day
stages. Kosaka (1927) found that doses greater than j^ SED would give
100 per cent prenatal death if given between the fourth and tenth days
while more than 1 SED was required to accomplish the same result when
administered after the tenth day.

Wilson and Karr (1951) concentrated their study on irradiation at day
10 after fertilization in the rat and examined for dead embryos 1-5 days
following irradiation. Their results for the last observation day are
shown in Table 13-5. These figures and the summation for all observa-
tion days indicate that while 50 r does not increase the percentage of
prenatal death at all and 100 r not significantly so (t = 1.4 for the sum of
all observation days), the significant (t = 5.3) killing action of 200 r
should result in at least 88 per cent reduction in average litter size at
birth (more if, as is likely, there is further death between day 15 and
term). For approximately comparable stages in the mouse (days 7^ and
83^) and the same dose, the reduction in average litter size at birth is
only 29 per cent. It is of interest in Wilson's data that while mortality
due to 200 r is spread over a 4-day interval beginning 2 days after irradia-
tion, all death with 400 r occurs within 24 hours of exposure. In con-
trast, Kosaka (1928b), in his work on rats, found that even doses which
kill 100 per cent of the embryos— i.e., % SED and above on any of days
5-10 — do not cause death until 48 to 240 hours after irradiation. In an

RADIATION IN PRENATAL DEVELOPMENT

873

abstract, Wilson and Karr (1950) mention that 200 r applied on day 9 will
kill all embryos, which would indicate that stage to be even more critical
for prenatal viability than day 10.

While it is tempting to compare the results of Wilson and those of
Russell with a view to demonstrating a probable difference between rat

Table 13-5. Viability after Prenatal Irradiation in the Rat: Comparison of

the Results of Different Authors

Senior
author

Job, 1935
Wilson, 1950
Wilson, 1951

Job, 1935

Warkany, 1947
Warkany, 1947
Warkany, 1947
Job, 1935

Warkany, 1947
Warkany, 1947
Warkany, 1947
Warkany, 1947

Day

irrad.

9th- 15th

9

10

8th-llth

9rf
10
11
12th-16th

12
13
14
15

Dose, r

50

100 200

300

400

500

600 700

800

900

1000

1100

1200

Percentage of embryos or fetuses alive"

0
89(37)<> 100(2) 89(9) 12(8)

0(10)

Percentage of 9 9 with litter at term — average litter size'

I I

?%— 8.2 ?%— 7.4(237)
(41)

50%— 4.9(44)

79%— 3.0(45)

92%— 4.9(54)

?%— 11.0 ?%— 8.4(101)
(22)

72 %— 5. 2(249)

100%— 6.3(50)

92%— 7.4(81)

J

75%— 7.5(45)

" Observations recorded in this portion of the table were made as follows: in the work of Job el al., on day 18; in the experi-
ments of Wilson and Karr, on day 15 for 0-200 r, day 12 for 400 r.

b Figures in parentheses indicate the number of young on which percentage alive or average litter size is based.
c Based only on litters brought to term.
d See text footnote 3.

and mouse in susceptibility to prenatal killing, three further investiga-
tions, all on the rat, serve to show that even in the same species vastly
different results may be obtained, depending, perhaps, on genetic differ-
ences, details of method, or difficulties in early dosimetry. Job et al.
(1935) report 100 per cent resorption following exposure to only 95-200 r
(see Table 13-5). It must, however, be pointed out that females were
opened before irradiation to ascertain their pregnancy and, since no con-

874 RADIATION BIOLOGY

trol figures are cited, the operation itself may have contributed to the
prenatal death. It is otherwise hard to imagine how Warkany and
Schraffenberger (1947) could have obtained as large a yield as they did
with the very much higher doses of 190-1120 r. Summing all treatment
stages (days 9-15), 3 75 per cent of their 144 irradiated females had litters
at term and the average litter size was 5.26. Since results for different
doses are, in general, not reported separately, it is difficult to compare
stages. Table 13-5, however, makes it clear that, as in the mouse, there
is more prenatal death from irradiation of the earlier stages. In general
keeping with this trend is the report by Kosaka (1928b) that only }$ SED
suffices to kill all embryos irradiated during the "first stage" (days 5-10),
while during the second stage (days 11-15) 1 SED is required to produce
100 per cent mortality. Similar results are obtained in guinea pigs
(Kosaka, 1928c). In over-all sensitivity, Kosaka's rat embryos appear
intermediate between those of Job and those of Warkany.

In summary, it may be stated that extensive data on the mouse reveal
that, for comparable doses, irradiation during the period of major organo-
genesis causes considerably less prenatal death than does treatment
during the preimplantation period. Furthermore, within the period of
organogenesis, susceptibility to prenatal killing probably decreases fairly
rapidly with embryonic age. This is shown by the mouse results as well as
by individual investigations on the rat, although the latter differ greatly
amongst themselves with regard to absolute sensitivity of the embryo.

b. Neonatal. Only two investigators have reported in a quantitative
manner on death at birth following irradiation during the period of
organogenesis. In both cases, this is more frequent than prenatal death
observed by them in the same sample. Kaven (1938a) reports stillbirth
of over three-fourths of those newborns which had been irradiated
on day 10, 11, or 12 postfertilization with 178 r. Irradiation on the
immediately preceding or following day results in a smaller proportion
of death at birth.

A pooling of several experimental series (involving different genetic
strains) by Russell and co-workers is represented in Fig. 13-2. This
shows that, while 100 r and below has no effect on survival at birth, 200 r
(a dose comparable to Kaven 's) applied on any one of days 7% through
11}-^ inclusive causes neonatal mortality, with a sharp peak for days 9^
and 103^ (75 and 67 per cent respectively). Raising the dose to 300 r
increases neonatal mortality in general, the curve paralleling the 200 r
curve and the peak reaching 100 per cent (days 9}£ and 10^2) • These

3 Throughout this review an attempt will be made to name the stage according to
the actual time elapsed since fertilization — a procedure which is followed in the
publications of Wilson, and of Russell et al. Following this system, Warkany and
Schraffenberger 's "10th day" will be referred to as day 9 (if irradiation took place
in the afternoon, it was day 9J><$), etc

RADIATION IN PRENATAL DEVELOPMENT

875

data indicate that the slope of the dose-mortality curve for any given
stage is steep around 50 per cent lethality (see also Fig. 13-96). The
work of both Kaven and Russell makes it clear that mortality is markedly
dependent on the stage irradiated, even when stages are separated by only
ioo-

90-

80-

70-

<

uj 60

Q

_l
<

<

o

Ld

o

50-

40

30-

20-

10-

• 300 r
O 200 r

DAY IRRADIATED (POSTCONCEPTION)
Fig. 13-2. Incidence of neonatal death following irradiation with different doses at
different stages in the prenatal development of the mouse. {Data of Russell and
co-workers.)

24-hour intervals. For the data presented in Fig. 13-2, the LD5o at birth
varies as follows according to the stage irradiated:

For irradiation on days ^-S^ postconception, LD5o > 200 r.
For irradiation on days 9^ and 10^ postconception, LD50 < 200 r.
For irradiation on day 11^ postconception, 200r < LD5n < 300r.
For irradiation on days 12^-15% postconception, LD50 >300 r.

876

RADIATION BIOLOGY

2. BODY SIZE

Although only a few investigators report observations on weight or
length of their material, the information available is sufficient to demon-
strate an effect on body size of irradiation during the period of major
organogenesis. Kosaka (1928b, c, d) mentions generalized hypoplasia
from prenatally lethal doses in several rodents. Wilson and Karr (1951)
present data on prenatally observable growth retardation in rat embryos.
Raynaud and Frilley (1949a) and Russell (1950), both working on mice,

l.50-i

CONTROLS

1.40-

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l3i

DAY IRRADIATED ( POSTCONCEPTION)

• 200 r O 300r A 400 r
Fig. 13-3. Mean birth weights following irradiation with different doses at different
stages in the prenatal development of the mouse. (Russell, 1950.)

have measured size at term. Finally, there are two rather vague reports
of reduced body size in later life, in the rat (Hanson, 1923), and the
rabbit (Cohn, 1907).

Wilson and Karr (1951) found no effect on body weight 2-11 days after
irradiation with 50 r on day 10. With 100 r, however, there was a
marked growth retardation one day after irradiation, treated embryos
weighing 37 per cent less than controls. This high initial retardation
may have been due mostly to moribund embryos since the percentage
weight reduction in survivors decreased to between 6 and 15 per cent in
the succeeding period up to term. With 200 r, weight reduction
decreased to 20 per cent in survivors after an initial high of 39 per cent.
Finally, embryos irradiated with 400 r, all of which were already dead at
the time of first observation, appeared to have undergone at least some

RADIATION IN PRENATAL DEVELOPMENT 877

growth between irradiation and death. Of the animals which had
received 100 r, seven were raised and it was found that they made up
their initial weight deficiency (an average of 15 per cent at term) by the
age of 70 days.

Russell (1950) presents mean birth weights for groups of animals
irradiated in stages differing by 24-hour intervals and ranging from day
3^ to day 13^2 after fertilization (see Fig. 13-3). Several variables which
are known to affect birth weight are controlled: (1) newborns are genet-
ically uniform (Fi hybrids between two inbred strains); (2) the maternal
environment is genetically uniform; (3) parity is controlled, all animals
coming from second litters. Other variables, especially litter size, could
not be eliminated. A secondary control line was drawn in Fig. 13-3 to
provide a liberal allowance for the amount of milk sucked by the time of
weighing. It was found that mean birth weights for the 83^- through
13^2-day stages of treatment fell considerably below that control line.
The 200-r and 300-r curves were, in general, parallel with a mean differ-
ence of 0.2 g. Minima for both curves lay between the 10^2- and 11^2-
day stages. The short portion of the 400-r curve available paralleled the
300-r curve in the rising portion between days 12^ and 133^.

Holding the time of irradiation constant, Russell, Russell, and Major
(1951) obtained a more complete dose-weight series for day 11^2, which
is close to the stage of maximum susceptibility to growth retardation.
Points for different doses and for the control fall on an approximately
straight line, weight reduction per 100 r averaging 0.22 g over the three
available intervals (Fig. 13-9a).

The experiments of Raynaud and Frilley fall into a special category
since they involve very local irradiation with a narrow beam (0.5-3 mm
diameter) of very high dosage (100,000 r per exposure) in an attempt at
selective destruction of the pituitary. Five male mouse fetuses at term
which had been irradiated twice, at 12 days 6 hours and 13 days 6 hours
respectively, weighed, on the average, only about 60 per cent as much as
their control litter mates (1949a). The loss of weight due to direct
destruction of radiation-traversed head tissues does not account for the
total weight loss, which is considerable. This is also evident from the
fact that the irradiated animals showed a 10 per cent reduction in
xiphoid-anal length as well as the expected reduction in crown-anal
length. The results may be explained by secondary effects of pituitary
destruction, by such uncontrolled factors as scattered radiation to other
parts of the body, or, as the authors suggest, by indirect action of radia-
tion-produced toxins.

3. SEX RATIO

The only claim in the literature of an upset sex ratio following prenatal
irradiation is by Job et al. (1935), who state that treatment between the

878 RADIATION BIOLOGY

ninth and eleventh days with 35-50 r yielded 160.6 males per 100 females
(i.e., 61.6 per cent males) without modifying the litter size. Control
ratios are not reported and even significance of the difference from a 1 : 1
ratio cannot be calculated since the actual numbers for this group are not
given. (In other groups, similar with respect to stages irradiated, the
sex ratio appears normal; e.g., treatment between the eighth and eleventh
days with 27-90 r gave 141 males and 132 females.) It may, however,
be pointed out that a 1 : 1 sex ratio is not necessarily expected at birth
even if it exists at conception. Russell (1950) reports 61.0 per cent new-
born males in one group of controls (difference from 50 per cent males =
2.5 X S.E.) in comparison with which the excesses of males in both
broad experimental categories (preimplantation and postimplantation)
are nonsignificant.

Although it is, of course, quite conceivable that the sex ratio at birth
might be affected by prenatal irradiation — either through differential
mortality of one sex or through actual sex reversal (e.g., loss of one X
chromosome) — valid reports of such effects must to date be considered
lacking.

Job et al. further claim that males are more susceptible to the induction
of abnormalities than are females. In the group irradiated between the
eighth and eleventh days, which yielded all the morphological abnormali-
ties reported, 37.4 per cent of 123 males and 24.8 per cent of 109 females
treated with 36-90 r were deformed. The difference is at the 5 per cent
level of significance (t = 2.07). The authors state, however, that only
one of the abnormal animals had been irradiated on the eighth day, the
other seventy-two between the ninth and eleventh, and the percentage of
males obtained following irradiation on these days (as given without
actual figures) is 61.6. The sex incidence of 62.7 per cent males among
the abnormals then seems as expected on the basis of random distribution.
The authors' claim must thus be considered unproved. Again, however,
it is quite conceivable that differential sensitivity of the sexes to the
induction of abnormalities will be found.

4. MORPHOLOGY

Even within the relatively limited literature on the subject a vast
number of abnormalities has been reported as resulting from the irradia-
tion of mammalian embryos during the period of major organogenesis.
It is, therefore, unfortunate that some authors failed to realize the impor-
tance of timing accurately the stage at which the embryos were subjected
to treatment, while others neglected to provide such pertinent informa-
tion as total numbers (for the calculation of percentage incidence) or
exact dosage. However, not many of the later publications fail in this
respect, and the earlier ones, which do, were nevertheless useful in demon-
strating that there existed a fertile field for future work. The earliest

RADIATION IN PRENATAL DEVELOPMENT 879

report of abnormalities was by von Hippel and Pagenstecher (1907) who
obtained cataracts, microphthalmia, and lid coloboma in newborn rabbits
following irradiation with a high dose (21 H = Holzknecht units) on
days 7, 9, and 11, or 8, 10, and 12 after fertilization. Bagg (1922) men-
tions one case of "dislocation" of the spinal column in a rat fetus which
had been treated between the tenth and fourteenth days of intra-uterine
life with radium emanation. Several other fetuses in this group suffered
from extravasations in subcutaneous vessels and along meningeal sinuses
(Gudernatsch and Bagg, 1920). But since the same type of lesion could
be produced by injecting the mother 22 days before conception and since
it is unlikely that the activity of the circulating radiation source would be
maintained long enough to affect the embryos directly, an indirect non-
specific effect through lasting injury of the mother must be postulated in
this case. Hanson (1923) found that rat females in "later stages of preg-
nancy when given the proper dosage" of X rays produced litters in which
one or more of the young had serious eye defects, changes in the shape of
the skull, and paralysis of the limbs. When raised, they showed consider-
able growth retardation and nearly all proved sterile. De Nobele and
Lams (1927), reporting on what appears to be the same material as that
in which they studied radiation-induced prenatal mortality (1925)—
although it is possible that some additional animals are included in the
later publication — state that a few of the treated rat embryos which were
permitted to come to term were afflicted with microphthalmia, while
some of the irradiated guinea pig embryos later developed hydrocephalus
and dilatation of the lateral ventricles. Murphy and de Renyi (1930)
report foot abnormalities in all of five litters irradiated prenatally with 400
or 800 r. Since, unfortunately, the method consisted of giving a mixed
population of virgin and pregnant females a series of exposures and then
considering only those whose litters were cast within twenty-two days of
the last exposure, it is not known at what stages embryos were irradiated,
or even whether they were irradiated once or twice.

On turning to the more extensive and more carefully controlled work,
it appears that the field has been explored in two general ways : (a) com-
parison of the results of irradiating a number of different stages in order
to determine whether the changes produced are characteristic of the
treatment day (Kosaka, 1927, 1928a, b, c, d, e; Job, Leibold, and Fitz-
maurice, 1935; Kaven, 1938a; Warkany and Schraffenberger, 1947;
Russell, 1949, 1950; Russell and Russell, 1950b) ; and (6) concentration on
one or two stages (Raynaud and Frilley, 1943b, 1947a, b, c, 1949a, b;
Wilson and Karr, 1950, 1951; Russell et at., 1951), or a certain group of
abnormalities (Pagenstecher, 1916; Kaven, 1938b; Hicks, 1950) with the
ultimate aim of tracing the genesis of the malformations. All experi-
ments (groups a and b) will first be briefly outlined in turn. Following
this examination of each experiment, particularly with regard to the evi-

880 RADIATION BIOLOGY

deuce for critical periods, tabular comparisons between experiments will
be made in an attempt to derive some general conclusions about certain of
the abnormalities. Many others will have to be ignored, due to limita-
tions in scope of this review.

a. Experiments That Compare Results of Irradiating Different Stages.
The first investigator to attempt a survey of the gestation period with
regard to radiosensitivity was Kosaka who worked on mice (1927, 1928e),
rats (1928b), rabbits (1928a),4 and guinea pigs (1928c), irradiating during
known intervals (though not definite days) postconception with various
doses, ranging from \£ to 2 SED, and observing effects manifest 6, 12, 24,
48, 72, et seq., hours after irradiation. A large part of the work on all
animals was devoted to histological description of tissue damage which
was apparently not considered as malformation since special mention is
made (1928d) of the fact that malformations (microcephalus, deformity
of the extremities) occurred only in the rabbit. For any given stage of
irradiation, tissues were ranked in order of decreasing sensitivity, and this
rank order was found to change with the stage in a manner parallel to the
change in relative growth rates of the particular organs. Early in the
period of organogenesis, brain and spinal cord are most sensitive, retina
and mesoderm in second place. Shortly thereafter, spinal cord loses
much of its sensitivity, while retina joins brain in first place. During
the period of the fetus, thymus suddenly becomes extremely sensitive, and
liver and spleen move up on the list. A variety of other tissues in the
lower ranks of sensitivity must be omitted from mention here. Finally,
certain organs showed no marked effects from irradiation at any stage.
One interesting finding was that, while the processes of regeneration in
most tissues were similar in kind to the growth which had occurred just
before irradiation, cerebrum and retina responded to severe damage by
forming numerous ependymal canals believed analogous to the neural
tube of early stages.

Job et al. (1935) posed the question of whether certain periods in
development could be demonstrated to be critical either for the rat
embryo as a whole or for certain of its organ systems or individual
organs. After eliminating higher doses because of excessive mortality,
they obtained litters from sixty-six females which had received a single
dose of X rays of 90 r or less between the first and sixteenth days of
gestation. In all cases where irradiation had been before the eighth day
or after the eleventh day, the young were normal. Following irradiation
between eighth and eleventh days, 17 of the young (7 per cent) were
hydrocephalic, 14 (6 per cent) suffered from jaw abnormalities, and 52

4 Unlike Kosaka's other publications, the paper on rabbits lacks an English or
German summary. This reviewer did not obtain a translation of the Japanese text.
Scattered information about the rabbit results was obtained from the other papers of
the series, particularly the summary paper (Kosaka, 1928d).

RADIATION IN PRENATAL DEVELOPMENT 881

(22 per cent) had eye defects (about % bilateral) combined in 8 of the
animals with one of the other two abnormalities. t Breaking down this
generally sensitive period, it appeared that hydrocephalus resulted from
irradiation on the ninth day and jaw abnormalities on the eleventh day,
while most of the eye defects were produced by treatment on the tenth
day. Job et at. can thus be considered to have brought the first demon-
stration of well-defined critical periods for abnormalities in different
characters. Incidence for individual treatment days cannot be calcu-
lated since separate totals are not given, but the qualitative results are
included in Figs. 13-4 to 13-7 for purposes of comparison with the findings
of other investigators.

In an experiment essentially similar to the above, Kaven (1938a)
obtained litters from over 162 mouse females irradiated on any one of
days 7 to 195 with a constant dose of 178 r (at the level of the embryo or
fetus). Two improvements over Job's method were (1) the use of
genetically homogeneous material, and (2) observations on a large control
group (Kaven, 1938b). Abnormals are reported for each group sepa-
rately and give clear indications of the existence of critical periods even
though percentage incidence can usually be only estimated (since Kaven,
like Job, does not report the number of young observed within each
group). The qualitative results are included in Figs. 13-4 to 13-7. The
most frequently affected structure, the tail (abnormal in over 130 new-
borns), is sensitive only between days 9 and 14, with peak sensitivity
probably on day 11. Brain hernias were obtained exclusively from
irradiation on day 8. Although both the incidence and total number in
this group were small in the original series, a separate experiment (Kaven,
1938b — see p. 890), in which irradiation was given only on days 7 or 8,
established that the original cases had not been spurious. Four other
abnormalities are reported, in the survey series, from treatment during
the period of organogenesis: head malformation and head hemorrhage,
each appearing in one of eighteen litters irradiated on day 10; hydro-
cephalus (apparent a few days after birth), occurring in about 10 per cent
of the animals irradiated on day 12 and in lower proportions from the
treatment of days 13 and 10; and skin defects arising later in life as a
result of irradiation on days 13 and 14 postconception. (Other abnormal-
ities produced by Kaven — e.g., torsion of limbs, digital abnormalities-
were apparently not noticed by him but are apparent from the photo-

8 Kaven followed two systems of timing pregnancy: (1) by actual observation of
copulation, in which case his "first day" equals day %$ to 1%, depending on hour of
mating and hour of irradiation; (2) by looking for vaginal plugs, which, in the majority
of cases, represent fertilizations between 1 :00 and 4:00 a.m., so that, under this system,
his "first day" approximately equals day 134 to 1%, depending on hour of irradiation.
It is thus obvious that within a given one of his stage groups, pregnancies may differ
in age by a whole day. In general, the designation of his "first day" as day one is
probably representative of the majority of his cases and will be followed in this review.

882 RADIATION BIOLOGY

graphs.) The numbers of afflicted animals are small but assume impor-
tance in comparison with the results of other workers.

Warkany and Schraffenberger (1947) studied litters from 108 rat
females irradiated on any one of days 9 to 15 of gestation (referred to by
the authors as "10th to 16th" days)6 and reported well over a dozen
different abnormalities. Unfortunately they do not separate the results
for different doses but give only the dose range (often very wide, e.g.,
190-900 r) for each stage group. Since, at any one stage, an increase in
dose generally increases percentage incidence of a given abnormality
(Russell, 1950), it is obviously impossible to determine the most sensitive
stages in Warkany and Schraffenberger's experiment by comparing inci-
dence : for a peak may be due merely to a higher dose having been used.
Moreover, since, by raising the dose, a primordium or process can usually
be affected on days adjacent to that of maximum sensitivity (Russell,
1950), it is not surprising that with the large doses used (up to 1120 r) the
authors did not usually get very clearly defined critical periods. Even
so, a few emerge when incidences are calculated from their report and
when allowance is made for the dosage difficulties discussed above (see
Figs. 13-4 to 13-7). Thus, the peak for rib angulation is definitely on
day 14, the long bones of the front legs are sensitive almost exclusively on
day 12, and those of the hind legs mostly on day 12 except for the tibia
which has a peak sensitivity on day 10. These points and others become
interesting in comparison with other experiments (see pp. 891-893).

Like Kaven, Russell (1949, 1950) used genetically uniform mice, in her
case an Fi hybrid between two inbred strains to add vigor to uniformity.
The scope of the experiment was extended with respect to ( 1 ) the number
of stages surveyed, (2) doses used (200 r for all stage groups, 300 and
400 r at certain ones), and (3) types of abnormalities observed. The
skeleton was chosen for detailed study mainly because its various parts
are formed by processes whose periods of maximum activity, taken

6 See footnote 3.

Figs. 13-4 to 13-7. Results of several groups of investigators represented to show
critical periods for the induction of various abnormalities.

Wherever sufficient data are given in the original publication, representation here is
by (a) percentage incidence of abnormality (see scale in figures) and (b) magnitude of
dose required to produce abnormality. Thus, the wider and more heavily shaded a
band, the greater the sensitivity. Absence of a band at a particular stage indicates
that the abnormality did not occur in the irradiated group (see, e.g., 200 r results of
Russell), except where the serrated end of a band indicates that the dose series was
not continued to the stage in question (see, e.g., 300 and 400 r results of Russell, all
results of Wilson and Karr). For cases where exact incidence for a given stage and
dose cannot be calculated from the original data, representation is only roughly
quantitative. In general, + = 1-49%, >200 r; + + = 1-49%, 1-200 r; + + +
= 50-100%, >200 r; + + + +=50-100%, 1-200 r. (J = Job, Leibold, and
Fitzmaurice, 1935; K = Kaven, 1938a,b; R = Russell, 1950, 1949; Wi = Wilson and
Karr, 1951, 1950; Wa = Warkany and Schraffenberger, 1947.)

RADIATION IN PRENATAL DEVELOPMENT

883

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884

RADIATION BIOLOGY

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RADIATION IN PRENATAL DEVELOPMENT

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886

RADIATION BIOLOGY

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altogether, spread over a considerable portion of embryonic life. In
addition to skeletal abnormalities, changes noted on careful external
examination and on gross dissection were recorded. A large control
group (372 newborns) was studied in the same manner as 420 newborns
which had been irradiated during prenatal life. Since it is impossible to
give a complete report of changes observed (the checklist of characters
examined included about a hundred items), only the most striking ones
and those most useful for comparison with the results of other investi-
gators are represented in Figs. 13-4 to 13-7. These samples will serve to
illustrate the general type of result obtained.

Almost all abnormalities represented in Figs. 13-4 to 13-7 were found in
newborns or fetuses at term but are projected back upon the stage at
which they were apparently induced. Width of the bands represents
percentage incidence at term. Without going into details about the
abnormalities, several general features of Russell's results may be pointed
out. First, it is striking that with the general survey dose of 200 r,
abnormalities in a high proportion of the animals are obtained only from
embryos exposed after day 5)^ (no external and visceral and only 2 per
cent slight skeletal abnormalities from irradiation of earlier periods). At
the other end, day 13^ is probably the limit beyond which abnormalities
of the types looked for at term can no longer be obtained with 200 r.

RADIATION IN PRENATAL DEVELOPMENT 887

This broad division of the gestation period has already been mentioned
and the special features of the first phase (preimplantation) have been dis-
cussed. Second, it is apparent that critical periods7 for the induction of
almost all abnormalities are short. The 200-r series shows many of
them to be restricted to one particular stage (e.g., spina bifida, reduction
of the ilium, abnormalities of the basihyal, etc.), and even when they
span several stages, one often stands out as the main sensitive one on the
basis of incidence (e.g., anterior premaxillary fusion, vertebral jumbling,
rib fusion, coloboma). Third, the effective period of disturbance is
lengthened by raising the dose, indicating that a certain degree of sensi-
tivity exists at stages other than the most critical one. High doses are
thus not suitable for the mapping of critical periods except for abnormali-
ties which have high thresholds of induction (e.g., deformities in limbs
and girdles). The addition of higher dose series is useful, however, not
only for conclusions which can be drawn from dosage comparisons (dis-
cussed later in Sect. IIIC) but also for the confirmation of location and
shape (e.g., unimodal, or bimodal as in tail reduction, etc.) of any given
critical period.

b. Experiments That Concentrate on a Few Stages or on a Group of
Abnormalities. In contrast to the experiments discussed so far, Wilson
and Karr (1951) confined their X-ray treatment of rat embryos to day 10.
Survivors which had received 50, 100, or 200 r (18, 40, and 31 embryos,
respectively) were sacrificed 1-5 days later and examined for morpho-
logical changes. A small number of embryos in the 100-r group were
allowed to come to term and were observed postnatally. All the pre-
natal findings of aberrant growth in various organs, as well as damage to
the liver (classed by the authors with localized retardations rather than
with malformations), are represented in Figs. 13-4 to 13-7. It may be
noted that 50 r was ineffective except in the production of two cases of
slight microphthalmia. For most changes, incidence with 200 r is higher
than with 100 r. The eye was the most consistently affected of all
organs, giving 75 per cent abnormalities even with 100 r. Defects of the
extremities and liver damage cannot be diagnosed until the thirteenth or
fourteenth days, when these organs have reached a stage of differentiation
adequate for observation. Percentage incidences are, therefore, based on
considerably lower totals (14, 27, and 16 animals, respectively, in the
three dose groups) but, as they stand, they approximate eye defects in
frequency in the 200-r, though not in the 100-r group. Urogenital mal-
formations include (in order of decreasing frequency) incipient horseshoe

7 The critical period for a given change from normal development may be defined
as the developmental moment or interval during which radiation must be applied to
produce that change at some specified stage of observation, if the dose of radiation is
the lowest one that gives a detectable incidence of that change. The term "critical
period" does not necessarily imply that the immediate primordium of the character
malformed as a result of irradiation at a certain stage was damaged at that stage.

888 RADIATION BIOLOGY

kidneys, defects in the mesonephros, inhibition or duplication of a meta-
nephric bud, and epispadias. In the central nervous system, the brain
was the only part affected by irradiation on day 10; in all but one of the
cases, malformations involved the forebrain, mainly the telencephalon.
A tendency to reversed asymmetry in dominance of aortic arches, com-
bined in two of three cases with reversed tail curling, gave indications of
a certain degree of situs inversus. Abnormalities of the extremities were
usually more pronounced in, or limited to, the forelimbs. In mild cases
only the distal elements were affected, while distortions and deficiencies
extended more proximally in more severely afflicted individuals.

Set apart from the malformations are cases of localized retardation, i.e.,
tardiness in normal processes rather than aberrant growth. Criteria for
gauging retardation are considered not completely satisfactory, especially
where malformations are superimposed. Brain and urogenital organs
are most frequently retarded, but heart, aortic arches, and lungs may
also be affected. Also classed with retardation rather than malformation
is liver damage, which frequently involves a reduction (occasionally down
to complete absence) of hemopoietic elements. While the impairment is
only slight and transitory in the 100-r groups, the authors feel that in the
200-r groups it represents a significant functional loss and may be associ-
ated Avith the high rate of prenatal death. As judged by eight 100-r
animals allowed to go to term, there may be recovery from localized
retardations except in the eye, where microphthalmia may become
anophthalmia or malformation, and in the heart (one case).

A few results of irradiation on day 9 are mentioned by Wilson and Karr
in an abstract (1950) and used (1951) for occasional comparisons with
the day-10 results. While reversal of symmetry was only indicated
following irradiation on day 10, treatment on the preceding day may
give situs inversus totalis. Furthermore, there seems to be a very sharp
division for the type of central nervous system anomalies obtainable from
the two treatment stages, day-10 irradiation never yielding the class of
defects that results from faulty closure of the neural tube, while irradia-
tion on day 9 often produced meningocele, encephalocele, cranioschisis,
etc. More prevalent after day-9 exposure were aberrant cell islands of
neural origin, growing independently in the mesenchyme surrounding the
brain. These are described in more detail by Wilson, Brent, and Jordan
(1951) and may be comparable to the regenerative growths found by
Kosaka (1928b, c, see p. 880).

In one experimental series, Russell et al. (1951) concentrated on the
irradiation of day 11^. However, since this experiment also involved
irradiation under hypoxia, it will be reviewed in a different section (see
Sect. IIIB).

The experiments of Raynaud and Frilley fall into a class by themselves
since, because of the method employed, the results cannot be used to

RADIATION IN PRENATAL DEVELOPMENT 889

indicate differential sensitivity throughout the body. The mother's
uterus was exposed by laparotomy and a localized beam (0.5-3 mm diam-
eter— see 1943a for technique) directed from below at the head region of
the embryos, particularly at the base of the diencephalon. The trav-
ersed region also included the central shaft of the cranium, the buccal
cavity, and the primordium of the tongue. The dose range was 5000-
200,000 r. The investigators confined exposures to mouse embryos of a
certain age, namely, 12 days 6 hours, adding occasionally a second
irradiation 24 hours later. All observations were made just before term.
In three publications (1943b, 1947a, c), the authors report on effects in
structures directly traversed by the pencil of X rays; in another three
(1947b, 1949a, b), they describe observations on the reproductive system
which was not irradiated but could conceivably have been secondarily
affected as a result of pituitary damage.

As expected for the high doses used, there was considerable damage in
the radiation path. Externally, the animals treated with 5000 40,000 r
were microcephalic, often had open eyelids, and occasionally lacked ears,
tongue, salivary glands, or upper or lower jaws. After irradiation with
190,000 r, the head at term was only a small atrophic mass. Condensa-
tion of mesenchyme to precartilage was suspended and membranous
ossification suppressed even with 5000-40,000 r. The same dose range
produced extensive cerebral lesions, and complete destruction of the optic
nerve and of both layers of the retina. Following treatment with
60,000 r, the brain at term showed the vesicle configuration characteristic
of the stage of irradiation; while 190,000 r left only a necrotic mass with
the vesicles no longer recognizable. Complete destruction of the anterior
lobe of the pituitary could be achieved only with 200,000 r divided evenly
between 12 days 6 hours and 13 days 6 hours. It should be noted that, in
spite of the massive damage due to the high doses used, it is possible to
find in the data evidence of differential susceptibility of different struc-
tures. Thus, only a quarter (or less ?) of the dose required to inactivate
the anterior lobe will completely eliminate the pars nervosa; the choroid
plexus resists 60,000 r, a dose which causes almost complete destruction of
the rest of the brain; the lens of the eye resists doses which completely
destroy retina and optic nerve.

Several indirect changes were noted in regions presumably not trav-
ersed by radiation. Effects on body size have already been discussed
(see p. 877). In addition, there was marked atrophy of the adrenal
cortex and a reduction in liver glycogen. In spite of pituitary destruc-
tion, the histogenesis of the genital tract, including accessory structures,
and the cytological differentiation of the germ cells proceeded normally,
which led the authors to suggest that either the hypophysis exerts no
gonadotropic influence in development or that its function can be taken
over by maternal or placental hormones. There was, however, up to

890

RADIATION BIOLOGY

70 per cent reduction in the number of germ cells in both sexes and some-
what reduced masculinization in the external genitalia of males, which
would point to the lack of some normally available trophic influence or to
a general toxic effect (e.g., in the blood) caused by the localized radiation.
The experiment of Kaven (1938b) forms a transition between those
concentrated on a particular stage and those concentrated on a certain
group of abnormalities. His concern was with the latter — namely,
meningocele and possibly related changes — but his earlier experiment
(1938a) had yielded information on the effective treatment stage. The
pertinent features of Kaven's experiment may be read from Table 13-6,

Table 13-6. Incidence of Brain Abnormalities and of Mortality in Fetal
Stages and at Birth Foi lowing Irradiation of 7- or 8-day-old Mouse Embbyos

(Kaven, 1938b)

Irradiated day 7

Day observed : 14

Number of litters .

Number of fetuses

Per cent dead

Per cent extrakranielle

Dysencephalie11

Per cent meningocele" . .

Average litter size:

Total-

Expected at term''. . . .
Found at term

31

237
65.8

4.9
0

7.6

2.6

17

24

182
72.0

0

17.3

7.6
2.1

Birth

39

166
11.4

0
2.4

4.3

Irradiated dav 8

13-16

17-19

40

118

39

0

16

7

2

8

169
30.8

3.4
19.7

7.2
4.4

Birth

47

219
13.7

0
16.1

4.7

Control

Birth

306

2066

1.7

0
0

6.8

a Percentages for prenatal observations are based on living fetuses only, in order to
make them more comparable with results at birth, when the majority of prenatally
dead animals will not be counted.

b That is, living and without "extrakranielle Dysencephalie."

which was compiled from information scattered through his paper. The
results show that brain hernias (more correctly, meningoceles) at birth
are significantly more frequent following irradiation on day 8 than on
day 7 (they were never obtained from later stages — Kaven, 1938a), and
that this abnormality becomes macroscopically apparent only late in
prenatal development (all cases but one after the sixteenth day). A find-
ing entirely unexpected from the birth data was the presence, in dissected
uteri, of fetuses with "icepack" brains, i.e., pseudencephalics. This
abnormality, referred to by Kaven as "extrakranielle Dysencephalie," is
about five times as frequent in 13- to 16- as in 17- to 19-day fetuses and
is apparently missing in litters not observed until shortly after birth.

RADIATION IN PRENATAL DEVELOPMENT 891

Reduction in litter size at birth in the group irradiated on day 8 is com-
pletely accounted for when these abnormals are added to the number of
placental rests (but agreement for day 7 is not very good). Kaven sug-
gests that the various effects may be explained by embryos being in
slightly different stages of neural tube closure at the time of irradiation on
day 8. The developmentally (not necessarily chronologically) youngest
are most sensitive and die early (placental rests at time of observation).
This is supported by the finding that irradiation on the preceding day
(day 7) gives a greatly increased prenatal mortality. Slightly more
advanced embryos respond with development of pseudencephaly and die
shortly before term. Still more advanced embryos are at least externally
normal for a considerable period, then about 3 days before birth develop
meningocele and show reduced postnatal viability. Finally, the most
advanced embryos are not affected at all.

Pagenstecher (1916) concentrated his studies on rosette formation in
the retina, after von Hippel and Pagenstecher had demonstrated very
early (1907) that various eye abnormalities could be obtained from the
irradiation of rabbit embryos. In four fetuses, observed 3-4 days before
term, which had been irradiated between days 9 and 15 postfertilization,
he found three with rosettes (five out of eight eyes). He believed that
rosettes were the simplest type of retinal damage and that each repre-
sented the apex of an altered fetal fold of the retina.

Hicks (1950) irradiated rat and mouse females "in the second and third
week" of pregnancy (without closer timing of the stage of treatment) and
reports the resulting nervous system changes. Mention is made of the
fact that extraneural damage is not obtained with doses below 400 r.
Since this is in contradiction to all other work (already reviewed) on
mouse and rat for irradiation during the second week, it seems probable
that virtually all of Hicks' data came from exposure during the third
week. They will therefore be discussed in the section dealing with
irradiation during the period of the fetus.

c. Comparison of Experiments. In an attempt to derive more general
conclusions about certain of the abnormalities which have been obtained
through irradiation of embryos, Figs. 13-4, 5, 6, 7 have been constructed
from data contained in eight publications of five groups of investigators
reviewed in preceding sections. The figures include all abnormalities
reported by Job et al. (1935), Kaven (1938a, b), Warkany and Schraffen-
berger (1947), Wilson and Karr (1951, 1950) ; and a sample of the abnor-
malities reported by Russell (1950, 1949). The data of Kosaka, of
Pagenstecher, of Hicks, of Murphy and de Renyi, and of early authors
contain either insufficient information or none at all on stage of irradia-
tion. They, therefore, had to be omitted from the comparison since they
can contribute little or nothing to conclusions about critical periods.
Also omitted are the experiments of Raynaud and Frilley which involved

892 RADIATION BIOLOGY

localized radiation and could thus not be expected to indicate differential
sensitivities throughout the body.

It should be pointed out that several approximations have had to be
made to construct Figs. 13-4 to 13-7. They represent work on two differ-
ent animals — rat and mouse — which had to be matched according to
equivalent developmental stage. In most of the original publications,
data are not represented in a manner to indicate critical periods. Many
are incomplete: Job does not report totals for calculation of percentage
incident, Kaven only occasionally; Kaven's observations were apparently
not thorough since he missed limb defects clearly indicated in one of his
illustrations; Warkany and Schraffenberger list only wide dose ranges
(e.g., 190-900 r) instead of separating results according to dose. It must
also be remembered that the thorough work of Wilson and Karr was
confined mainly to day 10.

Despite the various limitations, agreement in most cases is good and
some of it is striking. Only some of the comparisons can be discussed
here. In the work of three authors, the critical periods for all long-bone
defects in the forelimbs seem to be strictly confined, both in mouse and
rat, to the stage when the anterior limb buds first become apparent as
rudder-shaped outgrowths. The total period for the hind limb is some-
what longer and the work of two authors indicates that the tibia is the
first bone to be sensitive, the fibula the last. Warkany and Russell agree
well, both on Polydactyly and on foot reduction (oligodactyly, syn-
dactyly, etc.), but for the latter there is an additional early critical period
in the rat (Wilson, Warkany) which has not appeared in the mouse data.
The general period sensitive to the induction of eye abnormalities is
similar in three experiments.

There is agreement between authors on apparent bimodalities in
critical periods of three characters (ribs, tail, palate). In the case of the
ribs, two different abnormalities — fusion and angulation — are obviously
represented without overlap in times of sensitivity. For the tail, such
distinction cannot be made from the end result, but very probably two
different developmental processes are affected (Russell, 1950). All cases
of cleft palate resulting from early treatment were found by Russell
(1949) to be associated with anterior premaxillary fusion, while those
following irradiation during the second period were independent of this,
thus probably indicating two entirely different precursor processes for
the abnormality. Warkany does not report any such distinction but,
since short upper jaw occurs in his material irradiated during the early
period, a similar situation may exist.

Russell found only one instance of situs inversus in the mouse, but this
corresponds well in critical stage with the more frequent cases of Wilson.
On the other hand, the urogenital malformations described by these two

RADIATION IN PRENATAL DEVELOPMENT 893

investigators are probably of different types and it is thus not surprising
that the critical periods differ.

C. THE PERIOD OF THE FETUS

Very few of the more recent investigators have concentrated their
attention on irradiation during the period of the fetus. Thus, the bulk
of Table 13-7 represents either early (often inadequate) work, or newer
experimental series in which the main emphasis lay on the period of
organogenesis but which, for one reason or another, were extended to
include the first few days of the period of the fetus. It will, therefore,
suffice to mention only the few highlights on which evidence seems
adequate.

1. MORTALITY

Mortality resulting from irradiation of fetal stages has been found to
occur mostly in the period between birth and two weeks of age (see
also Table 13-7). The mode is probably shortly after birth, but the fre-
quency there is considerably less than for equivalent doses used during
the period of major organogenesis (Kaven, 1938a, see p. 874; Russell, see
Fig. 13-2). Prenatal death has been noted only by a few authors
(Kosaka, 1928c; de Nobele and Lams, 1925; Schinz, 1923) for doses in the
neighborhood of 1 SED and above, i.e., even higher than those which will
cause early postnatal death. In early experiments where absolute doses
were not known (Burckhard, Trillmich, Saretzky), exposures equal to
those which would give a high degree of prenatal mortality when given
earlier in pregnancy were without effect when given during the period of
the fetus.

A kind of acute fatal radiation sickness may occur during the first week
or two following birth. Bagg (1922) reports that 7-ray treatment of
rats 2 or 3 days before term produces in approximately half the animals of
each litter symptoms of anemia, diffuse edema, and meningeal, spinal
cord and subcutaneous extravasations, leading to death at about a week
of age. Autopsy revealed fatty degeneration of the liver and desquama-
tion of the lining of the intestinal mucosa. Job et at. (1935) report
postnatal anemia, diarrhea, hemorrhagic exudates in eye and nose,
underweight, and abnormal nervousness following X-ray treatment of
rat fetuses with more than 1 "skin unit," but no effect from lower doses.
(It is not clear whether 1 skin unit = 1 SED.) Lacassagne and Coutard
(1923) found that fifty-three of fifty-four rabbits, which had been irradi-
ated 2-3 days before birth, died of ' ' purpura roentgenien. ' ' Death occurred
always on the tenth day after birth following a relatively invariable syn-
drome (hemorrhagic spots in skin and viscera ; extravasations from peri-
toneum, pleura, pericardium; edema; torpor; dyspnea; — note resemblance

894

RADIATION BIOLOGY

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RADIATION IN PRENATAL DEVELOPMENT

895

to Bagg's syndrome) , which was first noticeable externally on the fourth
day (Lacassagne, Lavedan, and Leobardy, 1922). Coagulation time was
progressively decreased, starting with the seventh day, and platelets were
absent. Lacassagne and Lavedan (1922) report blood counts on pre-

,•-5

IRRADIATED
RABBITS

CONTROLS

D D

RED BLOOD CORPUSCLES
POLYNUCLEAR WHITE CELLS
MONONUCLEAR WHITE CELLS

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POSTNATAL

POSTCONCEPTION

AGE, DAYS
Fig. 13-8. Average counts of red blood corpuscles polynuclear white cells, and
mononuclear white cells in rabbits irradiated in utero 2-3 days before term.
Counts were begun following irradiation on day 29 postconception and were con-
tinued daily until death. (Graphs constructed from data presented by Lacassagne and
Lavedan.)

sumably the same group of animals. Figure 13-8 was constructed from
their data. The red count of irradiated animals is normal until the
fourth day after birth when it takes a sharp drop and continues to fall.
The effect on total white count is more immediate, consisting of almost

896 RADIATION BIOLOGY

complete inhibition of the very sharp rise which normally takes place at
birth: while control counts increase from 350 to 5800 by the tenth day,
irradiated animals stay at the fetal level except for a temporary rise to
1100 on the fifth day. The deficiency of polynuclear cells is somewhat
greater than that of mononuclear cells, the former averaging only about
29 per cent of the total white count for the period from birth to ten days
instead of the normal 43 per cent.

2. MORPHOLOGY

Irradiation during the period of the fetus does not lead to the striking
changes or malformations in newborns which are produced by irradiation
with comparable or even lower doses during the period of major organo-
genesis (Job, 1935; Kaven, 1938a; Russell, 1950). It is, however,
erroneous to assume that exposure during the latter part of pregnancy is
without morphological effects. It must be remembered that there is less
chance for any damage to express itself as a gross change by the time of
birth, for not only is the irradiation-to-observation interval shorter, but
the rate of development averages considerably less over that period than
over the interval from earlier exposure to birth. On these grounds, it
may be expected that gross changes will become expressed later in life and
these have indeed been reported by a number of authors (Bagg, 1922;
Kosaka, 1928e; Kaven, 1938a; Russell, 1950; Hicks, 1950; Grobman,
personal communication). Several studies on fertility point in the same
direction (Parkes, 1927; Kosaka, 1928e; Hanson, 1923). The immediate
damage which may lead to later gross changes was demonstrated by
Kosaka (1928b, c) and Hicks (1950). General growth retardation has
also been reported (Kosaka, 1928e; Job et al., 1935). Some of the studies
enumerated will be briefly described in the rest of this section.

Delayed morphological effects of irradiation during the fetal period were
first noticed by Bagg (1922) in survivors from the group in which acute
radiation death had been described (see p. 893). Externally, these
animals exhibited opaque pupil and atrophied lens. At autopsy at about
a year of age, Bagg also noted smallness of the cerebral hemispheres,
especially the neopallium (occasionally even complete absence of the
cortex), and arrest of the gonads, ovaries and testes alike. Kosaka
(1928e) also found very marked hypoplasia of the cerebrum and the
gonads, and occasionally a smaller degree of arrest in lung, liver, heart,
and kidney in surviving mice (or rats ?) irradiated after the fourteenth
day postconception with % SED. Both Kaven (1938a) and Russell
(1950) report later development of skin defects and of cataracts resulting
from irradiation during the period of the fetus but it is not known whether
that period is critical for the production of these changes since mice
exposed earlier were not kept alive in large enough numbers after birth.
Working on the same Fi hybrid used by Russell, Grobman autopsied

RADIATION IN PRENATAL DEVELOPMENT 897

twenty-nine mice 37-49 days old which had been irradiated with 300 r
on day 143-^ or 153^ postconception. He found reduction in size of
gonads and secondary sex glands, absence of the corpus callosum, and
absence of the gallbladder. Defects in the reproductive system could
also be produced by earlier irradiation (day 113^2, 123^, or 133^).

Sterility has frequently been observed to result from irradiation during
fetal stages. Parkes (1927) raised twenty-five mice treated between days
10 and 18 postconception (mostly days 14 to 17). Only four of sixteen
females and neither of two males tested proved fertile. Kosaka (1928e)
found that, while irradiation between the seventh and thirteenth days did
not affect the fertility of survivors, irradiation after the fourteenth day
completely sterilized all but two males (totals not given). Even these
two (irradiated on the seventeenth day) were only temporarily fertile.
Sterility in all males was due to failure of sperm formation. Some
females too were completely sterilized but the majority was poorly
fertile and showed reduction in the number of mature ova formed.
Hanson (1923) mentions that "nearly all" rats (no numbers given)
irradiated in fetal stages proved sterile. Nurnberger (1920) found
sterility in the only one of his guinea pigs (a male) which survived the
early postnatal period.

Two investigators have made observations on immediate effects of
irradiation during the period of the fetus. Kosaka (1928b, c) reports
findings on rat and guinea pig fetuses from fourteen and eighty-five
litters, respectively, sectioned 6, 12, 24, 48, 72 hours . . . (etc., to term)
after irradiation with ^-2 SED during the fetal period. In the rat, he
finds most tissue damage in the brain, retina, and thymus; liver and
spleen take second place, and skin third. In the guinea pig, results are
similar for irradiation in early fetal stages. Later in gestation, when the
guinea pig fetus is really comparable to a postnatal rat or mouse, brain
and retina lose some of their sensitivity but thymus remains in first
place. Hicks (1950) irradiated rat and mouse females in late pregnancy
(unfortunately without timing the stage of treatment) and observed the
resulting nervous system changes ^ to 96 hours after irradiation, as well
as at one day to several months after birth. He found 100 per cent
brain damage and usually also damage in the retina, cord, and ganglia
following doses of 200, 400, or 600 r. With 150 r, the degree and incidence
of damage was less; and no necrosis at all was found below 100 r. Acute
stages of necrosis were observable as early as 2 hours following irradiation.
The periventricular neuroblasts and other regions of rapid growth in the
brain, the neuroblastic layer of the retina, and the dorsal gray columns of
the cord were affected. Changes seen 2 to several days after irradiation
were classed as early malformations. These consist of the formation of
ependymal canals and rosettes in brain, cord, and retina, and are there-
fore interesting in comparison with the results of Kosaka, of Wilson, and

898

RADIATION BIOLOGY

of Pagenstecher. Finally, the author lists almost a dozen "late mal-
developments," namely, nervous system changes found in the postnatal
period. Among these are: virtual absence of corpus callosum (also found
by Grobman — see p. 897), jumbling and reduction of the hippocampus,
malformations of the ventricles. It is interesting that extraneural
damage could be obtained only with the highest doses and that even then
it was rare except in the thymus (cf. Kosaka).

All morphological changes — immediate and delayed — reported for
irradiation during the period of the fetus have been summarized in
Table 13-8.

Table 13-8.

Morphological Changes Following Irradiation during the Period

of the Fetus

Observa-

Author

Affected organ

tion

Gonads

Brain

Retina

Lens

Thymus

Skin

+

+

+

Miscellaneous

Immediate

Kosaka
Hicks

+ +
+ + +

+ +
+ +

+ + +

+

Liver, spleen
Cord, ganglia

Delayed

Bagg
Kosaka

Kaven

Russell

Grobman

Parkes

Hanson

Niirnberger

Hicks

+ +
+ + +

+ +
Sterility
Sterility
Sterility

Cerebrum
Cerebrum

9

Rosettes

Cataract,
atrophy

Cataract
Cataract

Occas. liver,
lung, heart,
kidneys

Corpus callos.

Corpus callos.,
hippocampus,
etc.

Gall bladder
Cord

3. TIME OF PARTURITION

There are a number of scattered reports in the literature to the effect
that irradiation during pregnancy causes a delay in parturition. Since
most of these statements are concerned with irradiation during the
period of the fetus, they will be examined in this section. In rabbits, a
delay of 17 hours to 2 days was found by Sebileau (1906a, b — three cases
out of seven), of 3-4 days by Niirnberger (1920 — three cases out of three),
and of 1-2 days by Lacassagne and Coutard (1923 — three cases out of
nine). In guinea pigs, one instance of slight delay is mentioned by von
Klot (1911) ; de Nobele and Lams (1925) record one case of 3 and one of
8 days delay; and Kosaka (1928c) states that dead fetuses may occasion-
ally be expelled after the expected time. The same is true in rats
irradiated in the third week of pregnancy (Kosaka, 1928b) where the
young may be stillborn 1-3 days late.

RADIATION IN PRENATAL DEVELOPMENT 899

In a class by itself is the claim by Levine (1927) that the irradiation of
pregnant mice lengthened the mating-to-birth interval by from 3 to 41
days in twenty-two of twenty-six cases. Examination of his method
reveals that the author did not realize that (1) conception may occur a
considerable interval after a male and female mouse are caged together,
and (2) the first litter conceived may be completely lost as a result of
early irradiation, so that the litter observed at birth stems from a later
conception. Even where errors are not as crass as this, it should be borne
in mind that any claim of delay must be supported by adequate control
data, since there is considerable variation in length of gestation period
depending on genetic constitution, and even individual variation within
an inbred strain. Moreover, suckling a previous litter may markedly
lengthen the gestation period of the litter being carried. This fact was
not known to earlier authors.

Russell (1950) found that 88 per cent of her control mice and 80
per cent of the irradiated females of the same inbred strain delivered by
approximately 19^ days after conception. The rest of the control
group was born within the following 16 hours and there were no indica-
tions (from the developmental state of the fetuses) that the balance of the
irradiated animals, which was delivered by caesarean section at 19^ days,
would have been born later than that. There is thus no evidence of
delayed parturition following irradiation during the preimplantation
stages and the period of major organogenesis, but critical data are needed
to answer this question for irradiation during the period of the fetus.

4. ISOTOPE STUDIES

It remains to mention two recent studies in which radioactive isotopes
were injected into pregnant females and the effects on the resulting young
studied. It should be kept in mind that isotope treatment differs from
external irradiation in two ways: (1) the period of effective irradiation
may extend for a considerable time following injection; (2) the radiation
source may be concentrated in certain tissues or organs because of
chemical affinities of the isotope or other reasons. Response, therefore,
does not necessarily represent relative degrees of radiosensitivity through-
out the body.

Burstone (1951) studied the effect of localized electron bombardment
produced by the metabolism of P32 in fetal tooth primordia of inbred
mice. Mothers were injected 2-6 days before expected parturition with
5-17 juc/g and the teeth of the young studied 6-28 days after birth. The
following are some of the findings: (1) fetal teeth are more radiosensitive
than the teeth of newborns (or P32 concentrates in them to a greater
degree) ; also, injections 4-5 days before term were more effective than
injections 1-2 days before term; (2) the development of the third molar is
completely inhibited, while that of the first and second molars, which had

900 RADIATION BIOLOGY

already commenced before injection, is greatly modified; (3) newly
differentiated cells are more radiosensitive as shown by greater damage in
the second than the first molar and by effects within different cell layers
of each tooth; (4) doses of 10-17 /zc/g produce general postnatal growth
retardation which is' greater for earlier than for later injections.

Finkel (1947) injected Sr89 and plutonium into pregnant females
(stages not stated). While very little plutonium reached the fetus, Sr89
concentration in the fetus actually exceeded that of the mother if injec-
tion was shortly before term. Both treatments increased the percentage
of stillbirths, and Sr89 produced retardation of growth, fragility, bending
and shortening of the long bones, anemia, and osteogenic sarcoma.

III. MECHANISMS OF RADIATION EFFECT
ON THE MAMMALIAN EMBRYO AND FETUS8

A. INFLUENCE OF THE MATERNAL ORGANISM

One question that has been raised several times in the literature is
whether radiation affects the conceptus directly or produces an altered
condition in the maternal organism which indirectly damages the embryo
or fetus. Archangelsky (1923) suggested that this altered condition
might consist of changes in uterine tissues, alterations of hormonal output
of the ovaries, or the production of "roentgen leukotoxins" first postu-
lated by Linser and Helber (1905). Obviously, any debilitated condition
of the mother could affect viability of the embryo, and several reports
from the early investigations in this field — when preoccupation with this
particular point was greatest — seem indeed to indicate an indirect influ-
ence on viability of the embryo. In most of this early work, however, the
doses were extremely high, often high enough to kill the mother before
expected term, and factors of shielding may have been incompletely
understood. Fellner and Neumann (1907) and Saretzky (1908), who
shielded the uterus and irradiated the ovaries of rabbits in the first half of
pregnancy, both reported death of embryos (resorption and abortion,
respectively). Von Hippel and Pagenstecher (1907) obtained almost as
poor a yield of living young at term when the abdomen was shielded as
when total-body irradiation was given to rabbits 7-12 days pregnant.
They also injected blood from an irradiated animal into one nonirradiated
doe presumed to be 9 days pregnant and report that no young were born
and that maternal toxins were therefore responsible for prenatal death.
Cohn (1907) shielded all but the head of rabbits 10-20 days pregnant and

8 Since this chapter went to press, the mechanisms of radiation effect on the mam-
malian embryo have been more extensively and systematically discussed in another
publication (L. B. Russell and W. L. Russell, 1954).

RADIATION IN PRENATAL DEVELOPMENT 901

observed poor postnatal development of the young. (This may very
well have been due to a disturbance of maternal lactation brought about
through radiation injury of the pituitary.) Driessen (1924), who irradi-
ated only the left half of the abdomen, found that the more severely
affected embryos were as often in the shielded horn as in the exposed one.
The work of Wilson and Karr (1950, 1951), in which only individual
implantation sites were exposed, should make it clear that a high inci-
dence of prenatal death can be caused, even with relatively low doses, by
direct action on the embryo, though indirect effects of certain degenera-
tive changes produced in the placenta (Kosaka, 1927) can even here not
be definitely excluded. The possibility of interaction between embryos of
the same pregnancy has not been considered in any experiment to date.
It is, however, quite conceivable that radiation-induced death of several
individuals within a uterus may adversely affect the remainder of the
litter, the viability of which was not directly influenced by radiation.

As far as the production of abnormalities is concerned, there seems little
doubt that the maternal body need not act as an intermediary in causing
damage to the embryo, for similar changes have been produced in ovip-
arous forms. Moreover, in mammals, the methods of several investi-
gators, reporting a variety of malformations, have included shielding of
either the anterior half of the mother (Murphy and de Renyi) ; or of all
nonabdominal regions (Kaven; Warkany and Schraffenberger) ; or of
everything except a selected implantation site (Wilson and collaborators) ;
or part of an embryo (Raynaud and Frilley). In investigations where
part or all of the mother was exposed, it is, of course, not inconceivable
that a few of the embryonic abnormalities had an indirect causation, but
two pieces of circumstantial evidence, added to the evidence derived from
analogy with oviparous forms, make it likely that the majority of abnor-
malities are due to the action of radiation on the embryo itself: (1) since
"radiation sickness" of the mother is not sharply limited to a day, any
major influence of maternal pathological conditions on the development
of the embryo would presumably result in a much more blurred relation
between time of disturbance and effect than is actually encountered in the
well-defined critical periods (Russell, 1950) ; (2) it is unlikely that effects
via the mother would become apparent within 2 hours (Hicks, 1950, for
changes in neuroblasts).

The only serious evidence against the above arguments is presented by
Job et at. (1935) who obtained only normal offspring (192) from 22 rat
females irradiated with shielding of the anterior half, while 23 other
females, irradiated in the same stages of pregnancy and receiving approxi-
mately the same dose of whole-body irradiation, produced 73 abnormal
young among 175. This result is puzzling in the light of later evidence
by Warkany and Schraffenberger, and by Wilson and collaborators, who
dealt with the same organism in comparable stages of pregnancy.

902 RADIATION BIOLOGY

B. NATURE OF THE PRIMARY DAMAGE AND INTERMEDIATE EFFECTS

Assuming that at least a large part of the damage must be due to the
action of radiation on the embryo itself, there arises the more difficult
task of elucidating the nature of the primary damage and the pathways
that lead to the observed malformations. Although there is little direct
evidence on these points available for mammalian embryos, the hypoth-
eses are limited by two end results which have been discussed in preceding
sections and must now be considered well established: (1) irradiation at a
given stage causes observable changes only in certain specific characters,
and (2) a primordium affected at any given stage responds in consistent
ways. These points will be considered in turn.

(1) The first end result must be due either to differential primary
damage, or, if the primary damage was randomly distributed throughout
the body, a differential intermediate effect. A few available facts indi-
cate a possible basis for selectiveness at either step. There is evidence to
show that some of the processes specifically affected at any given stage
are those engaged in a rapid rate of change at the time. A few examples
are: vertebral jumbling — beginnings of primitive streak activity (Russell,
1949) ; brain hernia and pseudencephaly — early brain differentiation
(Kaven, 1938b) ; microphthalmia — early optic evagination (Wilson and
Karr, 1951); coloboma — invagination of optic vesicles (Russell, 1950);
digital reductions — limb bud rudder formation (Russell, 1950); second
phase for tail reduction — tail bud formation (Russell, 1950). There are
also a few examples which show that sensitivity is not always predictable
from the visibly fastest rate of change in a primordium: e.g., the critical
periods for Polydactyly and the first phase of tail reduction occur before
limb buds and tail bud, respectively, have made their appearance (Russell,
1950) ; a high incidence of liver damage is produced by irradiating at a
stage when the hepatic primordium is not yet indicated morphologically
(Wilson and Karr, 1951). In these cases some process related develop-
mentally (e.g., an organizer), and not necessarily by cellular ancestry,
may be sensitive at the time of irradiation.

It is conceivable that the primary intracellular damage — e.g., chromo-
some breakage, injury of the spindle-forming mechanism — occurs with
uneven distribution, possibly dependent on relative proportions of cells
in mitosis in various regions. In addition to differential distribution of
the primary intracellular (and the subsequent cellular) damage, various
intermediate selective mechanisms on a higher level can be postulated.
Thus, for example, the different degrees of differentiation reached by
various precursors at the time of irradiation may be correlated with
different capacities for regeneration. Or, even a slight upset in cellular
balance in a region providing trophic influences for a certain primordium

RADIATION IN PRENATAL DEVELOPMENT 903

may inhibit proper differentiation of the latter but not at all affect other
processes.

(2) Having discussed distribution of damage within the body, it is
necessary to examine the quality of the change in affected cells in order
to arrive at an explanation for the consistency of end results. In the
reviewer's opinion, this consistency for any given primordium and stage
of irradiation is almost certainly due to an invariable response of cells
(e.g., death) which have undergone a random type of primary change
within them (e.g., any of a number of chromosome aberrations). The
alternative hypothesis, namely, that consistency results from primary
damage selective within the cell, is hardly tenable. Although it is
possible to' postulate mechanisms that would give a certain degree of
intracellular specificity, e.g., greater susceptibility of terminal regions of
chromosomes, or a maximum breakage frequency in the longest chromo-
some, this type of directed damage cannot be expected to change with
stage and type of precursor nor can it be sufficient to account for
consistency in a vast number of phenotypic changes. This argument
automatically eliminates Wilson and Karr's (1950) suggestion of "subtle
genie alterations" leading to failure "to follow the prescribed course in
differentiation" — quite apart from the fact that several reasons make
gene mutations unlikely as an effective primary change.

Among the possible classes of primary damage are chromosome break-
age, damage to the mitotic mechanism, and gene mutations. The last
named is, on the basis of frequency alone, unlikely to cause many abnor-
malities. Thus, with currently available mammalian germinal mutation
rates (W. L. Russell, 1951), and assuming the same somatic rate and
2 X 20,000 loci, 200 r would give an average of two mutations per cell.
The average degree of dominance of these radiation-induced mutations is
presumably quite low and only very few cells would thus be affected in
diploid tissues. (Of course, embryonic material can be used for the study
of somatic mutations, but special genetic techniques are necessary for this
purpose — Russell and Major, 1952.)

A number of mechanisms may be postulated to account for the con-
sistent nature of the final change, in spite of the random quality within
the cell of the primary damage. These include (1) dominant lethal
action of chromosomal aberrations which lead to aneuploidy after mitosis
and thus to cellular death; (2) retardation in mitotic rhythm brought
about by any damage to the mitotic mechanism or by certain types of
chromosome aberrations; and (3) change in developmental potency,
provided there is only one possible abnormal path for the cell, regardless of
the type of aberration it contains ; this condition is probably met only in a
small number of instances, if ever. That selective cellular death occurs
following prenatal irradiation and often after only short intervals has

904 RADIATION BIOLOGY

been shown by Hicks (1950) and by Kosaka (1927, 1928b, c, d), who
describe its various histological manifestations in detail. Temporary
mitotic inhibition is harder to demonstrate but the work on other organ-
isms, reviewed in Chap. 10, makes it likely that it can occur in mammals
also. In this connection, it should be pointed out that arrests in whole
tissues or organs, which have been reported by Wilson and Karr (1951),
and which may later turn into abnormality, need not be due to mitotic
inhibition but may be caused by death of a certain proportion of cells at
a stage preceding observation. Abnormalities involving an apparent
increase rather than defect, e.g., Polydactyly, or the formation of an
extra thoracic rib, are quite reconcilable with the idea that the initial
change is death or retardation of cells rather than acceleration: thus the
selective elimination of a region may divide a primordium, and result in
"twinning." Radiation effects on viability (except during cleavage
stages, which form a special problem because of the apparently great
regulatory powers of blastomeres) may be thought of as resulting from
damage to various key tissues or organs (e.g., liver, as suggested by
Wilson and Karr, 1951). It is then obvious that incidence as well as
time of mortality would vary with the stage irradiated, i.e., with the
different critical periods existing at the time of treatment.

The work of Russell, Russell, and Major (1951) has shown that hypoxia
protects markedly against radiation-induced abnormalities. By obtain-
ing results at a particular stage for a variety of dosages both in 5 per cent
oxygen (+ 95 per cent helium) and in air, it could be demonstrated that
the magnitude of protection is approximately similar for all of six charac-
ters so far tabulated (see Fig. 13-9). Since hypoxia has been shown to
protect against chromosome aberrations (Giles, Baker, and others) and
mitotic inhibition (Gaulden and Nix, and others), it may be suggested
that the protection against developmental abnormalities is on the level of
the intracellular damage, but the results do not provide evidence in favor
of any one kind of intracellular damage. The fact that various characters
are protected to an approximately equal degree makes it likely that
different sensitive primordia are affected in a similar manner by radia-
tion to yield abnormalities. It is also noteworthy from the point of view
of the discussion which follows that, both in incidence and degree of
change, reduction in oxygen is equivalent to treatment with a lower dose.

C. DOSAGE RELATIONS; VARIABILITY

The commonest results obtained on increasing the dose are: (1)
increase in incidence, (2) increase in degree, and (3) extension of the
period during which radiation will yield the given abnormality (Russell,
1950, 1949). Exceptions to these points will be discussed. If the prob-
ability of affecting a potentially sensitive cell in a given precursor of n
cells is p, then the proportions of animals with 0, 1, 2, 3, . . . ,n cells of

RADIATION IN PRENATAL DEVELOPMENT

IOO-» •-

905

400

O O 5% 02 + 95 % He ;

AIR

Fig. 13-9. Quantitative comparison between the effects of prenatal irradiation of the

mouse in an atmosphere of 5% 02 + 95% He (O O) and in air (• •). All

treatments (0, 100, 200, 300, and 400 r) were administered on day 11^ postconception
and all observations made at birth. (Russell, Russell, and Major, 1951.) a = mean
birth weight; b = mortality; c = tail length; d = tail shape; e = forefeet; / = hind
feet.

906 RADIATION BIOLOGY

the precursor affected are given by the successive terms of the binomial

expansion [(1 — p) + p]". It may be assumed that precursors with a

x
proportion of from 0 to - cells altered can still develop normally, while

ft

x + 1
those with a proportion of from — to all cells altered will give rise to

a changed character, the degree of abnormality being determined by the
number of stricken cells. The larger p becomes on raising the dose,
(1) the relatively smaller will be the proportions of animals in the classes
with 0, 1, 2, . . . , x cells affected, i.e., the greater the incidence of
abnormality, and (2) the larger will be the mean, pn, i.e., the higher the
average degree of abnormality. The result, mentioned in the preceding
section, that irradiation under hypoxia is equivalent in end result to
lower dose treatment in air, indicates that for any given dose, p is lower
under hypoxia, since n and x are presumably constant. The fact that
irradiation at a stage subsequent to that of maximum sensitivity may still
yield abnormality with a higher dose can be explained either by decrease,
with age of precursor, in p for a given dose because of some biological
reason, e.g., slowing in rate of mitosis, or merely by increase in n (pro-

x
vided - > pn), or by a combination of these factors.

The above statistical considerations on incidence and degree show that
there could be variability in results even in perfectly uniform material, a
fact which has not generally been recognized. As it is, however, such
variability would be superimposed on three biological variables: (1)
genetic variability, which can, however, easily be controlled by the use of
inbred material ; (2) environmentally determined variability existing even
in genetically homogenous material of a given developmental age;
(3) differences in developmental age between and within litters of a given
chronological age (Allen and MacDowell, 1940) ; these probably account
in part for the finding that certain abnormalities are obtainable with low
incidence by irradiating on the days adjacent to the main critical period.

The effect of genetic constitution on radiosensitivity to the induction of
certain abnormalities has been studied by Russell and Russell (1950b),
choosing characters (homeotic shifts in vertebral borders and related
changes in the thorax) in which there is normal variability between as
well as within inbred strains. Genetic constitution determines the loca-
tion of the strain on a scale of developmental potencies, while environ-
mental factors, mostly intangible, cause individuals to be distributed
about this mean. Because of thresholds of expression, the finally
observed characters fall into alternate categories. Data indicate that
differences between three strains in the final visible result of a given dose
of radiation — and thus in the apparent ease of radiation shift — are not
attributable to differences in the underlying effect. For example, 200 r

RADIATION IN PRENATAL DEVELOPMENT 907

on day 8H postcohception increases the presacral number to 27 in 100 per
cent of BalbC strain animals, in only 3 per cent of (C57 X NB)Fi's, and
in 0 per cent of 129 strain mice. However, control results show the
BalbC and 129 distributions to be situated across the 2%q thresholds,
respectively, while the (C57 X NB)Fi distribution crosses neither thresh-
old. The positions are such that equal radiation-induced shifts in
the mean, on the scale of underlying processes, would account for the
results. In the course of this experiment it also became apparent that
for the changes studied, i.e., quantitative characters in which even slight
shifts are observable, a dose as low as 25 r may be shown to have an
effect on prenatal development.

Although the commonest result of an increase in dose is increased
incidence and degree of abnormality, there are a few exceptions which
point to the possible existence of interesting developmental pathways.
In the case of microphthalmia, for instance, the results of both Russell
(1950) and Wilson and Karr (1951) indicate a lowering of incidence as the
dose is raised. This can be explained by some other abnormality "com-
peting" with microphthalmia at the higher dose but produced only to a
slight extent or not at all at the lower. — Certain primordia are apparently
capable of only one type of response since degree does not change with
dose level even though incidence does. Examples are Polydactyly, and
hydro-ureter (Russell, 1950).

D. COMPARISON WITH OTHER AGENTS AFFECTING DEVELOPMENT

Radiation-induced developmental abnormalities may be compared with
those produced by other agents (Haskins, 1948; Gillman et al, 1948;
Hamburgh, 1952; Waddington and Carter, 1952; Fraser and Fainstat,
1951a; Kalter and Fraser, 1952; see also review by Fraser and Fainstat,
1951b). Haskins' (1948) results for nitrogen mustard treatment between
days 13 and 16 of rat gestation are similar to (but less extensive than)
those obtained by Warkany and Schraffenberger following irradiation at
the same stages. On the other hand, Waddington and Carter (1952) point
out that trypan blue (used also by Gillman et al. and by Hamburgh) pro-
duces a smaller variety of abnormalities than found by Russell for irradia-
tion at the comparable stage, and the same seems true of the action of
cortisone (Fraser and Fainstat) and of 17-hydroxycorticosterone (Kalter
and Fraser). This may well be due to chemical affinity of these deleteri-
ous agents for certain primordia. It is also of interest that the high
proportion of tail abnormalities found by Waddington and Carter and
by Hamburgh for trypan blue injection on day 7 points to the persistence
of the deleterious action until somewhat later stages, at which X-ray
results have shown the tail to become considerably more sensitive. On
the other hand, cleft palate can be produced by the injections of cortisone
at stages following closure of the nasomaxillary fissure. Fraser and

908 RADIATION BIOLOGY

Fainstat suggest a degenerative change instead of developmental inter-
ference, which seems to occur as a result of other agents.

Comparison of radiation-induced developmental abnormalities with
the changes produced by mutant genes has been made by Russell (1949,
1950). Although a particular mutant gene is probably present in every
cell of the body, primary gene action is in most cases circumscribed by
conditions arising in the course of differentiation. It may be limited to
one process, depending on the identity of which the ultimate pheno-
typic effect may be either circumscribed or widespread. Or the primary
gene action may be widespread, in which case the ultimate expression is
even more likely to be a whole syndrome of changes. In any case, it is
unlikely that gene action, evoked, as it is, by certain conditions in differ-
entiation, should exactly parallel the pattern of radiation effect which is
probably determined by some generalized state (e.g., high rate of mitosis)
of a variety of precusors. It is, therefore, not surprising that there are
no cases of perfect correspondence between all the changes brought about
by a given gene and the various abnormalities produced by radiation at a
given stage. On the other hand, similarity of certain details may point
to the weak links in some of the various developmental chains coexisting
at a given moment and should indicate that at least some of the secondary
gene effect occurs at the time indicated by the radiation effect. Another
possible difference between mutant action and radiation is that, within
the affected tissue, the gene may act in every cell, radiation probably only
in a certain proportion. This latter property of radiation damage may
also differentiate it from the action of physiological poisons, e.g., certain
chemicals, and explain its failure to date to produce many of the classical
abnormalities — cyclopia, twinning, otocephaly.

IV. CLINICAL LITERATURE ON THE EFFECTS OF RADIATION

ON EMBRYO AND FETUS;

HUMAN IMPLICATIONS OF EXPERIMENTAL WORK

The case literature on the subject of prenatal radiation effect is large
and diffuse but various summaries facilitate a survey of the field, even
though they suffer from a certain amount of overlap. In order of
appearance, these are by von Klot (1911), Driessen (1924), Murphy
(1929), Goldstein and Murphy (1929) (the same material is also found in
Murphy, 1947), Gauss (quoting a thesis by Kraemer, 1930), Flaskamp
(1930), Schall (1933 — thorough tabular representation), Miller et al.
(1936), and Jones and Neill (1944). Cases not included in the above sum-
maries are by Lindenfeld (1913) and Murphy et al. (1942). Russell and
Russell (1952) have discussed the implications for medical practice of
findings in experimental animals.

Although reports of normal children following prenatal irradiation are

RADIATION IN PRENATAL DEVELOPMENT 909

given, among others, by Robinson (1927 — twenty-three cases collected
from the literature), Lacomme (1931 — two cases), Jones and Neill (1944 —
seven definite and twenty-eight questionable cases), and Hobbs (1950 —
one case), and have been stressed by various authors as showing that
damage need not invariably or even frequently follow prenatal irradiation,
there can, of course, be no question, both from experimental and clinical
findings, that the human embryo is subject to severe radiation injury.
The types of human abnormalities enumerated in the case literature
include microcephaly, blindness, microphthalmia, coloboma, cataract,
chorioretinitis, ankyloblepharon, strabismus, nystagmus, mental defi-
ciency, hydrocephaly, coordination defects, mongolism, spina bifida,
skull malformations, ossification defects of the head, cleft palate, ear
abnormalities, deformed arms, club feet, hypophalangea, genital deform-
ities, and general mental and physical subnormality. Microcephaly is
present alone or in combination with other changes in 16 of 28 abnormal
cases listed by Goldstein and Murphy (1929), and in 27 of 38 tabulated
by Schall (1933). It is probable that this classification includes a
variety of head abnormalities. Next on the list in Schall's tabulation
are various eye defects (14 of 38 cases). Central nervous system abnor-
malities other than microcephaly are frequent as a group. Thus, the
tissues believed to be radioresistant in adult humans are especially sensi-
tive in the embryo.

By equating human and mouse gestation periods developmentally
(rather than chronologically, since, e.g., the first one-fourth of mouse
prenatal life is equivalent to only the first one-thirtieth of human) , as has
been done by Otis (1952), it should be possible to predict when the
critical period for the production of abnormalities determined in the
mouse (see preceding sections) should occur in man. This type of predic-
tion is substantiated by one of the few human cases where stage of irradia-
tion is accurately known (Feldweg, 1927) : irradiation during the fourth
or fifth week caused arm abnormalities of the newborn similar to those
which can be produced in the mouse by treatment at the corresponding
stage, day 10^2- The period of major organogenesis in man, correspond-
ing to days 6^ to 13^2 m the mouse, comprises weeks 2-6. It may be
predicted from the animal results that irradiation during those stages will
lead to a high percentage of conspicuous abnormalities, while later
irradiation will yield a lower incidence, at least insofar as immediately
recognizable changes are concerned. Confirmation comes from the
tabulation of Kraemer (1930) who found that all of 11 cases of irradiation
within the first two months resulted in damage, while only 7 out of 11
(64 per cent) irradiated between the third and fifth months and 3 out of
13 (23 per cent) irradiated between the sixth and tenth months were
abnormal at or near term. In Schall's (1933) tabulation, which, in
contrast to Kraemer's, lists only positive cases, 57 per cent (20 of 35) were

910 RADIATION BIOLOGY

irradiated between conception and the third month, 31 per cent between
the third and fifth months, and 11 per cent after the fifth month. In
Goldstein and Murphy's compilation (1929), 79 per cent of the abnormals
come from pregnancies irradiated before the fifth month.

Russell and Russell (1952) have suggested that, since during at least
the early part of the period of major organogenesis, many women are not
yet aware that they are pregnant, pelvic irradiation of women of child-
bearing age should whenever possible be restricted to the two weeks
following a menstruation as there is little chance of an unsuspected preg-
nancy during that time. This type of timing should be feasible for
diagnostic irradiation (see following paragraph for discussion of dose), if
not always for therapeutic. The dangers of irradiation after the period of
major organogenesis, which, predicting from the animal results, consist
of the production of possibly delayed and less obvious but, from a
human point, at least as undesirable effects (e.g., changes in mental
abilities, sterility), occur at a time when there need no longer be any
doubt of pregnancy. They are, therefore, already avoided in good cur-
rent practice.

It was mentioned earlier (see p. 907) that in experimental work a dose
as low as 25 r, the lowest used, may be shown to have an effect on prenatal
development (Russell and Russell, 1950b), and there is no evidence that
lower doses would not also cause damage detectable in man where even
subtle defects are likely to be recognized. This is of significance in
medical diagnostic practice since Sonnenblick et at. (1951) found that
about one-fifth of sixty-three fluoroscope machines tested emitted^ more
than 30 r per minute, and Bell's results (1943) indicate that the length of
exposure for a standard gastrointestinal series is 4^2 to 12 minutes, during
which time about 50 r may be received at a depth of 10 cm for a panel
dose of 30 r/minute. On the other hand, doses received by the uterus
in radiographs are comparatively negligible (Mobius, 1951).

The question of feasibility of radiation-induced therapeutic abortion
arose very soon after the beginning of radiological practice. As early as
1911, von Klot discusses its pros and cons at considerable length. Dries-
sen (1924) cites seven references which claim success and seven others
which state X rays to be unsuitable for the induction of abortion in man.
Murphy (1929) showed that the abortion rate following postconceptional
irradiation was probably no higher than in the population at large. In
his review, Schall (1933) states that radiation abortion has finally been
discontinued in medical practice because of inconstancy of results; but,
three years later, Mayer, Harris, and Wimpfheimer (1936) recommend
the method and claim a very high percentage of success. The statement
has occasionally been made (Robinson, 1927, and others) that doses high
enough to cause maldevelopment usually also terminate pregnancy and
thus do not present an appreciable hazard. Whether the unreliability

RADIATION IN PRENATAL DEVELOPMENT 911

in success of X-ray abortion is due to variability in the physical factors
of irradiation or to biological variability in the subjects cannot here be
determined, but it is clear, both from the animal results discussed in
earlier sections and the human cases summarized above, that damaged
embryos or fetuses do come to term in a large number of instances. Sur-
vival of these monsters is, moreover, fairly frequent. Murphy (1929)
found no stillbirths among seventy-four cases of postconceptional irradi-
ation which went to (or near) term; yet 34 per cent of the children were
grossly deformed. Postnatal viability may be estimated from the lists
of Goldstein and Murphy (1929) and of Schall (1933) : only 7 of 26 (27 per
cent) and 11 of 38 (29 per cent), respectively, of the malformed children
died between birth and two and a half years of age ; the rest, alive at the
last report, ranged from several weeks to twelve years with about two-
thirds of the group (in Schall's list) two or over.

The possibility of damaging the human conceptus indirectly through
radiation injury of the mother has been stressed by Flaskamp (1929, 1930)
and by Faerber (1933), who report abnormal children following irradiation
during pregnancy to nonabdominal sites or with the uterus shielded.
Schall (1933) evaluates the four cases cited by Flaskamp and three addi-
tional ones and comes to the conclusion that, in at least five of the seven,
the uterus probably received scattered radiation. Results with experi-
mental animals (discussed earlier) make it likely that at least the
majority of the abnormalities are due to the action of radiation on the
embryo itself and would thus be avoided by appropriate shielding if non-
abdominal regions of a pregnant woman had to be irradiated.

SUMMARY

1. Radiation has proved to be an excellent tool in mammalian experi-
mental embryology and radiation-embryological studies on mammals
have yielded results which have important clinical implications.

2. Most of the experimental work in the field has been concerned with
rodents and with radiation from an external source. Various degrees of
shielding and localization of the beam have been used, ranging from none
at all to the exposure of only selected parts of individual embryos.
Almost all of the early experiments, and even a few of the more recent
ones, suffer greatly from lack of timing of the developmental stage at
which radiation was applied.

3. Recent experiments have shown that the stages at which radiation
is applied to the embryo can be conveniently grouped by broad end result
into (a) the preimplantation period, (6) the period of major organogenesis,
and (c) the period of the fetus. Experiments concerning these three
periods are reviewed in turn.

4. Irradiation during the preimplantation period causes a high rate

912 RADIATION BIOLOGY

of prenatal mortality. In the mouse, the incidence for a dose of 200 r is
43-79 per cent, depending on the stage irradiated. Death may involve
the whole litter, terminating pregnancy prior to implantation, or it may
occur in individuals in surviving pregnancies both before or shortly after
implantation. The relative proportions of the different types of death
depend on the stage irradiated. Virtually all survivors are normal.

5. For comparable doses, irradiation during the early part of the
period of major organogenesis causes considerably less prenatal death
than does treatment during the preimplantation period and, from then
on, susceptibility to prenatal killing probably decreases rapidly with
embryonic age. On the other hand, neonatal death assumes consider-
able importance and, in the mouse, is at a peak following irradiation on
days 93^> and 10^. The LD50 at birth for prenatal irradiation varies
markedly with the stage irradiated, even when stages are separated by
only 24-hour intervals.

6. A vast number of varied abnormalities have been reported as result-
ing from the irradiation of mammalian embryos during the period of
major organogenesis. A few critical experiments have shown that sensi-
tive periods for the induction of almost all abnormalities are short: with
200 r, many of them are restricted to one particular day, or, if they span
several days, one often stands out as the main sensitive one on the basis
of incidence. There are several cases of good agreement of different
authors on the critical periods for certain characters.

7. Irradiation during the period of the fetus does not cause the striking
changes in newborns produced by comparable doses during the period of
major organogenesis. Immediate histological damage can, however, be
demonstrated and this leads to gross changes which become expressed
later in life. The most frequently reported defects are in the brain,
retina, gonads, lens, skin, and thymus.

8. Proportional growth retardation leading to decreased body size can
be produced by irradiation at almost any stage after implantation. In
the mouse, the maximum reduction in birthweight for any given dose
follows irradiation between days 10^2 and HJ^. Birthweight depression
for treatment on day 11L2 is directly proportional to dose, averaging
0.22 g per 100 r.

9. Claims of upset sex ratio following prenatal irradiation and of
differential susceptibility of the sexes to the induction of abnormalities
cannot be considered proved, although it is conceivable that such effects
will be found. The same is true of claims that prenatal irradiation pro-
longs the gestation period.

10. Although high doses to the mother may indirectly affect the viabil-
ity of the conceptus, there seems little doubt that most of the major
embryonic abnormalities are due to the action of radiation directly on
the embryo.

RADIATION IN PRENATAL DEVELOPMENT 913

11. The finding that irradiation at a given stage causes changes only
in certain characters may be explained by either or both of the following:
(1) differentially distributed cellular damage; (2) selective mechanisms on
a higher level. The basis for (1) may be differential mitotic activity.
The fact that a precursor affected at any given stage responds in con-
sistent ways may be explained by the invariable response of cells which
have undergone a random primary change within them. The most
plausible mechanism is cell lethal action of somatic chromosomal aber-
rations which lead to aneuploidy after mitosis.

12. Hypoxia protects markedly against radiation-induced abnormal-
ities to an approximately equal extent in several characters. Protection
is probably on the level of the intracellular damage.

13. Although there are several exceptions, the commonest results
obtained on increasing the dose are increase in incidence, increase in
degree, and extension of the period during which radiation will yield the
given abnormality. These results are as expected on the assumption
that a threshold proportion of cells must be affected in a primordium in
order to produce any abnormality and that, above this threshold, the
number of cells affected determines degree of abnormality. Simple sta-
tistical considerations show that there could be variability in result even
in perfectly uniform material. This is superimposed on genetic variabil-
ity, environmental variability, and subtle differences in developmental
age among embryos of a given chronological age at the time of exposure.

14. There are certain similarities but also marked differences between
the action of radiation on the one hand and of other deleterious agents or
of genes on the other.

15. The clinical literature includes many cases in which abnormal
children have been born following irradiation of the pregnant mother.
In applying experimental results to man, it becomes apparent that
critical periods for the majority of gross abnormalities occur at a time
which corresponds to weeks 2-6 in human gestation. During at least
part of this time, pregnancy may still be unsuspected. It has, therefore,
been recommended that, whenever possible, irradiation involving the
uterus in women of childbearing age should be restricted to the two weeks
following the menses, to preclude the possibility of fertilization having
taken place. Experimentally, doses as low as 25 r have been shown to
be effective in producing developmental changes if applied at the critical
time. This is within the dose range used in diagnostic fluoroscopy.

914 RADIATION BIOLOGY

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par les rayons de rontgen. Arch, electricite med., 343: 321-334.
Robinson, M. R. (1927) The effect of a castration dose of roentgen rays upon the

rabbit ovary. An experimental study with a clinical evaluation of the problem

of ovarian irradiation. Am. J. Roentgenol. Radium Therapy, 18: 1-25.
Russell, Liane Brauch (1949) X-ray induced developmental abnormalities in the

mouse and their use in the analysis of embryological patterns. Thesis, University

of Chicago.

(1950) X-ray induced developmental abnormalities in the mouse and their

use in the analysis of embryological patterns. I. External and gross visceral
changes. J. Exptl. Zool., 114: 545-602.

and Mary H. Major (1952) A preliminary report on radiation-induced

presumed somatic mutations in the house mouse. Genetics, 36: 621.

and W. L. Russell (1950a) The effects of radiation on the preimplantation

stages of the mouse embryo. Anat. Record, 108: 521.

and (1950b) Changes in the relative proportions of different axial

skeletal types within inbred strains of mice brought about by X-irradiation at
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and (1952) Radiation hazards to the embryo and fetus. Radiology,

58: 369-376.

and (1954) An analysis of the changing radiation response of the

developing mouse embryo. J. Cellular Comp. Physiol., Suppl. (In press.)

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Therapie im Kindesalter. Georg Thieme, Leipzig, pp. 567-577.

918 RADIATION BIOLOGY

Schinz, Hans R. (1923) Der Rontgenabort. Zugleich ein Beitrag zum spontanen
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Manuscript received by the editor, June 3, 1952

CHAPTER 14

The Pathological Physiology of Radiation Injury

in the Mammal. I. Physical and Biological

Factors in Radiation Action

Harvp:y M. Patt and Austin M. Brues

Division of Biological and Medical Research, Argonne National Laboratory,

Lemont, III.

Introduction. Nature of the biological response to radiation. Radiation quality and
quantity. Biological factors in radiosensitivity: Cell and tissue sensitivity — Species
sensitivity — Individual sensitivity. Induced radiosensitivity or resistence: Temperature
and metabolic rate — Intermediary radiochemical events. Summary. References.

INTRODUCTION

When in 1895 Professor Rontgen reported on a "new form of radiation "
to the Wurzburg Physico-Medical Society, the stage was set for an
energetic, though initially somewhat chaotic, development in biology.
The discovery of radioactivity by Becquerel in 1896 lent further impetus
to this development, although the connection between the two forms of
radiation was not immediately apparent. Alopecia following X-ray
photography was reported by Daniel (1896) four months after Rontgen's
announcement, and the severe superficial injuries of a journeyman
exhibitor of X-ray machines were eloquently described in the August 12,
1896, issue of the Electrical Review. Radiation dermatitis and epilation
were observed by Leppin (1896) and Stevens (1896) at about the same
time. In 1897, Freund reported the successful removal of a hairy mole
with X rays, and Walsh described the unpleasant symptoms of acute
radiation illness. Two years later a report on treatment of skin cancer
with X rays appeared (Stenbeck, 1899), and in 1902 the earliest case of
radiation-induced cancer was recorded (Frieben). Perhaps the first
observation of an internal effect may be attributed to Senn, who described
a decrease in size of the spleen in 1903.

As a consequence of these observations and the pressure of growing
clinical usage, the biological action of ionizing radiation on animals and
man was widely studied, and, although the experiments took place
sporadically, with great variation in methods of exposure and dosage,
many fundamental facts soon emerged. Among these were the selective

919

920 RADIATION BIOLOGY

action of radiation on different parts of the same cell and on different
cells; the relation between differentiation, mitotic activity, and radio-
sensitivity; the importance of intensity-duration considerations in the
radiation effect; and the latency of most biological responses.

Although many of the important effects of ionizing radiation were
reported over 25 years ago, radiation dosimetry unfortunately did not
keep pace with the biological approach. Advances in radiation physics
and the availability of many different sources of radiation, culminating in
the development of the cyclotron and other accelerators and the chain
reactor, have now made possible a quantitative reinvestigation and
amplification of the earlier qualitative observations and have, in addition,
opened new avenues for investigation. It is our intention to present a
comprehensive, though not necessarily encyclopedic, appraisal of the
pathological physiology of radiation injury in the mammalian organism.
The existence of several excellent reviews of the early literature has
greatly simplified our task. Although we have drawn upon some early
observations, our source material has largely been restricted to the more
recent papers and to the Manhattan Project and Atomic Energy Com-
mission documents. For convenience, the presentation has been divided
into two parts, the first being concerned with the physical and biological
factors in radiation action and the second with the specific aspects of the
physiology of radiation injury.

NATURE OF THE BIOLOGICAL RESPONSE TO RADIATION

High-energy radiations dissipate their energy in tissue by ionization
and excitation. The dependence of biological effectiveness upon the
specific ionization or ion density of a particular radiation lends strong
support to the idea that its action is related in some manner to direct
local release of energy, presumably through ejection of electrons from
the atoms through which it passes. While the role of excitation is not
well defined, it may constitute an important secondary, if not primary,
process. An interesting aspect of the energy absorption is the relatively
small absolute amount that is required to produce widespread effects.
One thousand roentgens, a lethal dose for most mammals, corresponds to
an energy absorption of only 2 X 10-3 calorie per gram. We may
examine the problem in another way by computing the fraction of
molecules within a cell that is likely to be modified as a direct result of
the radiation. In a cell containing about 1014 molecules, 1000 r would
be expected to ionize only 107 molecules. Although it is likely that
many more molecules will be affected indirectly in consequence of the
energy transformations resulting from the absorption of radiation, the
total number of altered molecules is probably a small fraction of those
present within a cell. It would seem, moreover, that not all the damaged

\

PHYSICAL AND BIOLOGICAL FACTORS 921

molecules are of critical importance in the cell economy since most of
these would be water molecules. These considerations raise the question
of the localization and amplification of the early critical events.
Although amplification could be accomplished by inactivation of genes or
enzymes, by some disturbance in the synthesis or assembly of essential
substrates, or by the formation of toxic substances, the nature of the
early physicochemical events is obscure and poses the most formidable
problem in radiobiology.

Two main concepts of radiation effects on living systems have been
formulated. These are described most frequently as the theories of
direct and indirect action and have been presented in some detail by
Lea (1946) and others. Direct action postulates that ionization occurs in
specific molecules that constitute a sensitive region or vital target. This
idea in its classical sense is compatible with the single-hit type of effect
that is characterized by its exponential relation to radiation dosage, its
independence of dosage rate, and the inverse relation between its effi-
ciency and ion density. It is perhaps best expressed in the effects on
dried biological materials.

Both the direct and indirect concepts assume that chemical changes are
induced by ionizing radiation, either by the alteration or inactivation of
important molecular species or by the production of substances that
influence cell metabolism (Zirkle, 1949). The indirect effect assumes
that the initial chemical changes are due to highly reactive substances,
mainly oxidants, that are formed at random in the aqueous environment
and subsequently react with critical entities. Thus, localization of an
effect within the living system is secondary and depends upon the nature
of the acceptors as well as upon the original distribution of ions, the
number of reactive substances formed, and the kinetics of their diffusion.
The indirect type of action is dose rate dependent and varies, generally,
directly with ion density. Organic molecules may, therefore, be modified
by direct ionization and excitation or by reaction with the decomposi-
tion products of water. While it has not been possible to demonstrate
these phenomena directly in irradiated biological material, there is
sufficient circumstantial evidence to justify the assumption that they
occur.

Perhaps the outstanding characteristic of radiobiological responses is
their diversity, a not unreasonable situation when we consider that the
radiant energy is absorbed at random in a heterogeneous and highly
integrated system. Thus it is not surprising that many different effects
are observed even on the cellular level and that not all these are in direct
consequence of the radiation. It is probable that many of the seemingly
discordant observations found in the literature are real and merely reflect
the inherent complexity of the chain of events concerned in the radiation
response and that of the responding organism.

922 RADIATION BIOLOGY

This diversity of action increases the probability of nonspecificity, and
so we observe that no single response is peculiarly specific for radiation
injury, while many agents (such as urethane, nitrogen mustard, or
benzol) are capable of mimicking radiation effects. Thus many of the
effects of nitrogen mustard on enzymes and cells are indistinguishable
from those of radiation and point to similar chemical processes (DeLong,
1950). Yet radiomimesis has its limitations in that no single agent
duplicates all the radiation-induced reactions. In part at least this
difference is one of spatial distribution; owing to natural barriers to the
diffusion of a chemical agent from its point of entry and to conditions of
its degradation, it would be almost impossible for such a radiomimetic
agent to attain the uniform distribution of penetrating radiation.

Depending upon the character of the radiation and the manner of
exposure, a variable period intervenes between irradiation and the various
observed effects. This interval may be a matter of minutes for suppres-
sion of cell division, hours for lymphopenia, days for neutropenia and
hemorrhage, weeks for anemia, or months for induction of tumors. Some
of these intervals may be correlated with the life span, mitotic rate, and
metabolic activity of the cells concerned with the effect. Many critical
events, presumably chemical in nature, must occur during this "latent"
period.

RADIATION QUALITY AND QUANTITY

As an environmental change or stimulus, radiation in general conforms
to the familiar concepts of threshold, summation, intensity-duration,
and adaptation. The energy required to produce different biological
effects varies considerably. Retardation of growth of Phy corny ces has
been observed after only 0.001 r, while 1 r is sufficient to inhibit the
activity of aged preparations of adenosinetriphosphatase (Forssberg,
1943; Barron, Dickman, et al., 1949). Hematologic changes in mammals
are seen with dosages of 25 to 50 r. Yet other effects such as the inhibi-
tion of contraction of striated muscle or immediate mammalian death
may require many thousands of roentgens.

In general for any particular radiobiological response, there are limits
within which the quantity of radiation will influence the effect in terms of
latent period, severity, and recovery. In the mouse, total-body gamma-
ray dosages of 140,000 r delivered in 20 min lead to immediate death
(Henshaw, Snider, et al, 1946). With the same dose rate, 70,000 r will
kill in 1 to 5 hours, and 35,000 r in 7 to 62 hours. On the other hand,
after dosages between 3500 and 14,000 r, there is no difference in survival
time; all mice live 4 to 5 days. When the quantity of radiation is
decreased still further to the minimum LDioo and below, survival time
again increases, most deaths occurring at about two weeks. Similar
findings have been observed with X rays and point to a distinct separa-

PHYSICAL AND BIOLOGICAL FACTORS 923

tion in the mode of killing which depends upon the dosage of radiation
(Quastler, 1945a; Quastler et al., 1951; Henshaw, 1944). Death follow-
ing median lethal irradiation is the result of several factors, the most
prominent of which are leucopenia, anemia, hemorrhage, bacteremia, and
intestinal damage. The shocklike syndrome that appears with higher
dosages and results in death within several days is initiated primarily as
a consequence of intestinal injury. Finally, with still more massive
dosages of radiation killing occurs within a matter of minutes or hours
and is associated with disturbances of the nervous system.

A number of factors influence the threshold for different radiobiological
effects. On the physical side these include the quality of the ionizing
radiation, its intensity or rate of delivery, and the manner of exposure,
i.e., local or total-body, external or internal, and single, continuous, or
fractionated. It is generally agreed that a, j8, y, and X radiations and
neutrons produce more or less similar physiological effects. However,
their efficiency varies considerably since the effects of ionizing radiations
depend not only upon the amount of energy absorbed per unit of volume
and of time but also upon its distribution along the individual ion tracks
(Zirkle, 1943; Gray, 1946). Specific ionization (ionization density)
increases progressively from fast /3 rays to y rays, hard X rays, soft X rays,
fast neutrons, and a rays, and for most systems the biological effective-
ness increases with increase in the ionization density. The time factor
of irradiation and the dosage distribution among different tissues may, of
course, contribute to the observed differences in efficiency and should be
considered in their evaluation. The y- to X-ray ratio of effectiveness has
been found to vary from 1.3 to 2.0 for a number of biological effects
(Sugiura, 1939; Lasnitzki and Lea, 1940; Mottram and Gay, 1940).
When the irradiation periods are similar, the efficiency ratio for acute
lethality in mice appears to be about 1.3. This is consistent with the
finding that 200-kv X rays are about 1.3 times as effective as 20-Mv
X rays, which have a specific ionization comparable with that of hard 7
radiation (Quastler and Clark, 1945; Quastler and Lanzl, 1950). The
200-kv X rays are also about 1.4 times as efficient as Na24 /3 rays for acute
killing of mice (Snyder and Kisieleski, 1950) ; it is worth noting, however,
that the actual ratio may be somewhat lower, since the time factor of
irradiation varied in these experiments. For acute effects in mammals,
1 n of fast neutrons (2.5 rep) is equivalent to 3 to 32 r, depending on
the criterion (Lawrence and Lawrence, 1936; Aebersold and Lawrence,
1942; Lampe and Hodges, 1943; Mitchell, 1947; Evans, 1947; Henshaw
et al, 1947; Gray and Read, 1948; Dowdy, 1949). The effectiveness
ratios are generally somewhat larger for periodic or protracted exposures
than for acute, presumably because of a slower recovery from neutron
irradiation. However, for certain effects that are related presumably to
very small volumes (gene mutations or killing of small microorganisms)

924 RADIATION BIOLOGY

the efficiency varies inversely with specific ionization. It is inferred that
only a few ions are required for these all-or-none effects, additional ioniza-
tion being wasted.

Further evidence pointing to the similarity of the effects produced by
the different radiations is given by additivity studies. Obviously, incom-
plete additivity of two radiations would indicate some difference in
mechanism, whereas complete additivity would suggest that at least the
events that are directly responsible for observed effects are identical.
When the energy distribution is similar and the exposure times are rela-
tively brief, complete additivity of the various radiations is observed
(Stapleton and Zirkle, 1946; Zirkle, 1950). If the exposure time is pro-
longed (24 to 48 hours instead of 1 hour) additivity of y rays and fast
neutrons is incomplete (Mitchell, 1947). This may be related to differ-
ences in the recovery times, for there is reason to believe that recovery
may be most rapid with 0 and y rays, less with conventional X rays, and
least with neutrons (Quastler and Lanzl, 1950; Lasnitzki, 1948). This
point requires further amplification. Incomplete additivity is also
observed after external irradiation with beta and hard gamma rays
(Raper, 1947). In view of the great difference in tissue distribution of
these two radiations given externally, one would hardly anticipate more
than a partial additivity. It is of interest that an isotope with osseous
distribution and one with reticuloendothelial distribution are synergistic
with respect to lethality (Friedell and Christie, 1951).

In the production of some biological effects (e.g., gene mutations or
lethal effects in Triton) a given dose produces the same degree of effect
regardless of the rate or time during which it is delivered (Lea, 1946;
Brunst and Sheremetieva-Brunst, 1949). In a few instances, the radia-
tion effect diminishes with very brief exposure times : the lethal effect on
Drosophila eggs exposed to 165 r is reduced by 20 per cent when the
radiation is delivered in 0.4 second instead of 1.2 seconds (Sievert and
Forssberg, 1936). For most responses, especially in mammals, the
effectiveness of a given dose decreases as the rate of exposure decreases.
Yet the events in even this category display reciprocity over a part of
the intensity spectrum, and there is little information relating to very
intense instantaneous exposures, such as might be encountered in an
atomic explosion. Reduction in the biological effect with protraction of
the irradiation is generally explained by assuming that the recovery rate
becomes appreciable during the exposure. Increasing the period of
exposure 10 times reduces the lethal action in mice for a given dosage of
gamma radiation to about 70 per cent (Henshaw et al, 1947). The acute
LD50 of gamma radiation for mice is 840 r when the dosage rate is 30 r per
minute and 1200 r when the rate is reduced to 3 r per minute (Henshaw
et al, 1946). In man, the dosage required to produce an equivalent
cutaneous reaction is doubled when the period of irradiation is increased

PHYSICAL AND BIOLOGICAL FACTORS 925

about 30 times, e.g., from 435 r per minute to 15 r per minute (MacKee
et al., 1943).

When a particular dose is given in several fractions, it is generally
less effective than when it is delivered at one time. Fractionation has
been studied by exposing animals at different daily doses over a specific
interval and comparing the effect produced by the same total dose given
in a single exposure or by continuing the exposures until death. Such
studies provide valuable information concerning lethal mechanisms and
recovery constants and bear directly on the problem of radiation tolerance
(MacComb and Quimby, 1936; Quimby and MacComb, 1937; Ellinger,
1943; Hagen and Simmons, 1947; Sacher et al., 1949; Ellinger and
Barnett, 1950; Sacher, 1950). Differences in rates of recovery may
account for the lower relative efficiency of 20-Mv roentgen rays with
fractionation as compared with that of 200-kv X rays (Quastler and
Lanzl, 1950) or neutrons (Henshaw et al., 1947; Evans, 1947). Although
recovery constants can be calculated for certain radiation effects, they
apparently cannot be used to predict quantitatively the recovery pattern
following various modes of irradiation (Sacher, 1950).

Another aspect of the intensity-duration factor is the production of
different effects with the same total dose. Acute and delayed toxic
effects have been described in chick embryos, chicks, and ducklings
(Karnofsky et al., 1950; Jacquez and Karnofsky, 1950; Steamer, 1951).
Early deaths, within 24 hours, occur with dose rates above 5 to 10 r per
' minute and are relatively independent of a total dose above 800 to 1000 r.
There is evidence that early lethality is associated with renal failure since
uricemia precedes death by several hours (Steamer et al., 1950). Delayed
deaths take place within one to two weeks following irradiation, and the
pathological physiology parallels that seen in mammals. From dose
fractionation studies it appears that the over-all time of exposure is more
important than the dose rate in eliciting the early killing in chicks and
ducklings (Steamer and Christian, 1951). The significance of the over-all
time of exposure has been indicated in other experiments. Dose rate as
such is of less importance than the dose per fraction or the intervals
between fractions in determining the lethal effect of X radiation on chick
fibroblasts (Paterson and Thompson, 1948).

The response to radiation depends upon the portion of the cell or
organism that is irradiated. It has been shown by fractional irradiation
of parts of cells that ions formed in the cell nucleus are more effective in
the production of certain biological effects than those formed elsewhere
(Henshaw, 1938; Zirkle, 1932). Several hundred roentgens applied
locally may be relatively innocuous, whereas a similar dose administered
to all or most of the body leads to widespread effects. Irradiation of the
abdomen is more efficient than irradiation of the thorax, and penetrating
radiations are more effective than the superficial in producing acute

926 RADIATION BIOLOGY

toxicity (Raper, 1947; Bond et al., 1950). It is also apparent that
different mechanisms contribute to lethality, depending upon the area
that is irradiated.

The severity of cutaneous erythema is related to the size of the radia-
tion field, and two fields a distance apart show less injury than areas that
are closer together (Jolles, 1941; Jolles and Mitchell, 1947; Jolles, 1950).
Partial irradiation of the frog's corneal epithelium with 3600 r results in
the same over-all inhibition of mitotic division as total irradiation of the
cornea with 900 r (Strelin, 1950). Thus, injury and recovery are, to some
extent, dependent upon chemical interaction between the irradiated and
adjacent nonirradiated areas. There is also reason to believe that injury
to specific sites is more severe after a total-body exposure than after
local irradiation. This may be attributed to the liberation of non-
specific toxic materials from irradiated tissue and/or to a sparing action
of nonirradiated tissue. Although there is some evidence of active
circulating factors following irradiation, the significance of such humoral
agents is not fully appreciated (Barnes and Furth, 1943; Ahlstrom et al.,
1947; Van Dyke and Huff, 1949; Lawrence et al, 1947; Ellinger, 1951).
Parabiosis, cross-circulation, and early transfusion have been shown to
diminish radiation toxicity (Barnes and Furth, 1943; Van Dyke and
Huff, 1949; Salisbury et al., 1951, Brecher and Cronkite, 1951, Swisher
and Furth, 1951).

Shielding of relatively small volumes of tissues can decrease the
severity of an otherwise total-body exposure. For example, with the
spleen shielded and the remainder of the body irradiated, the 30-day
LD50 for mice is increased from 550 to 975 r (Jacobson et al., 1949). Pro-
tection of the head, the extremities, or other small areas mil also diminish
mortality (Abrams and Kaplan, 1951; Gershon-Cohen et al., 1951;
Allen, 1951). Recovery of hematopoietic tissue is also more rapid after
subtotal irradiation (Boffil and Miletzky, 1946; Rekers, 1949; Jacobson
et al., 1951). The mechanism of these protective effects is poorly under-
stood; Jacobson et al. (1950) have suggested that the mesenchymal tissues
in certain shielded areas may supply a factor that facilitates regeneration
of blood-forming tissue. It is of interest, in this connection, that spleen
transplants or the injection of marrow suspensions or of spleen homo-
genates are effective in reducing radiation mortality in certain species
(Jacobson et al., 1951; Lorenz et al., 1951; Cole et al., 1952). In contrast
to protection by shielding is the synergism that results when certain
radioisotopes, which differ in localization, are administered in combina-
tion. When the reticulo-endothelial system of rats is irradiated with
colloidal Au198 and the bone marrow with P32, lethality is potentiated
(Friedell and Christie, 1951). On the other hand, only an additive
effect is obtained when two bone-seeking radioisotopes are injected simul-
taneously (Salerno et al., 1952).

PHYSICAL AND BIOLOGICAL FACTORS 927

BIOLOGICAL FACTORS IN RADIOSENSITIVITY

Many of our concepts concerning the nature of radiation action are
derived from studies of radiosensitivity and of the factors that influence
it. The selective action of radiation on different parts of the same cell
and on different cells and the relation between differentiation, mitotic
activity, and radiosensitivity were described during the first decade
after Rontgen's discovery of X rays. There followed numerous attempts
to modify sensitivity experimentally. These included the effects of
temperature, oxygen, blood flow, and hydration (Schwarz, 1909; Hol-
thusen, 1921; Petry, 1922; Mottram, 1924; Jolly, 1924). Temperature
studies were especially prominent, since temperature was a convenient
tool, and it appeared that the radiation response was dependent upon
metabolic activity. Unfortunately, there were few attempts to dis-
tinguish the events occurring during irradiation from those taking place
after the exposure. Moreover, the early studies were restricted largely
to isolated cells and tissues. These considerations, along with increasing
knowledge of radiochemical reactions, have led, in recent years, to a
renewed interest in the factors of radiosensitivity.

CELL AND TISSUE SENSITIVITY

The inherent difference in sensitivity of various cells and tissues
attracted early attention, and in 1906 Bergonie and Tribondeau formu-
lated the principle that actively proliferating tissues are the most sensitive
to radiation and that the radiosensitivity of a tissue varies inversely with
the degree of differentiation. While this view has been generally
accepted, it is true only in a broad sense for there are many notable
exceptions. In contrast to the large body of information pertaining to
cell and tissue sensitivity is the paucity of early investigations concerned
with the comparative lethal dose for animals of the same and different
species and with the factors that influence the lethal dose. Our knowl-
edge relating to this aspect of the problem is, therefore, largely a
product of studies that have been conducted during the past 10 to
15 years.

Before discussing the radiosensitivities of particular cells and tissues, a
discussion of what is meant by radiosensitivity is in order. If atrophy of
tissues is referred to, it is obvious that this may be merely a consequence
of inhibition of growth of tissues, which are continually regenerating to
compensate for cells thrown off. This is particularly true of blood-form-
ing organs. Interference with other synthetic processes (e.g., secretory
activities) may be less obvious on cursory examination. In general,
radiosensitivity of cells, as discussed here, will refer to destruction or
degeneration of cells as living entities, and it may be well to recall that
these events are prone to occur at the time of cell division.

928 RADIATION BIOLOGY

For a given cell, the nucleus is a more sensitive indicator of damage than
the cytoplasm, and the cell in mitosis is usually more susceptible to
injury than the cell at rest. But even in mitosis there are differences,
for cells in prophase and metaphase are the most sensitive to radiation
injury (Sparrow, 1951). It is known that the absorption of radiation
depends upon the atomic constitution of the absorbing medium. While
nuclear sensitivity may perhaps be attributed to the differential absorp-
tion of radiation by the nucleus and cytoplasm, tissues with nearly the
same cross section can vary greatly in their susceptibility. It is evident,
therefore, that other intrinsic factors are involved. Although it has
been demonstrated by fractional irradiation that ions produced in the
cell nucleus are more effective biologically than those produced elsewhere
(Zirkle, 1932; Henshaw, 1938), nuclear injury in certain cells may be a
result of toxic factors originating in the cytoplasm (Duryee, 1949). A
normal cell injected with cytoplasm from an irradiated cell has been
shown to exhibit typical radiation effects. Thus, the special sensitivity
of the nucleus to radiation may, under certain conditions, be more
apparent than real, at least in terms of the initiating mechanism.

The sensitivity of the cell during division may reside in some facet
of its instability, since mitosis is characterized by a number of physico-
chemical changes, in, for example, chromosomal mass and surface,
viscosity, permeability, conductivity, and energy requirements. Poly-
merization and depolymerization of nucleic acids, as well as changes in
the relative amounts of nuclear and cytoplasmic nucleic acids, are thought
to occur during the mitotic cycle, and such changes may affect sensitivity.
Recent work with pepsin-albumin films reveals that form, even on the
molecular scale, can greatly influence the radiation response (Mazia and
Blumenthal, 1948). The physical and chemical factors that contribute
to nuclear sensitivity are numerous and only partially understood. They
are discussed in some detail in the excellent review by Sparrow (1951).

Recovery phenomena have been evoked to explain the difference in
response of slowly and rapidly dividing cells. It is believed that the
slowly dividing cell has a greater chance to recover since the death of a
cell frequently occurs at mitosis from structural alterations incurred
sometime before (Lasnitzki, 1943a). However, with a and neutron
irradiation, or with high dosages of X rays, degenerated cells appear in
appreciable numbers before resumption of the mitotic process (Lasnitzki,
1943b; Spear and Tansley, 1944; Tansley et al., 1948). Although there
is ample evidence that radiation interferes with the synthesis of desoxy-
ribonucleic acid (DNA), which could explain the postirradiation mitotic
inhibition, the disturbance in rapidly growing and in adult tissues does
not differ greatly (Mitchell, 1942; Hevesy, 1945, 1949; Holmes, 1949;
Kelly and Jones, 1950). Unfortunately, there are few, if any, data
relating the recovery of nucleic acid formation to the mitotic activity of

PHYSICAL AND BIOLOGICAL FACTORS 929

tissues known to differ in sensitivity. It has been suggested that radio-
sensitivity may be related to the ratio of the nuclear and cytoplasmic
nucleic acids, since the highest ratio is found in lymphoid tissue and the
lowest in resistant cells (Brues and Rietz, 1951). Proliferating tissues
probably show, in general, an increase in DNA per cell (Price and Laird,
1950). It is well known, however, that ribonucleic acid (RNA) also
tends to be increased in proliferating cells.

That certain types of recovery may be faster in cells which are pre-
sumed to have a more rapid turnover is indicated by the finding that
intestinal epithelium and lymphoid tissue exhibit less interference of
mitotic activity after sublethal X irradiation than the less sensitive skin
and adrenal tissue (Knowlton and Hempelmann, 1949). On the other
hand, although thyroid administration increases the mitotic index of
mouse epidermis, and presumably its rate of metabolism, it does not
affect the response to irradiation, similar changes being observed in the
epidermis of control and thyroid-fed irradiated mice (W. W. Smith, 1951).

In general, the hematopoietic and germinal tissues are the most sensi-
tive to radiation. These are followed by the intestinal epithelium, skin,
and connective tissue. Bone and glands are relatively radioresistant
while muscle and nerve are the least sensitive (Warren, 1942; Henshaw
and Snider, 1946; Warren and Bowers, 1950). There seems to be no
relation between the susceptibility of different tissues and their basal
oxygen consumption. Brain and kidney have higher rates of respiration
than spleen; yet the former are relatively radioresistant, while the latter
is radiosensitive. Although it has been shown that polyploidy protects
certain simple organisms against radiation damage (Latarjet and
Ephrussi, 1949; Clark and Kelly, 1950), there does not appear to be any
reason to think that this is an important factor in radiosensitivity of the
several animal tissues. Polyploid cells occur in the liver, but they are
the exception rather than the rule and probably do not account for the
apparent resistance of this organ to radiation.

Although radiosensitivity appears to be related to the life span, growth
rate, and differentiation of tissue, this is only part of the story. For
example, with the onset of mitosis germinating wheat seedlings become
more resistant to the growth-retarding effect of X radiation (Henshaw
and Francis, 1935), and in the eggs of Drosophila sensitivity does not
exactly parallel the rate of division (Packard, 1930). Furthermore,
radiosensitivity is not correlated with the number of premitotic or
mitotic nuclei in the roots of Vicia faba (Mottram, 1935b). The most
sensitive cells in the testis are the spermatogonia rather than the sperma-
tocytes, which manifest greater mitotic activity (Bloom, 1947). The
rapidly growing squamous cell epithelioma is fairly resistant to radiation,
while the more slowly growing basal-cell tumor is sensitive (Packard,
1930). There are other exceptions; the regenerating liver does not show

930 RADIATION BIOLOGY

evidence of increased sensitivity (Braes and Rietz, 1951), and erythro-
blast vulnerability to radiation injury is not enhanced by an increase in
mitotic activity; in fact, the hyperplastic erythroid tissue shows less
injury than the normal (Jacobson et at., 1948).

There are equally convincing arguments in regard to primitiveness and
radiosensitivity. Susceptibility of the developing ovum of the rabbit
does not depend exclusively on its differentiation (Bloom, 1947). More-
over, the primitive reticular cells are exceedingly resistant although the
blast cells, which can develop from reticular cells, are quite sensitive
(Bloom, 1947; Tullis, 1949). Finally, the highly differentiated nerve cell
is radioresistant while the polymorphonuclear leukocyte, though well
differentiated, is fairly sensitive (Warren, 1942). It would appear that
many factors act to influence responsiveness of tissue under various con-
ditions of growth and differentiation.

SPECIES SENSITIVITY

The lethal dose of ionizing radiation for the whole animal varies not
only among the different species but also among animals of the same
species. The dosage of total-body X radiation required to kill 50 per
cent of adult warm-blooded mammals within 30 days ranges from 200 to
800 r (Dowdy, 1949). In Table 14-1 are presented the acute lethal

Table 14-1. Comparison of the 30-day LD6o for a Total-body Dosage of Hard

X Rays"
Animal LD50, r

Guinea pig 200-400

Swine 275

Dog 325

Goat 350

Monkey 500

Mouse 400-600

Rat 600-700

Hamster 700

Rabbit 800

a These dosages are approximate and may vary from time to time in the same or
different laboratories (refer to text).

dosages for a number of laboratory animals. The absolute roentgen
values will differ somewhat depending upon the strain, conditions of
exposure, and maintenance, but the order of sensitivity remains the
same. The LD50 for man is questionable; best estimates place it between
the LD50 for the goat and mouse. Of interest is the radioresistance of the
bat, a hibernating mammal, whose life span under laboratory conditions
is shortened only after dosages of about 15,000 r (D. E. Smith et at.,
1951). On the other hand, the LD50 for the goldfish is only 850 r (Prosser
et al., 1947a).

PHYSICAL AND BIOLOGICAL FACTORS

931

Species sensitivity to penetrating radiation is not well correlated with
body size or with metabolic rate, although these factors, as will be dis-
cussed later, may be important in individual animals. There is little
difference in the basal heat production of the guinea pig and rat although
the LD5o for the guinea pig is lower than that for the rat by a factor of
about 2. Since a number of factors are involved in radiation death,
species sensitivity may reflect, in part at least, the particular suscepti-
bility of the different animals to the diverse mechanisms leading to
morbidity, e.g., toxins, leukopenia, bacteremia, hemorrhage, impaired
nutrition, and shock. There is a suggestion that species sensitivity may
be related to differences in the rates of recovery from radiation injury
since the mean survival time after median lethal irradiation is greater for
the more sensitive species (Sacher, unpublished observation, 1951). It
is of interest that a number of physiological and histological changes
reflect the amount of radiation and not the lethal effect; i.e., they are
more nearly independent of species (Bloom, 1947; Brues and Rietz,
1948; De Bruyn, 1948).

In contrast to the well-established differences in species sensitivity to
penetrating radiations are the apparently negligible differences in the
acute lethal effects of total-surface /3 irradiation when a correction is
made for body size, i.e., for the proportion of the body mass irradiated.
The median lethal dose of external 0 irradiation varies from 4700 rep for
the mouse to 17,000 rep for the rabbit; the total integrated dose, however,
is directly proportional to the body mass (Raper, 1947). That the
lethal action of surface 0 rays is dependent upon a total mass or volume
effect is assumed from the fact that all the energy is absorbed in a super-
ficial layer of tissue whose mass is small relative to the mass of the animal.
The possibility that some compensating situation, also varying with
body mass, may be active is not ruled out. The lethal mechanisms are
undoubtedly different for the superficial and penetrating types of radia-
tions (Table 14-2); for example, the blood picture is unaffected by a

Table 14-2. Dosage of External Beta (P32) and Gamma (Ta182) Radiation
Required to Kill 50 Per Cent of Animals within 45 Days

(After Raper, 1947)

Animal

0 radiation, rep

y radiation, r

Baby rat

Mouse

Rat

2,200
4,700
7,500
7,750
17,000

510

840
1280

Guinea pig

Rabbit

310

1500

lethal dose of external (3 irradiation. With internally deposited (3
emitters, or a emitters, however, the picture more or less resembles that

932 RADIATION BIOLOGY

following penetrating external irradiation (Bloom, 1947; Prosser et al,
1947b). The wider the distribution of an internal emitter, the greater is
the similarity of the clinical picture to that associated with penetrating
radiation.

INDIVIDUAL SENSITIVITY

Individuals in an apparently uniform population do not respond equally
to radiation. Curves relating mortality to dose are of the sigmoid type
and are quite steep, especially for mammals (Ellinger, 1945; Boche and
Bishop, 1946). Variations in mortality of from 0 to nearly 100 per cent
may occur in the LD50 range (Clark and Uncapher, 1949). As might be
anticipated, the dose range between just lethal and completely lethal is
greater in hybrid than in inbred strains, and, in addition, the LD5o varies
under different laboratory conditions. Besides the variation in mortality,
there are impressive differences in survival time during the acute period
following median lethal irradiation. In the rat, deaths within 30 days
are most frequent between the fourth and eighth and between the tenth
and fifteenth postirradiation days (Hagen and Simmons, 1947). Mor-
tality waves have also been observed in rabbits and chickens (Karnofsky
et al, 1950; Jacquez and Karnofsky, 1950; Steamer, 1951; Hagen and
Sacher, 1946). In these animals the first peak of deaths occurs during
the first day after irradiation, even with an LD50- The existence of
several processes that may lead to death in the acute period is suggested
by these findings. Reference has already been made to the apparent
transition from one mechanism of death to another with increasing
amounts of supralethal radiation. Although there are wide variations in
mortality, it is rather surprising that there are few histologic differences
between animals dying after an LD50 of radiation and those surviving the
same dose (Bloom, 1947).

Radiosensitivity of the embryo varies with its age (Karnofsky et al,
1950; Wilson, 1935; Wilson and Karr, 1950). The chick embryo mani-
fests an increasing sensitivity to early lethality (death within 24 hours)
during the third to seventh day of incubation, and is most sensitive on
the eighth to tenth day, after which its response is stabilized at a slightly
more resistant level (Karnofsky et al, 1950). Depression of growth is
also related directly to the age of the chick embryo at the time of irradia-
tion. Interestingly enough, the number of dividing cells in the non-
irradiated embryo decreases markedly between the third and eighth days
of incubation (O'Connor, 1950). Oxygen consumption is unchanged,
but there is a threefold decrease in aerobic glycolysis during this interval.
Irradiation of the pregnant mouse before implantation of the embryo
leads to a high prenatal fetal mortality with only a negligible incidence
of abnormality in animals surviving to term (Russell and Russell, 1950).
The earliest stages after mating are the most sensitive. When the preg-

PHYSICAL AND BIOLOGICAL FACTORS 933

nant mouse is exposed during the postimplantation stages, the relative
magnitude of effects is reversed, abnormality exceeding mortality. There
is a striking separation in sensitivity to lethal action. Exposure to 200 r
on the ninth day of gestation is completely lethal while 400 r is required
on the tenth day of gestation (Wilson and Karr, 1950). This may be
related to implantation of the embryo or to beginning differentiation.
Malformations of the skeleton and destruction of the developing nervous
system are prominent sequelae of irradiation during the latter two-thirds
of pregnancy (Russell, 1950; Hicks, 1950). In contrast, adult bone and
nervous tissue are relatively radioresistant (see Chap. 13).

There is remarkably little difference in the LD50 for delayed deaths
among the chick embryo, baby chick, and adult chicken, the embryo
being, in fact, somewhat more resistant, perhaps because of the sterility
of the internal environment of the egg (Karnofsky et al., 1950; Jacquez
and Karnofsky, 1950). This is apparently not the case in the mammal.
The mammalian fetus is more susceptible than either the young or adult
animal, and the former is, in general, more sensitive than the latter.
There are some exceptions, however. Mice under 15 days of age are less
sensitive and survive longer than 30-day-old animals, which manifest a
greatly increased susceptibility (Quastler, 1945b; Abrams, 1951; Furth
and Furth, 1936). Sensitivity to the lethal action of X rays decreases
rapidly with increasing age beyond 30 days, and little difference is appar-
ent between two- and three-month-old animals. The acute median
lethal dose of y rays does not differ appreciably for mice varying in age
from 1.5 to 12 months (Zirkle et al., 1946). The relative resistance of the
newborn mouse may be related to protective influences associated with
suckling (Abrams, 1951). Conversely, the sensitivity of 30-day-old
mice may be a function of changes associated with puberty. It is of
interest that maximal susceptibility to radiation lymphoma also occurs
during the puberal period in mice (Kaplan, 1948). In contrast to these
results, the amount of X radiation required for minimal depression of
femoral growth following local exposure of the epiphyseal region is linear
with age (Hinkel, 1942). Minimal stunting is seen with 700 r in one-
month-old rats, whereas 2200 r is required at six months of age. The
younger rats, however, recover more quickly than older animals.

Females may be somewhat more resistant than males, i.e., by about
50 r, but a sex difference is not apparent in all species or in different
strains of the same species (Hagen and Simmons, 1947; Hagen and
Sacher, 1946; Abrams, 1951; Zirkle et al, 1946). The role of body
weight in radiosensitivity is not well established, although it appears that
heavier animals tend to be less sensitive. It is not clear, however,
whether this is a reflection of weight or of age. The median lethal dose of
X rays is not very dependent upon weight in rats of approximately the
same age (Hagen and Simmons, 1947). However, rats of the same age;

934 RADIATION BIOLOGY

differing in weight because of dietary restrictions, vary in their sensitivity
to neutron irradiation, the smaller rats being more sensitive (Ely and
Ross, 1947). Rabbits of either sex weighing more than 2 kg are reported
to be somewhat less sensitive than animals weighing less than 2 kg (Hagen
and Sacher, 1946). The heavier rabbits survive longer in any given
dose group. It has also been observed that survival time is related to
body weight in mice (Quastler, 1945b). An explanation of the protective
influence of weight or age is not obvious ; it is probably not attributable to
a simple relation between the rate of growth and sensitivity. Differences
in the amount and distribution of body fat can account for small differ-
ences in sensitivity on the basis of the low effective atomic number of fat
(Spiers, 1946).

A number of constitutional factors must influence the breakdown
following irradiation. It is thought that preirradiation tests of func-
tional capacity would be useful in sorting out many of these influences,
thereby eliminating much of the so-called "inherent variability." Unfor-
tunately, there is little information relating to this aspect of the problem,
which is of both practical and theoretical importance.

INDUCED RADIO-SENSITIVITY OR RESISTANCE

Many procedures have been employed in an attempt to modify the
response to radiation. Some of these act in a general way to change
the response of the organism to stress. Others are concerned more
specifically with the decisive events responsible for injury to critical
physiological systems or for recovery from such injury. Some of the
procedures within the more specific category may be considered to act
primarily upon the biological system, while others probably influence the
pathways of energy dissipation within it.

Needless to say, the criterion will influence any evaluation of radio-
sensitivity factors. The death of an irradiated animal may be a con-
siderable distance downstream from death of any particular cell or group
of cells in the same animal. Since the death of an animal is more com-
plex than the death of a cell, it is not surprising that more factors are
found to influence sensitivity of the animal than of the cell. Likewise, it
is necessary to distinguish the modification of immediate and delayed,
or acute and chronic, effects. Protection against acute lethal action
does not necessarily imply protection against the more chronic sequelae
in so far as different mechanisms may be involved in their development.

In general, any agent that changes the body economy sufficiently modi-
fies the lethal action of radiation. Adrenal insufficiency (Cronkite and
Chapman, 1950), protein depletion (Jennings, 1949; Elson and Lamerton,
1949), vitamin deficiency (Johnson et al., 1946), infection (Taliaferro and
Taliaferro, 1951; Shechmeister and Bond, 1951), trauma (Brooks and

PHYSICAL AND BIOLOGICAL FACTORS 935

Evans, 1950), and exhaustive exercise (Stapleton and Curtis, 1946;
Kimeldorf et al, 1950) increase susceptibility to radiation as to most
noxious stimuli. Some degree of radioresistance may be acquired as a
result of previous exposures to /3 or X radiation (Raper, 1947; Cronkite
et al, 1950; Bloom, 1950), pretreatment with horse serum (Hektoen,
1918; Graham et al, 1950), estrogens (Treadwell et al, 1943; Patt,
Straube, et al, 1949), and adrenal cortical extract (Graham et al, 1950),
and by immunization (Kohn, 1949). There are no adequate explanations
of these phenomena. In the case of the estrogens it is believed that the
decreased toxicity is related to stimulation of myeloid activity following
the initial estrogen-produced depression since the pretreated animals
show rapid recovery from radiation granulocytopenia but not from
lymphopenia (Patt, Straube, et al, 1949). Along these lines, an increased
resistance of erythropoietic tissues has been observed following the pro-
duction of a regenerative anemia by phlebotomy, administration of phen-
ylhydrazine (Jacobson et al., 1948), or exposure to high altitude (Schack
and MacDuffee, 1949).

TEMPERATURE AND METABOLIC RATE

There is general agreement that the changes incidental to irradiation
are dependent upon the rate of metabolism. This is consistent with
observations made in other types of injury and is not unreasonable in
view of the apparent nonspecificity of radiobiological effects. An
increased metabolic activity after exposure to ionizing radiation induced
by thyroid extract or dinitrophenol (W. W. Smith and F. Smith, 1951;
Blount and Smith, 1949), a cold environment in nonacclimatized mammals
(W. W. Smith et al., 1949 ; Hempelmann et al., 1949), or exhaustive exercise
(Stapleton and Curtis, 1946; Kimeldorf et al, 1950) enhances lethality,
while depression of metabolic rate, at least in poikilotherms and isolated
tissues (Patt and Swift, 1948; Allen et al, 1950; Schrek, 1946; Cook,
1939), decreases the rate of development of radiation damage. Surpris-
ingly, administration of thiouracil and thyroidectomy appear to have
little influence on lethality in adult mice and rats (W. W. Smith and F.
Smith, 1951; Blount and Smith, 1949; Hempelmann et al, 1949; Haley
et al, 1950).

Evidence that relates to the effect of temperature during exposure
to radiation is equivocal, and virtually every conceivable effect has been
described. Sensitivity of the eggs of Ascaris (Holthusen, 1921) and of
Drosophila (Packard, 1930) is increased by elevation in temperature
during irradiation. Yet temperature is without influence on E. coli
(Lea, 1946), thymic cell suspensions (Schrek, 1946), wheat seedlings
(Henshaw and Francis, 1935), or eggs of the frog and hen (Ancel and
Vintemberger, 1927). Moreover, cold is reported to increase the sensi-
tivity of the roots of Vicia faba and of certain tumor cells to the growth-

93(5 RADIATION BIOLOGY

retarding effects of irradiation and also to increase the number of
dominant lethal mutations in irradiated Drosophila (Mottram, 1935a;
Crabtree and Cramer, 1933; Baker and Sgourakis, 1950). In larger
animals the results are equally confusing. Sensitivity of newborn mice
and rats to irradiation of the whole body or of the skin is decreased when
the ambient temperature is lowered during exposure (Lacassagne, 1942;
Hempelmann et al., 1949; Evans, Goodrich, and Slaughter, 1941; Evans,
Robbie, et al., 1941). Chilling of the adult animal does not increase the
survival rate, however (Hempelmann et al., 1949). Moreover, lethality
and cytological damage in the frog and tadpole are not influenced by a
change in body temperature during irradiation (Patt and Swift, 1948;
Aliens al., 1950).

Evaluation of temperature effects during irradiation is complicated
since temperature may alter the quality as well as the quantity of cellular
activity, which, in turn, may modify the response to irradiation. Fur-
ther, reactions of injury and of recovery must rapidly follow the primary
events that are associated with the absorption of energy, and it is probable
that these secondary metabolic reactions appear in some degree even
before the irradiation is terminated. Since the time course and tempera-
ture coefficients of the subsequent biochemical changes may vary for the
different effects and systems, it is perhaps not surprising that many types
of temperature responses have been described. When there is little
metabolic activity initially, as in the unfertilized eggs of Nereis, the
temperature coefficient of the events occurring during irradiation, which
result ultimately in a lethal action, is found to be about 1.1 (Redfield
et al, 1924).

Certain temperature effects may be attributed more directly to specific
changes, e.g., in blood flow or in oxygen tension. Sensitivity to radiation
is dependent upon the blood flow to the exposed area, and the beneficial
effects of chilling the skin can be ascribed to changes in the vascular bed
(Carty, 1930). Likewise, the decreased lethality of the chilled newborn
mammal may be a consequence of lowered oxygen tension resulting from
the relatively greater depression of breathing than of tissue respiration
in the cold. In contrast, the increase in dominant lethal mutations in
Drosophila irradiated in oxygen at 2°C over those treated in oxygen at
27°C has been related to the higher oxygen tension within the irradiated
sperm at the lower temperature (Baker and Sgourakis, 1950). Actually,
these two views are not incompatible.

While it is true that fern spores and certain pollens show an enhanced
radioresistance at liquid air temperatures, perhaps because of altered
formation and diffusion of free radicals or of some change in the con-
figuration of organic molecules, there is reasonable evidence that less
extreme temperature changes do not directly influence the immediate
radiation reactions in living systems. Temperature independence over

PHYSICAL AND BIOLOGICAL FACTORS 937

the narrow range compatible with life is consistent with the concept of
indirect as well as of direct action since the free radicals must have zero or
nearly zero energies of activation. On the other hand, the events sub-
sequent to irradiation appear to be rate-sensitive ; yet, since recovery may
also be influenced, the ultimate outcome is not always changed.

Consistent with these concepts are the findings that treatment of mice
with desiccated thyroid both before and after irradiation gives the same
enhanced effect as treatment begun after the exposure, while pretreatment
alone fails to modify lethality (W. W. Smith and F. Smith, 1951). It is
also noteworthy that survival is not altered by severe preirradiation
exercise (Patt, Blackford, et al., 1951) or by irradiating animals under
Nembutal anesthesia (Hempelmann et al., 1949; Patt, Blackford, et al.,
1951). Although lethality is diminished when mice are given a heavy
dose of morphine before exposure to X rays, protection is probably a
consequence of the hypoxia resulting from depression of the respiratory
center (Kahn, 1951). In contrast, the lethal effect is augmented when
rats are anesthetized with urethane, a radiomimetic agent, and then
irradiated (Henry, 1949). This has not been observed in mice, however
(Paterson and Matthews, 1951).

INTERMEDIARY RADIOCHEMICAL EVENTS

The aberrations in cell chemistry that follow irradiation are not easily
resolved since the pathways of energy dissipation and the structure and
properties of biologically important molecules are still obscure. From
energy considerations and the widespread nature of radiation damage
there is reason to believe that the primary disturbance centers around
enzymes, genes, and other key molecules that are involved in the assembly
and synthesis of essential substrates. Presumably, certain of these
entities are inactivated or modified by direct ionization and excitation
and/or by the products of irradiated water. It has been postulated by
Barron (1946) and Barron and Dickman (1949) that the critically
involved enzymes are those requiring sulfhydryl groups for their activity
and that these are reversibly oxidized by low dosages and irreversibly
denatured by higher dosages. While sulfhydryl enzymes may be
inhibited by ionizing radiations under certain conditions, evidence for
their selective inhibition in vivo is incomplete (Dinning et al., 1950;
Dubois et al., 1950; LeMay, 1951). It is not known, moreover,
whether changes when observed are the cause or the effect of cell
injury and death.

The abundance of water in biological materials and the demonstration
of activated water reactions in simple chemical systems have naturally
led to much speculation concerning their implication for radiobiology.
As stated by Weiss (1947), irradiated water is essentially an oxidation-
reduction system that consists primarily of free hydrogen atoms and

938 RADIATION BIOLOGY

hydroxyl radicals. Although free radicals result from the ionization of
water, they may also be formed in the ambient liquid when ionization
occurs directly in a biological particle. Depending upon the initial
spatial distribution of the ions and upon the presence and configuration
of dissolved substances, recombination or further reaction of the radicals
takes place. Alpha rays may produce peroxides (presumably from
hydroxyl radicals) even in pure water, whereas with y and X rays,
peroxides do not appear unless dissolved oxygen is present (Bonet-
Maury and Lefort, 1948; Allen, 1948). Obviously, purity conditions
such as these do not obtain in biological systems, where many acceptors
are available for reaction with the products of activated water. More-
over, the products of irradiated water may not be of equal consequence
inside and outside of a cell. It is well to recall, in addition, that the cell
represents a nonhomogeneous system with varying gradients of concen-
tration and solubility.

The activated-water concept of radiation action has received ample
confirmation in a variety of in vitro systems in which the dilution effect
described by Dale (1947) (independence of ionic yield and solute con-
centration except at very low and high concentrations) and the protection
effect (competition between solutes for the activated solvent molecules)
have been demonstrated. As the solute concentration increases or with
increase in radiation dosage, there is a transition from the activated-
water type of reaction to the direct-hit type characteristic of nonaqueous
systems. Demonstration of these phenomena in vivo is not an easy
task, however. The inability to detect an immediate oxidation of
sulfhydryl groups in tissues obtained from heavily irradiated animals is
consistent with theoretical considerations which reveal that only an
exceedingly small fraction of the available sulfhydryl reservoir could be
oxidized even in the absence of the naturally occurring protective sub-
stances (Patt, Straube, et al., 1950). Perhaps the most convincing evi-
dence, admittedly indirect, in support of the theory of in vivo effects of
activated water comes from studies with anaerobiosis and with protective
substances. It will be remembered that direct effects of radiation on
solutes also play a role, but one that cannot be well understood in the
present state of knowledge regarding exact physicochemical effects and
the nature of the biological targets.

Water. It was recognized long ago that desiccation of biological
systems favors radioresistance. Cells exposed to ionizing radiation are
frequently seen to swell and this may be a factor in, or manifestation of,
cell death (Failla, 1940; Buchsbaum and Zirkle, 1949). A hypothesis
has been advanced to account for this hydration phenomenon, i.e., ion-
ization and subsequent intra- and extracellular partition of ions, followed
by removal of the latter with consequent increase in cellular osmotic
pressure (Failla, 1940). However, there is little evidence in support of

PHYSICAL AND BIOLOGICAL FACTORS 939

an increase in intracellular water following lethal irradiation of the whole
animal (Soberman et al., 1951). While dehydration may decrease radio-
sensitivity by minimizing cell swelling, a more likely explanation seems
to be the interference with activated-water reactions either as a result of
a decrease in the available water or an increase in the concentration of
protective substances. The dependence of sensitivity on the degree of
hydration has been demonstrated in seeds (Petry, 1922), seedlings (Hen-
shaw and Francis, 1935; Wertz, 1940), isolated tissues (Chambers, 1941),
and tumor cells (Failla, 1940). Yet it has been difficult to determine
whether this relation holds in the intact animal. Mice deprived of water
for 24 to 40 hours before X irradiation tend to live somewhat longer, but
the proportion surviving is not altered materially (France, 1946). Frogs
kept in dry, individual containers for 3 days lose about 35 per cent of their
water, yet are no more resistant to irradiation (Patt and Tyree, unpub-
lished observations, 1949). Even though appreciable water remains,
some quantitative difference at this level of dehydration should be
anticipated. Perhaps the answer lies in the relative concentrations and
distributions of free and bound water under these conditions of water
deprivation.

Hydrogen Ion Concentration. The local changes in pH that probably
occur in the neighborhood of an ion track may play a role in the develop-
ment of radiation injury. The formation of hydrogen peroxide has been
shown to depend upon the hydrogen ion concentration, and changes in
pH have been observed in irradiated solutions (Frilley, 1947). Varia-
tion in pH during irradiation alters the radiosensitivity of germinating
fern spores, Drosophila eggs, and paramecia, sensitivity maxima being
observed at definite concentrations of acid or base (Zirkle, 1936, 1940,
1941). Zirkle has compared this behavior with the effect of irradiation
on proteins in vitro, where maximal flocculation is observed at a pH near
the isoelectric point. This is considered to be consistent with the
hypothesis that total sensitivity is due to the added effects of several
reactions having maximum yields at different acidities. There is little
information in connection with pH effects in isolated mammalian tissue
or in the whole mammalian organism. Change in the pH of thymic cell
suspensions from 7.8 to 6.6 does not influence the action of X rays
(Schrek, 1946).

Oxygen Effects. The effect of ischemia on cutaneous radiosensitivity
was observed by Schwarz in 1909. The relative resistance of Ascaris
eggs during anaerobiosis was described subsequently by Holthusen (1921),
who attributed the protection to the absence of cell division. Several
years later Mottram (1924) and Jolly (1924) investigated the effect of
blood flow on the response of skin and lymphatic tissue to X rays, and in
1933 Crabtree and Cramer presented a detailed account of the influence
of anoxia and related chemical factors on the radiosensitivity of tumor

940 RADIATION BIOLOGY

cells. The possible relation of this aspect of radiosensitivity to the
reactions of activated water was suggested by Thoday and Read (1947,
1949) and Read (1950). They confirmed the observation of Mottram
(1935a) that oxygen deprivation decreases the growth reduction of broad
bean roots following X irradiation and observed, in addition, that
oxygen lack did not afford significant protection following a irradiation.
This is a biological parallel to the radiochemical reactions involving
oxygen in aqueous solutions, since oxygen lack would be expected to
exert a minor influence on the formation of the hydroperoxyl radical
and hydrogen peroxide by a rays.

Deprivation of oxygen during exposure to y and X radiations dimin-
ishes their action in a variety of biological systems. This is true for
yeast (Anderson and Turkowitz, 1941), E. coli (Hollaender et al., 1951),
barley seeds (Hayden and Smith, 1949), Vicia faba (Mottram, 1935a;
Thoday and Read, 1947), Drosophila (Baker and Sgourakis, 1950),
Tradescantia (Giles and Riley, 1950), tumor cells (Crabtree and Cramer,
1933), and mice and rats (Lacassagne, 1942; Dowdy et al., 1950). Radio-
resistance is said to be the same whether anaerobiosis is induced by
nitrogen, helium, argon, or hydrogen. Reduced oxygen tension has been
shown to decrease a number of radiation effects, including lethality
(Dowdy et al., 1950), growth reduction (Thoday and Read, 1947),
chromosome aberrations (Giles and Riley, 1950), and sex-linked lethal
mutations (Baker and Sgourakis, 1950). However, under conditions in
which the radiation effect is considered to arise from the direct ionization
of a biological particle, the oxygen level is without influence (Hewitt and
Read, 1950).

It is well known that radiation injury to specific sites parallels the
blood flow during exposure (Schwarz, 1909; Carty, 1930; Mottram, 1924;
Jolly, 1924; Evans et al., 1942). Thus, damage to a limb is greatly
diminished when the limb circulation is blocked, and tumor sensitivity
varies with its vascularity. Mechanical retardation of breathing also
increases resistance to irradiation (Evans et al., 1942). Since a factor
common to all these examples is a reduced oxygen tension, it is assumed
that anoxia accounts for the modification of sensitivity. While this
seems reasonable from the preponderant evidence in support of anaerobio-
sis cited previously, it has not been proved.

The question naturally arises as to whether anoxia modifies radiation
injury by interfering with the radiochemical reactions that involve free
oxygen or by inducing a more specific biological effect, e.g., on enzymes,
metabolism, or cell division. The precise mechanism must be regarded
as unsettled ; there is reason to think, however, that both modes of action
may be involved in the oxygen effect. Although Dowdy et al. (1950) have
shown that the 30-day LD5o for rats X-irradiated in 5 per cent oxygen is
about twice that for rats exposed in air, it is not known whether breathing

PHYSICAL AND BIOLOGICAL FACTORS 941

a 5 per cent oxygen mixture for several minutes before and during irradia-
tion lowers the blood and tissue oxygen tension sufficiently to modify
free radical and peroxide formation to an appreciable degree. Mice are
also protected against lethal irradiation by anoxic anoxia (Dowdy et al.,
1950). However, the range in which protection is evident is quite
narrow, protection being observed with 7 per cent but not with 10 per
cent oxygen, while a 5 per cent level is lethal during the period of irradia-
tion. As contrasted with these observations, the increase in chromosome
aberration frequency in Tradescantia is linear between 0 and 10 per cent
oxygen, after which the rise is somewhat more gradual, tending to level
off above 20 per cent (Giles and Beatty, 1950).

Several chemical procedures, which bear directly on the events associ-
ated with the oxygen effect, have been employed. Histotoxic anoxia pro-
duced by cyanide apparently increases the sensitivity of tumor tissue and
of Vicia faba to radiation (Crabtree and Cramer, 1933). On the other
hand, cyanide exerts some protective action against X rays in the mouse
(Bacq et al., 1950; Bacq, 1951), while it is without influence in the rat and
frog (Patt and Swift, 1948; Dowdy et al., 1950). Iodoacetic acid and
sodium fluoride which, like cyanide and oxygen deficiency, inhibit aerobic
glycolysis, do not affect the sensitivity of tumor cells (Crabtree and
Cramer, 1933). The failure of cyanide-induced anoxia to protect the
rat and the protection afforded this animal by anoxic anoxia have been
taken to indicate that radiation injury is related to the tissue oxygen
tension (Dowdy et al., 1950). On the other hand, it has been suggested
that cyanide may protect the mouse by inhibiting the formation of
peroxides by X rays and perhaps by forming a loose bond with certain
enzymes (cytochrome reductase and catalase) to prevent their oxidation
(Bacq et al., 1950; Bacq, 1951). The important species difference
between the mouse and the rat in regard to cyanide as well as its potentia-
tion of toxicity in plant and tumor tissue cannot be taken lightly. Poten-
tiation of tissue sensitivity by cyanide may be due to an increase in
oxygen tension as a consequence of inhibition of the cytochrome system
or to inactivation of catalase. Also unexplained is the finding that para-
aminopropiophenone enhances the resistance of mice and rats when it is
administered before X irradiation, while sodium nitrite, which produces a
rather similar methemoglobinemia, gives equivocal protection (Storer
and Coon, 1950; Herve et al, 1950).

Chemical Protection. That oxidative reactions are important com-
ponents of radiation action is also suggested from studies with protective
substances. Thus, the in vitro oxidation of sulfhydryl enzymes following
low doses of radiation can be prevented or reversed by glutathione
(Barron et al., 1949), and radiation effects in tetanus toxin (Ephrati,
1948) and bacteria (Hollaender et al., 1951; Forssberg, 1950) can be
diminished by a variety of reducing substances. A number of oxidizable

042 RADIATION BIOLOGY

entities also have the capacity to dimmish many of the effects of X rays
in mammals. Cysteine (Patt, Tyree, et ah, 1949; Patt, Smith, et ah,
1950; Smith, Patt, Tyree, and Straube, 1950; Patt, Smith, and Jackson,
1950), glutathione (Patt, Smith, Tyree, and Straube, 1950; Chapman
and Cronkite, 1950; Chapman et ah, 1950), and BAL (Smith, Patt, and
Tyree, 1950) have been shown to reduce acute toxicity in mice, rats, and
dogs. Some degree of protection has been seen also with thiourea
(Limperos and Mosher, 1950; Mole et ah, 1950), dithiophosphonate
(Mole et ah, 1950), and massive amounts of glucose (Loiseleur and Velley,
1950a; Baclesse and Loiseleur, 1947) and ethanol (Paterson and
Matthews, 1951; Patt, Mayer, and Smith, 1951a). The action of these
protective substances against repeated low-dose irradiation has not, to
our knowledge, been evaluated. Protection against the chronic sequelae
of irradiation is also largely undetermined.

It is perhaps significant that cystine, methionine, and ascorbic acid do
not modify sensitivity in the mammal (Patt, Smith, Tyree, and Straube,
1950). While all reducing agents do not appear to protect against the
acute lethal action of ionizing radiations, e.g., ascorbic acid, hydrosulfite,
tetraborohydride, mercaptosuccinate, and thiosulfate, this may be a
consequence of their temporal and spatial distribution and biological life
(Patt, Blackford, et ah, 1951; Patt, Mayer, and Smith, 1951a).

The time course of the protection against the acute effects of radiation
may follow a somewhat different pattern in the intact animal and in
certain in vitro systems. In general, however, these substances must be
given before irradiation to be effective. Thus, the optimal time of
cysteine administration in rats and mice is immediately before exposure,
and there is no postirradiation effect (Patt, Smith, Tyree, and Straube,
1950). Although some improvement of survival has been observed in a
small series of rabbits after treatment with cysteine plus ascorbic acid
during the first two postexposure hours (Loiseleur and Velley, 1950b),
this has not been verified in either rabbits or mice (Patt, Mayer, and
Smith, 1951b). On the other hand, the survival of thymocytes irradi-
ated in vitro is increased to the same degree whether cysteine is added
one minute before or after the exposure (Patt, Blackford, and Straube,
1952). Cysteine addition 15 to 30 minutes before X irradiation is
optimal, however. There is no satisfactory explanation for the apparent
discrepancy other than to implicate possible differences in the kinetics of
the reactions with cysteine and in the time constants for the develop-
ment of irreversible injury in the thymic cell suspension and in the intact
animal.

It has been suggested that glutathione protects animals by promoting
regenerative mechanisms and not by preventing cellular destruction
(Cronkite et ah, 1951). The basis for this interesting concept rests in
the similarity of changes in organ weights, peripheral blood counts, and

PHYSICAL AND BIOLOGICAL FACTORS 943

histologic appearance of certain radiosensitive tissues in treated and
untreated mice during the first few days after exposure to a nearly com-
pletely lethal dose. These findings do not, however, constitute incon-
trovertible proof of failure to prevent injury. There is reason to believe
that histologic changes, in general, reflect the amount of radiation and
not the lethal effect on the animal and, moreover, that the threshold point
of maximal change may be below that required for acute lethality (Bloom,
1947; De Bruyn, 1948). These considerations can also explain the failure
to observe a protective effect of glutathione on organ weights (Cronkite
et at., 1951) and of glutathione and cysteine on the uptake of iron in the
red blood cells of irradiated animals (Hennessy and Huff, 1950; Hennessy
and Folsom, 1950; Hennessy, Folsom, and Glover, 1950). For example,
the anticipated difference in the organ weights between 600 and 800 r,
which represents the degree of protection by glutathione on the basis of
survival data (Chapman, Sipe, et al., 1950), is only about 5 per cent
(Carter et al., 1950). The failure to detect a difference in the heterophil
and erythrocyte levels in the peripheral blood of glutathione-treated and
untreated mice is not readily explained, however (Cronkite et al., 1951).

As contrasted with the view that protection in the mammal results
from some facilitation of recovery rather than prevention of injury, there
is a body of evidence indicating that sulfhydryl compounds can diminish
cellular destruction incident to X-ray exposure. Cysteine has been
shown to decrease damage to the skin (Forssberg, 1950) and lens (von
Sallmann et al., 1951) after local irradiation and to increase the resistance
of tumor cells (Straube et al., 1950; Hall, 1951). Thymic cells have also
been protected in vitro over a dosage range of 50 to 2000 r (Patt, Black-
ford, and Straube, 1952). Furthermore, there are indications that
cysteine can modify significantly the hematologic changes in irradiated
rats (Patt, Smith, and Jackson, 1950; Rosenthal et al, 1950). Indeed,
the changes that have been observed in rats irradiated with 800 r after
cysteine treatment compare favorably with those observed after a sub-
lethal exposure of 300 r (Patt, Smith, and Jackson, 1950). These con-
siderations suggest that cysteine may raise the threshold for radiation
effects in general, with the probable exception of those attributable to
direct ionization.

The mechanism of chemical protection is poorly understood. Protec-
tive substances may increase the resistance to X rays by modifying
activated water reactions, the nature of the biological targets, or the
redox equilibrium of the cell. It is of interest that a given amount of
cysteine accounts for a rather constant percentage of the biological
effect of the radiation over a wide dose range (Patt, Blackford, and
Straube, 1952; Patt, Mayer, Straube, and Jackson, in press). This has
been observed in survival studies with thymic cells and with mice, but,
unfortunately, can be explained equally well by postulating an interaction

944 RADIATION BIOLOGY

with either the radiation or its toxic intermediates or with the biological
system.

It is not known whether reducing substances and oxygen deprivation
protect in the same manner. There is, of course, no compelling reason
to believe that the mode of action of either factor is identical in all living
systems or indeed in a specific system under different conditions. X-ray
inactivation of ribonuclease is prevented by glutathione but not by oxygen
lack (Holmes, 1950). It is perhaps significant that resistance of the rat
to the acute lethal action of X rays is increased to the same degree by
anoxic anoxia or cysteine (Dowdy et al., 1950; Smith, Patt, Tyree, and
Straube, 1950) and that the effect of cysteine is enhanced in mice breath-
ing 10 per cent oxygen during the irradiation, although the latter by itself
does not offer any protection (Mayer and Patt, 1953). Certain additive
effects have been observed in bacteria. The sensitivity of aerobically
grown E. coli irradiated in phosphate buffer is reduced beyond the value
obtained with anoxia alone when cysteine is present during the exposure
(Hollaender et al., 1952). This is not the case, however, when the organ-
isms have been grown anaerobically. Thymic cells resemble the anaer-
obic bacteria in the lack of additivity (Patt, Blackford, and Straube,
1952). These considerations suggest perhaps that some of the protec-
tive substances may act by diminishing the availability of oxygen in the
biological system. This does not imply, however, that oxygen depriva-
tion per se is necessarily the decisive event in the protection afforded by
such agents. Information relating to the relative effectiveness of oxygen
and protective substances for the different qualities of radiation, to their
additivity or lack of additivity for a specific radiation quality, and their
temperature dependence should aid materially in the elucidation of the
mechanism of action. A survey of some of the more important chemical
and metabolic factors that have been employed in studies of animal radio-
sensitivity is presented in Table 14-3. Although it is not yet possible
to classify these factors in terms either of their biochemical action or of
radiation action, their significance is obvious since they provide a real
basis for the study of the immediate chemical effects of ionization and
excitation in living systems. The more recent contributions in this field
have been reviewed by Patt (1953).

SUMMARY

The physical and biological factors affecting sensitivity to ionizing radi-
ations have been discussed with emphasis directed toward the response
of mammalian tissue. It has been shown that the biological effect may
be influenced by the quality, quantity, manner, and completeness of
irradiation. Although it is generally true that radiosensitivity is related
to growth rate, it is clear that the relation is not a simple one. In view

PHYSICAL AND BIOLOGICAL FACTORS

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PHYSICAL AND BIOLOGICAL FACTORS 947

of the complexity of cell physiology and the ubiquitous nature of the
energy absorption, it is not surprising that the effects of radiation are
manifested in many ways and that many factors have the capacity of
influencing the radiation responses.

The events that are immediately associated with the absorption of
radiation are fairly independent of temperature over a wide range. How-
ever, the subsequent reactions of injury and recovery are subject to tem-
perature influence and metabolic activity. The existence of activated-
water reactions in vivo as well as in vitro is suggested by the rather
parallel influence of hypoxia and protective substances on the behavior of
aqueous solutions and living systems to radiation. The mode of action
of these modifying factors has not been settled, however. There is
reason to think that the chemical effects of ionization and excitation are
responsible for the early cytological damage following moderate dosages
of radiation and that this, in turn, bears a causal relation to the over-all
injury in the mammalian organism.

REFERENCES

(Information regarding availability of government reports indicated by an asterisk
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950 RADIATION BIOLOGY

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PHYSICAL AND BIOLOGICAL FACTORS 951

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952 RADIATION BIOLOGY

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954 RADIATION BIOLOGY

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PHYSICAL AND BIOLOGICAL FACTORS 955

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956 RADIATION BIOLOGY

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PHYSICAL AND BIOLOGICAL FACTORS 957

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958 RADIATION BIOLOGY

(1943) Radiobiological importance of specific ionization. USAEC Report

MDDC-444.*

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Manuscript received by the editor Apr. 17, 1952

CHAPTER 15

The Pathological Physiology of Radiation Injury
in the Mammal. II. Specific Aspects of the Physiology

of Radiation Injury

Harvey M. Patt and Austin M. Brues

Division of Biological and Medical Research, Argonne National Laboratory,

Lemont, III.

Introduction. Blood and hema,topoiesis. Abnormal bleeding. Body fluids: Blood
and plasma volume — Lymph and tissue fluid — Constituents of plasma and lymph.
Cardiovascular system. Gastrointestinal tract. Liver. Kidney. Endocrines: Adrenal
— Thyroid — Pituitary. Nervous System. Miscellaneous tissues and organs. Metabo-
lism and tissue breakdown. Immunity and injection. Nonspecific physiological
stresses. Acute radiation syndrome. References.

INTRODUCTION

The physical and biological factors that influence the response to high
energy radiation have been considered in Chap. 14. We are concerned
here with the physiological aspects of irradiation of specific sites or of
the whole organism. In general, the development and course of injury
are similar whether ionization and excitation result from penetrating
external radiations or from internally deposited radioactive materials.
While biological effectiveness differs for the several radiation qualities,
it is difficult to discern by physiological or histological means whether
irradiation has been accomplished by slow or fast neutrons, X or 7 rays
when a median lethal dosage is employed. External /3 irradiation is dis-
tinguished by the superficial nature of the damage, and the results of
poisoning with internal emitters are modified by the organ distribution
and trajectory of the radiating particles.

It is clear that no single change is peculiarly specific for radiation
injury. Since the ionizing energy is dissipated in a heterogeneous and
highly integrated system, many different effects may ensue, and not all
of these are a direct consequence of exposure. Thus, we must contend
not only with the physicochemical complexities of energy absorption, but
with the many ramifications of function in the organism seeking to retain
physiological balance. Although the early biochemical and cytological
effects probably bear a causal relation to the gross pathological physi-

959

960 RADIATION BIOLOGY

ology, the nature of this relation is poorly understood. The manifesta-
tions of the radiation syndrome and possible modes of their development
will be the subject of the following discussion.

BLOOD AND HEMATOPOIESIS

The radiosensitivity of blood-forming tissues and the consequent
hazard of blood damage have attracted considerable attention since the
classic work of Heineke in 1903. The vast early literature on this sub-
ject has been reviewed by Dunlap (1942). Although blood and blood
formation are discussed in some detail by Jacobson in Chap. 16, present
concepts merit brief consideration here since the hematologic picture is
an integral part of the radiation syndrome.

Peripheral blood changes depend, for the most part, upon the radio-
sensitivity of the parent or precursor cells, their ability to recover from
injury, and the life span or rate of utilization of the mature elements in
the blood. From morphological studies on acutely irradiated animals,
it may be seen that changes in the peripheral blood cells reflect to a con-
siderable extent alterations in the bone marrow, lymph nodes, and
spleen (Lawrence, Dowdy, and Valentine, 1948; Lawrence, Valentine,
and Dowdy, 1948; Bloom and Jacobson, 1948; Jacobson, Marks, and
Lorenz, 1949; Suter, 1947; Steamer et at., 1947a). This is not, however,
readily apparent after chronic irradiation (Jacobson, Marks, and Lorenz,
1949). While the site of action appears to be mainly in the blood-form-
ing organs, indirect or remote effects on hematopoietic tissue and direct
effects on the morphological constituents of peripheral blood are not
unknown. The alteration of lymphoid tissue in areas distant from the
site of irradiation is an example of the indirect type of action (Barnes and
Furth, 1943; Leblond and Segal, 1942). The utilization, destruction,
and production of blood cells following irradiation of the whole animal are
undoubtedly influenced indirectly by certain general derangements of body
economy. Thus, infection may lead to an increased demand, damaged
epithelial surfaces to an increased loss, nutritional disturbances to a
decreased formation of substances necessary for normal hematopoiesis,
and circulatory changes to a decreased effective blood cell mass.

Blood exposed in vitro to moderate dosages of radiation, i.e., in the
lethal range for mammals, shows only slight changes. A direct cytocidal
effect on lymphocytes from the thymus and spleen has been observed in
the test tube with as little as 50 r and may contribute to the early decrease
in circulating lymphocytes (Schrek, 1946). Mature granulocytes, on
the other hand, are quite resistant and are not affected directly by radi-
ation except in excessive dosage. Degenerate cells are not found gener-
ally in smears of mammalian blood made during the first few days after
irradiation ; they may be noted, however, in smears prepared from chicken

PHYSIOLOGY OF RADIATION INJURY 961

blood (Murray et al., 1948). Hemolysis has been seen in blood sub-
jected to heavy X irradiation (Ting and Zirkle, 1940; Liechti and Wil-
brandt, 1941; Frankenthal and Back, 1944; Halberstaedter and Leibo-
vvitz, 1947). Erythrocytes irradiated in vitro with polonium a particles
first shrink and then swell before hemolysis; these changes require large
dosages (5 X 105 rep) (Buchsbaum and Zirkle, 1949). Osmotic and
mechanical fragility of canine erythrocytes is apparently unaltered fol-
lowing exposure in vivo to X rays or neutrons (Davis et al., 1949; Ross
and Ely, 1947), but the thermal fragility of rat erythrocytes is reported
to increase during the first few hours after exposure of the whole animal
to 500 to 600 r (Goldschmidt et al., 1951). It is not known whether this
represents a direct effect of X rays on the red cell.

That radiation anemia may be due, in part, to a hemolytic reaction is
suggested by the rapid decline of red blood cells, which is in excess of that
resulting solely from the absence of erythrogenesis, the active phago-
cytosis of erythrocytes, and the increased excretion of bile pigments
(Davis et al., 1949; Schwartz et al., 1947; Prosser, Painter, Lisco, et al.,
1947). These facts do not in any sense constitute definite proof of a
direct action of radiation on the mature erythrocyte. The early anemia
may be partially explained by the loss of red cells from the circulation, by
the increase in plasma volume, and, to some extent, by the failure of
replacement. Large numbers of erythrocytes have been shown to appear
in the lymph within several days after irradiation, and extensive erythro-
phagocytosis and hemosiderosis have been seen in the lymph sinuses of
irradiated animals (Furth, Andrews, et al., 1951). Destruction of
erythrocytes may also be accomplished by the toxic materials that are
liberated from damaged tissues or from infectious organisms. While
increased excretion of bile pigments is suggestive of red cell destruction,
this may also be attributed to cessation of red cell production or to some
aberration in the incorporation of hemoglobin into young erythroid cells.

Although each of the ionizing radiations affects the hematopoietic
organs in the same qualitative manner, their relative effectiveness varies
(Lawrence and Lawrence, 1936; Jacobson and Marks, 1946). The
X-ray-to-neutron ratio for the acute lymphocyte and granulocyte
response in rabbits is around 6.0 (Jacobson and Marks, 1946). Only
minimal blood changes are seen after external irradiation with /3 rays
(Raper and Barnes, 1951a). This is consistent with the localization of
radiation effects. Granulocytes may increase several weeks after external
/3-ray exposure of rabbits, but it is noteworthy that granulocytosis
coincides with infection of the ulcerated skin. On the other hand, exten-
sive damage to the hematopoietic tissues and concomitant changes in
peripheral blood have been observed following poisoning with internal
emitters (Bloom and Jacobson, 1948; Jacobson, Marks, and Lorenz,
1949). Differences in the hematologic effects seen after administration

962 RADIATION BIOLOGY

of various radioelements may be attributed to their anatomical distribu-
tion and the specific ionization and energy of the radiating particles.

The histological picture is characterized by disappearance of mitotic
figures, cell degeneration, and death, leading, a few days later, to aplasia
of the marrow and lymphoid tissue (Bloom and Jacobson, 1948). Edema
and hemorrhage with subsequent fatty infiltration of the hypoplastic
marrow cavity may occur several days after irradiation (Lutwak-Mann
and Gunz, 1949). The early chemical changes in the blood-forming
tissues are obscure. A transitory increase in oxygen consumption,
carbon dioxide production, and hemin synthesis has been seen in marrow
homogenates prepared from rabbits immediately after 800 r of X irradia-
tion (Altman et al., 1951). Synthesis of RNA and DNA by rabbit bone
marrow is inhibited, however, while that of protein is unaffected, during
the five hours following similar irradiation (Abrams, 1951). An early
increase in respiration has not been observed in irradiated rat thymus
(Barron, 1946). Respiration and anaerobic glycolysis of marrow and
thymus decline below the preirradiation level during the first few days
after exposure; recovery is apparent around the tenth day (Lutwak-Mann
and Gunz, 1949; Barron, 1946). Nucleic acid phosphorus in the marrow
reaches a minimum value between the second and fourth days and is still
below normal at two weeks (Lutwak-Mann and Gunz, 1949). It is of
interest that fatty acid synthesis is increased soon after X irradiation;
this may account, in part, for the deposition of fat in the hypoplastic
marrow cavity (Altman et al., 1951).

Greatest sensitivity in the marrow is shown by the erythrocytic pre-
cursors (M. A. Bloom, 1948; Rosenthal, Pickering, and Goldschmidt,
1950; Denstad, 1941). Precursors of granulocytes and platelets are
somewhat less sensitive, and the primitive reticular cells are quite
resistant. Erythropoietic sensitivity is reflected physiologically by an
early decrease in the uptake of iron by bone marrow. Twenty-four
hours after exposure to 250 r, iron uptake in the rat may be decreased by a
factor of 10 (Hennessy and Huff, 1950). In contrast to these findings,
recovery from a standard anemia produced by phlebotomy immediately
prior to irradiation with 200 r is only slightly less rapid in irradiated than
in control cats (Valentine and Pearce, 1951). On the basis of functional
impairment, it also appears that erythropoietic tissue is less sensitive to
radiation injury than myelopoietic, but this may be a consequence of the
preferential influence of phlebotomy on the relative recovery rates of the
two tissues. Since cells of the reticulum appear to be the common
progenitor, recovery may depend upon the factors involved in their transi-
tion to the various cell types. Formation of new cells from the reticular
cells is said to be inhibited completely during the first week following
median lethal irradiation (Rosenthal, Pickering, and Goldschmidt, 1950).

The blood-forming organs become actively hematopoietic within one to

PHYSIOLOGY OF RADIATION INJURY 963

two weeks, but several months may elapse before regeneration is com-
plete (Bloom and Jacobson, 1948). Overcompensation or secondary
aplasia, sometimes seen during the recovery phase, can give rise to blood
dyscrasias, which have been encountered especially after repeated irradia-
tion. It is well known that a minimal exposure becomes important if
repeated frequently enough to result in a significant accumulated dose.
Definite hematological changes may appear in guinea pigs after exposure
to as little as 1.1 r per day (Lorenz et al., 1946). On the other hand, there
is no evidence of any permanent functional impairment of the hemato-
poietic tissues when cats are subjected to a whole-body irradiation with
200 r every four months over a period of a year and a half (Valentine,
Pearce, and Lawrence, 1951).

From the point of view of the time course of events in peripheral blood,
the most important consideration appears to be the rate of utilization or
life span of the various cellular elements. This increases progressively
from the lymphocyte to the granulocyte, reticulocyte, thrombocyte, and
erythrocyte, and represents the order in which changes occur in the
peripheral blood after exposure to penetrating radiation in single or
divided doses (Lawrence, Dowdy, and Valentine, 1948; Jacobson, Marks,
and Lorenz, 1949; Suter, 1947). Significant reduction in the number of
lymphocytes has been detected after a single total-body exposure to 25 r.
Regeneration, however, does not follow the same pattern since recovery is
ordinarily seen first in the reticulocytes, granulocytes, and thrombocytes,
then in the erythrocytes and lymphocytes.

Recovery of hematopoietic tissue is more rapid after subtotal irradia-
tion (Boffil and Miletzky, 1946; Jacobson, Simmons, Marks, Gaston, et al.,
1951; Rekers, 1949). The observations of Jacobson and his colleagues
(Jacobson, Marks, Robson, et al., 1949; Jacobson, Simmons, et al., 1950;
Jacobson, Simmons, Marks, Gaston, et al., 1951) are of particular sig-
nificance in this connection. They have shown that shielding the spleen
in mice and rats or the appendix in rabbits lessens the severity of the
blood changes and enhances recovery. Splenectomy performed prior to
irradiation does not modify the hematologic response to X rays (Steamer
et al., 1947b), although it does increase the severity of the anemia that
occurs after poisoning with Sr89 (Jacobson, Simmons, and Block, 1949).
Anemia fails to develop when Sr89 is given to the normal mouse, appar-
ently because of the intense ectopic erythropoiesis in the spleen. The
mechanism of these protective effects is not understood; it has been
suggested (Jacobson, Robson, and Marks, 1950) that mesenchymal
tissues in shielded areas may supply a factor that facilitates regeneration
of blood-forming tissue. It is of interest that spleen transplants (Jacob-
son, Simmons, Marks, and Eldredge, 1951; Jacobson, Simmons, Marks,
Gaston, et al., 1951) and spleen homogenates (Cole et al., 1952) facilitate
recovery and reduce radiation mortality in mice. Injection of lympho-

964 RADIATION BIOLOGY

cytes does not protect rats against radiation damage (Campbell and
Ross, 1952). Although transplants of bone marrow do not influence
recovery appreciably in dogs (Rekers, 1949; Rekers et al., 1950) and rats
(Talbot and Pinson, 1951), intravenous or intraperitoneal administration
to certain species is highly effective and, in fact, heterologous marrow
(guinea pig to mouse) has considerable effectiveness (Lorenz et al., 1951,
1952). The latter does not necessarily implicate a humoral substance
since it has been shown that normal tissue can survive and proliferate in
an X-irradiated heterologous host (Toolan, 1951).

The transient granulocytosis that occurs in the rabbit, chicken, and
rat during the first hours after X irradiation has been attributed to the
mobilization of granulocytes from storage depots as part of the early
reaction to injury rather than to a primary stimulation of blood formation
(Bloom and Jacobson, 1948). Accelerated maturation of myeloid cells
has been noted, however, in the bone marrow of rats during the first
hours after total-body irradiation (Rosenthal, Pickering, and Gold-
schmidt, 1950). The heterophils or granulocytes are the only circulating
cells that are initially increased in number; monocytes and eosinophils,
as well as lymphocytes, platelets, and reticulocytes, are invariably
reduced. An "abortive rise" of heterophils, lymphocytes, and reticulo-
cytes followed by a decline has been observed about one week after acute
exposure to X rays or neutrons (Jacobson and Marks, 1946; Jacobson
et al, 1947; Jacobson, Marks, and Lorenz, 1949). There is no satis-
factory explanation of this phenomenon.

Hemoglobin concentration and hematocrit, in general, parallel the
changes in erythrocyte count. The anemia is, therefore, usually normo-
cytic and normochromic (Jacobson, Marks, and Lorenz, 1949). A macro-
cytic anemia has been described in the rat after median lethal exposure to
X radiation (Steamer et al., 1947a), and similar anemias have been seen
in man after poisoning with radioactive materials (Martland, 1926).
While radiation-induced anemia is due primarily to the cessation of
erythrocyte formation, extravasation of red cells and secondary damage
are also substantial factors in its pathogenesis. The role of anemia in
acute death occurring within two to three weeks after median lethal
irradiation is not clear. The degree of the anemia indicates that it is at
most of contributory significance during this period. Anemia may
become a critical factor in the deaths seen at later periods, sometimes
after a single exposure but more often after repeated irradiation.

The sedimentation rate of blood is increased after exposure to neutron
or X radiation or the injection of Sr89 or Pu239 (Prosser, Painter, Lisco,
et al, 1947; Ross and Ely, 1947). The increase in sedimentation can be
attributed only in small part to the decreased hematocrit and, therefore,
is most likely related to some unknown factor in the plasma (Prosser,
Painter, Lisco, et al., 1947). The evidence indicates that this is probably

PHYSIOLOGY OF RADIATION INJURY 965

not fibrinogen. Since the sedimentation rate in dogs may be increased by
as little as 100 r to the whole animal, it is doubtful that this is a simple
reflection of bacterial invasion.

In general, the sensitivity of blood and blood-forming tissues increases
from rabbits to rats, mice, and chicks and then to men, goats, guinea pigs,
and dogs (Jacobson, Marks and Lorenz, 1949). Since this is roughly the
order of increasing sensitivity to the acute lethal action of X rays, rather
comparable blood changes appear in each species after a median lethal
exposure. Differences in recovery exist, however; after comparable
hematologic injury recovery is more rapid in the rat than in the rabbit.
There are also indications that the acute lethal dose for a species may be
related to the sensitivity of its heterophils. Similar lymphopenias have
been observed in the rabbit and guinea pig following irradiation with
200 r (Brues and Rietz, 1948). The heterophils, on the other hand, are
decreased markedly only in the guinea pig with this dosage. Comparable
heterophil depression is not observed in the rabbit until its lethal range is
approached. That there may be some correlation between the heterophil
response and acute lethality is also indicated by the rapid recovery of
heterophils, but not of lymphocytes, in mice treated with estrogens prior
to irradiation (Patt, Straube, et al, 1949).

While consistent hematological differences between survivors and
decedents are not always apparent, the clinical signs of acute intoxication
generally parallel the hematological findings (Rekers, 1949; Ingram and
Mason, 1950). It does not necessarily follow that death is due to blood
damage, but the fact that morbidity and mortality can be influenced
appreciably by procedures that presumably promote recovery of the
blood-forming tissues suggests that such damage is a substantial factor
in the radiation syndrome.

A number of agents have been employed in an attempt to modify the
hematological response to radiation. Reference has already been made
to the effectiveness of spleen shielding (Jacobson, Marks, Robson, et al,
1949), spleen transplants and homogenates (Jacobson, Simmons, Marks,
and Eldredge, 1951; Cole et al, 1952), and bone marrow injections
(Lorenz et al, 1951, 1952). Increased resistance of erythropoietic tissues
has been noted following the production of a regenerative anemia by
phlebotomy, phenylhydrazine, and exposure to high altitude (Jacobson,
Marks, Gaston, Simmons, and Block, 1948; Schack and MacDuffee,
1949). It is of interest that hyperplastic erythroid tissue shows less
injury than normal tissue. Pretreatment with estrogens has been shown
to induce a rapid recovery from radiation granulocytopenia and to
minimize anemia in mice (Patt, Straube, et al, 1949). The estrogen
effect may be related to stimulation of activity, in this instance, of the
myeloid tissues, following an initial depression. Cysteine is an effective
prophylactic against the hematological changes induced in rats and mice

966 RADIATION BIOLOGY

by radiation (Patt et al., 1950, 1952). Its protective action on blood is
probably not specific for the hematopoietic tissues but appears rather to
be related to an increase in the threshold for radiation effects generally.
Enhanced recovery of blood-forming tissue is also apparent when gluta-
thione is administered before irradiation (Cronkite et al., 1951). Vita-
mins and other nutritional factors have been used experimentally with
rather disappointing results (Simmons et at., 1946; Adams and Lawrence,
1947; Steamer, 1948; Goldfeder et al., 1948; Cronkite, Tullis, et al,
1950; Carter, Busch, and Strang, 1950). Many of these substances are
apparently useful in clinical radiation sickness (Shorvon, 1949). Post-
irradiation anemia in dogs can be minimized by frequent transfusions of
whole blood ; granulocytopenia and thrombocytopenia are not influenced,
however, nor is survival (Allen et al., 1951).

ABNORMAL BLEEDING

Abnormal bleeding is usually, but not always, associated with the
panhematopenia of acute irradiation. The bleeding tendency has been
variously attributed to the thrombocytopenia (Shouse et al., 1931; Dun-
lap; 1942; Holden et al., 1949; Cronkite, 1950; Rosenthal and Benedek,
1950; Penick et al., 1951; Dillard et al., 1951), blood vessel damage
(Rekers and Field, 1948; Field and Rekers, 1949a), and the presence of a
circulating anticoagulant (Allen and Jacobson, 1947; Allen et al., 1948).
It is probable that each of these factors contributes to radiation hemor-
rhage, though to a different degree, depending upon species and conditions
of irradiation. While hemorrhage is an almost invariable feature of the
acute radiation syndrome, the extent and distribution of bleeding may
vary considerably among the different species and among animals of the
same species. Its severity depends, moreover, upon the completeness of
irradiation and the dosage and manner of exposure.

Purpura and petechiae, though evident in the mouse, rat, and rabbit,
are most severe in the dog, man, and probably the guinea pig; bleeding
from the body orifices is not uncommon in these species (Allen and
Jacobson, 1947; Allen et al., 1948; Rekers and Field, 1948; Field and
Rekers, 1949a; Kohn and Robinett, 1948; Rosenthal and Benedek, 1950;
Shields Warren and Bowers, 1950). Hemorrhagic manifestations are
first seen about a week after irradiation in the median lethal range and
reach their peak during the second and third weeks. Extravasation of
red cells results in a bloody lymph which becomes manifest several days
after exposure and reaches a maximum during the period of greatest
bleeding (Bigelow et al., 1951). Bleeding may be widespread but is more
often localized in various subcutaneous sites and in the gastrointestinal
and urinary tracts, heart, and lungs. In general, the sites most subject
to trauma bleed first. The distribution of hemorrhage apparently differs

PHYSIOLOGY OF RADIATION INJURY 967

in animals which vary in susceptibility, being more generalized in the
dog than in the rat (Allen et al, 1948; Kohn and Robinett, 1948). In the
dog, localized hemorrhages in the terminal state sometimes give rise to
myocardial and neurological signs (Prosser, Painter, Lisco, et al, 1947).

Exposure to ionizing radiation may result in a decreased coagulability
of whole blood in animals and in human beings (Prosser, Painter, Lisco,
et al, 1947; Allen et al, 1948; Field and Rekers, 1949a; Silverman, 1949).
Allen and his associates (Allen and Jacobson, 1947; Allen et al, 1948;
Jacobson, Marks, Gaston, Allen, and Block, 1948) have observed a
prolongation of clotting time in acutely irradiated dogs which is related
to the appearance of a circulating heparin-like substance. The hemor-
rhagic diathesis in dogs is considered to be a consequence, for the most
part, of the circulating heparin-like material, since various antiheparin
substances, e.g., toluidine blue and protamine, can restore coagulability to
normal values and prevent hemorrhage even in the presence of thrombo-
cytopenia. The coagulation defect may occur before significant reduc-
tion of the blood platelets and in the absence of any change in pro-
thrombin time and in blood calcium and fibrinogen. It is not altered
significantly by transfusions of whole blood (Allen et al, 1951) or adminis-
tration of vitamins K or C. A similar clotting defect has been described
in rabbits (Jacobson, Marks, Gaston, Allen, and Block, 1948) and in man
(T. R. Smith et al, 1948) after treatment with nitrogen mustards. The
origin of the heparin-like substance is unknown; although it could arise
from mast cells, the exact role of these cells after irradiation is uncertain
(Kelsall and Crabb, 1952). It is of interest that chronic irradiation with
low dosages, although sufficient to result in blood damage and death, does
not appreciably alter blood clotting (Jacobson, Marks, and Lorenz, 1949).

While the influence of toluidine blue or protamine on the development
and disappearance of radiation hemorrhage is dramatic, these agents do
not alter total survival appreciably. It is noteworthy that the median
lethal dosage for the rat can be reduced to that for the dog by injecting
heparin after irradiation (Kohn and Robinett, 1948) ; yet, the distribution
of hemorrhage in the heparinized irradiated rat is apparently unchanged
despite the increase in bleeding. This may imply a difference between
the rat and dog in the latent injuries that require "heparin-like sub-
stances" for their development; it should also be recalled that the hemor-
rhagic picture in traumatic shock varies greatly from species to species.
As contrasted with these observations, heparinization is said to decrease
the lung damage that appears in rabbits following repeated irradiation of
the thorax over a two- week period (Boys and Harris, 1943). Dicumarol
has also been employed to decrease pulmonary changes in man after local
irradiation (Macht and Perlberg, 1950). In this case the role of vascular
thrombosis in local tissue radiation damage may provide the explanation.

The existence of a circulating anticoagulant is questioned by some

968 RADIATION BIOLOGY

workers who fail to find consistent changes in the coagulability of whole
blood in a number of species, although the clotting time of plasma may
be increased and clot retraction diminished (Kohn and Robinett, 1948;
Kohn et al, 1948; Dixon, 1948; Holden et al, 1949; Cronkite, 1950;
Rosenthal and Benedek, 1950; Cohn, 1951). Cronkite (1950) believes
that the bleeding tendency and the coagulation defect when observed can
be attributed primarily to the severe thrombocytopenia and associated
phenomena. Whole hemophilic blood with normal platelet levels has
been found to accelerate the clotting of irradiated dog blood (Penick et al.,
1951). This is not the case with platelet-poor hemophilic plasma.
Platelet transfusions have been shown to reverse the coagulation defect
and to prevent bleeding in an irradiated dog (Dillard et al., 1951).
Multiple transfusions of whole blood are ineffectual, however (Allen
et al., 1951). We cannot account for the lack of agreement regarding the
etiology of radiation hemorrhage, i.e., thrombocytopenia vs. heparin-like
substances. Some of the seemingly discordant observations may be
attributed to differences in experimental techniques, e.g., in the with-
drawal of blood and the determination of the various components of the
clotting mechanism, or in the species of animal and dosage of radiation.
Recent evidence indicates that thrombocytopenia is the chief factor in
the pathogenesis of abnormal bleeding.

There is some evidence that alteration in vascular fragility also plays a
role in the hemorrhagic syndrome. This is largely indirect, however.
Petechiae may be induced more readily in irradiated than in nonirradi-
ated animals and bleeding is generally most severe at the points of great-
est trauma (Field and Rekers, 1949a; Allen et al., 1948). Bleeding cannot
always be related to thrombocytopenia and the delayed clotting reaction
(Field and Rekers, 1949a). Reference has already been made to the
flooding of lymph by erythrocytes, indicative perhaps of capillary damage
(Bigelow et al., 1951). Rutin, a flavonol glucoside that is considered by
some investigators to influence vascular fragility, has been shown to
hasten recovery of the skin of rats after local irradiation (Griffith et al.,
1947) and to minimize hemorrhage and improve the survival of dogs after
total-body irradiation (Rekers and Field, 1948; Field and Rekers, 1949a).
To be effective, rutin must be given for a week or so before or preferably
both before and after irradiation. Since reduction of hemorrhage occurs
with rutin despite depression of the blood platelets and prolongation of
clotting, it has been proposed (Field and Rekers, 1949a) that rutin either
decreases the rate of vascular disintegration or increases its repair. It is
not known whether hyaluronidase plays a role in altered capillary fra-
gility. Hyaluronidase inhibitors have been tried in a few irradiated dogs
without success (Field and Rekers, 1949b). Bleeding is said to be dimin-
ished in guinea pigs that are fed a diet supplemented with cabbage,
possibly because of its high content of vitamins P and C (Lourau and

PHYSIOLOGY OF RADIATION INJURY 909

Lartigue, 1950b). On the other hand, when beets are added to the basic
ration, the number of hemorrhages is increased and the LD50 is decreased
from 350 to 150 r. This may be attributed possibly to a relative defi-
ciency of vitamin C. Ascorbic acid does not alter the hemorrhagic state
or the survival of irradiated dogs although it may act synergistically with
flavonoids (Field and Rekers, 1949b). Flavonoids have been shown to
improve the survival of X-rayed guinea pigs and rats, but this has not
been verified by others, and the value of these substances requires further
elucidation (Kohn, Robinett, and Cupp, 1948; Clark et al., 1948; Sokoloff
et al, 1950; Cronkite et al, 1949).

It is of interest that rutin does not protect the dog against dosages
greater than the LD50 (Field and Rekers, 1949a). This suggests perhaps
that the hemorrhagic tendency in the dog is a minor factor in mortality
resulting from higher dosages of radiation. While hemorrhage may be
extensive at the LDioo, its control with toluidine blue or protamine does
not permit survival (Allen et al., 1948). Life may, however, be pro-
longed for a week or two, suggesting perhaps that the control of hemor-
rhage in less severely irradiated animals may contribute to recovery. It
is well known that many animals, including the dog, succumb with only
minor evidence of hemorrhage, again emphasizing the importance of
multiple factors.

BODY FLUIDS

Some disturbance in water and electrolyte metabolism may be antici-
pated following exposure of large areas of the body to ionizing radiation as
a result of the vomiting, diarrhea, altered food and water intake, tissue
breakdown, and bleeding, which are prominent features of the radiation
syndrome. The degree of redistribution of body fluids will depend, in
large measure, on the capacity of the homeostatic mechanisms, which may
also be affected by radiation, to maintain the delicate balance of the
internal environment. It is clear, therefore, that the body fluids reflect a
number of influences that may vary not only with dosage and from animal
to animal but also from time to time after irradiation in the same animal.
While it appears that early death after massive irradiation is accom-
panied by hemoconcentration and dehydration, disturbances of water
and electrolyte balance after median lethal dosages are, in general, com-
paratively mild and probably do not constitute an important cause of
death.

Blood and Plasma Volume. The blood volume remains relatively con-
stant or is only slightly diminished after irradiation of the whole body
with dosages that approximate the median lethal. This obtains in the
face of a greatly reduced erythrocyte volume because of the compensatory
increase in the volume of plasma (Prosser, Painter, Lisco, et. al., 1947;
Storey et al., 1950; Soberman et al., 1951). The decrease in red blood

970 RADIATION BIOLOGY

cell volume has been attributed to the failure of erythropoiesis, destruc-
tion of the mature cells, and internal bleeding. Cell and plasma volumes
have been determined in mice, rats, rabbits, and dogs, using P32 tagged
erythrocytes, iodinated plasma proteins, and the conventional Evans
blue technique. Following irradiation of the rabbit with 1000 r, there is
a parallel decline in cell and plasma volumes that begins on the third
day. By the tenth day, however, plasma volume rises, reaching a level
some 10 per cent above the normal during the third week when the red
cell volume is decreased by more than 40 per cent (Storey et al, 1950).
The blood volume is, therefore, only slightly diminished. In the irradi-
ated dog, plasma volume may be increased by more than 50 per cent when
the red cell volume and hematocrit are severely depressed (Soberman
et al., 1951). Rather similar findings have been observed in mice and
rats (Storey et al., 1950; France, 1946). Although there is some disagree-
ment in regard to the early decline in plasma volume, this may be attrib-
uted in some instances to whether the volume is expressed as a percentage
of the preirradiation body weight or of the body weight at the time of
determination (Storey et al., 1950; France, 1946). The rapidly falling
blood pressure that is seen frequently in rabbits during the first hours
after irradiation is not accompanied by significant changes in blood
volume (Painter et al., 1947).

When massive dosages are applied to the abdomen or whole body of
the dog, death nearly always occurs during the first few days with hemo-
concentration (Moon et al., 1941). With less severe, but nevertheless
lethal, exposure to X radiation or after injection of Sr89 or Pu239, the
usual rise in plasma volume is seen during the intermediate and acute
periods as the red cell loss becomes appreciable (Prosser, Painter, Lisco,
et al., 1947). Thus, as contrasted with traumatic shock, hemoconcentra-
tion and appreciable diminution of blood volume are not observed in the
irradiated animal unless the radiation dosage is well above the minimal

LUioo-

There is no information concerning the blood volume after superficial
irradiation, e.g., with 0 rays. A decrease in blood volume would perhaps
be anticipated because of plasma loss in the burned areas. It is possible,
however, that plasma loss might occur so slowly that compensatory
mechanisms, e.g., shift in fluids from other compartments or water
retention, could operate to prevent a change in blood volume. Similar
information is also lacking for neutron irradiation ; it is probable that the
effects with neutrons would resemble those seen after X irradiation.

Lymph and Tissue Fluid. The output of lymph from the thoracic duct
of cats is unchanged during the first hours after 1500 r X irradiation of
the whole body (Valentine et al., 1948). Lymph flow, as visualized by
thorotrast, is not affected following localized X irradiation of the hind
legs of rats (Hodes and Griffith, 1941). The volume of cutaneous lymph

PHYSIOLOGY OF RADIATION INJURY 971

flow in man is likewise unchanged by moderate amounts of local irradia-
tion and may actually decrease with larger dosages (Ane and Burch,
1941). However, an increased spread of intradermally injected Evans
blue dye, presumably indicative of an enhanced lymph flow, has been
seen in the rabbit within a few hours after irradiation (Painter et al., 1947).
Lymph output has not been measured in the later periods after exposure,
although there is some indication of an increased flow from the thoracic
duct in irradiated dogs (Bigelow et al., 1951). Reference has already
been made to extravasation of red cells into the lymph (Furth, Andrews,
et al., 1951).

Edema, especially of the face, neck, and extremities, may be observed
during the acute terminal period. While edema of the viscera, particu-
larly of the gastrointestinal tract, may also be noted in some animals,
edema is more often microscopic rather than macroscopic. Consistent
changes in water content have not been observed in muscle, adrenal,
kidney, and gastrointestinal tract (France, 1946; Beutel and Winter,
1935; Patt et al, 1947; Painter, 1948; Bowers and Scott, 1951b). In
chronically irradiated rats, muscle and bone water increase slightly after
the accumulation of nearly 1500 r over a period of 105 days (Brues et al.,
1946). Total-body water in mice is increased by about 8 per cent on the
fifth day after acute X irradiation (France, 1946), but there is little
change in the total water content of dogs following a nearly completely
lethal dosage of X rays (Soberman et al., 1951).

Extracellular fluid in rats, estimated from the thiocyanate space, has
been shown to increase by about 2 cc per 100 grams of body weight during
the first two weeks after whole-body X irradiation with 400 or 700 r. An
increase over the range of the control animals has also been noted eight
months after a single exposure to 750 r, and similar changes occur after
chronic irradiation with X rays or a rays from plutonium (France, 1946).
Although an early increase in the extracellular compartment has been
reported in a few irradiated dogs (Prosser, Painter, Lisco, et al, 1947),
consistent changes are not evident from other experiments in the same
dosage range (Soberman et al., 1951).

Sodium retention has been described in the rat during the first few days
following irradiation (Painter, 1948; Bennett et al., 1949), but similar
increases in sodium space do not occur in the dog after lethal irradiation
(Soberman et al., 1951). The lack of agreement may be related to differ-
ences in water intake between the two species, since the rat generally
manifests a marked polydipsia during the first post irradiation day while
water intake is reduced in the dog (Prosser, Painter, Lisco, et al., 1947).
On the other hand, there is reason to think that inanition, and not
polydipsia, is responsible for the increased sodium space in the rat since
similar changes have been seen in the gastrointestinal tracts of starved
and both starved and irradiated rats (Painter, 19-18). Whenever body

972 RADIATION BIOLOGY

tissue is catabolized, for example, in starvation, water is made available,
and this may be a factor in the animal which is anorexic as a result of
irradiation. Changes in the irradiated animal, however, are not entirely
comparable to those observed in starvation; there is apparently little
change in extra- and intracellular muscle water in the irradiated rat,
whereas, in the starved rat, the extracellular phase is expanded and the
intracellular contracted.

Water transfer is closely related to the exchange of its principal ionic
solutes and both solvent and solute must be considered in any evaluation
of hydration phenomena. Although there is evidence that heavy irradia-
tion may lead to cell swelling (Buchsbaum and Zirkle, 1949; Failla,
1940), simultaneous measurements of sodium, potassium, and chloride
indicate that intracellular fluid may actually be decreased at the expense
of the extracellular phase in the gastrointestinal tract of rats after mid-
lethal exposure of the whole body (Painter and Pullman, 1950). It is of
interest that there is a rather parallel loss of sodium and potassium from
radiosensitive tissues in the rat during the first day or two after X irradia-
tion (Bowers and Scott, 1951a, b). This is followed by a marked increase
in tissue sodium. There is also no evidence for an increase in intracellular
water in irradiated dogs (Soberman et al., 1951).

These considerations indicate that there is probably little serious dis-
turbance in water balance after acute lethal irradiation. Changes appear
to be more severe in rats than in dogs, which may be a result of the greater
diarrhea in the former. Water intake is reduced more than water loss in
most mammals. Since plasma volume may be increased in the presence
of an increase in extracellular and total-body water, the extra water may
be derived from metabolism or from a reduction of the respiratory loss,
which may be as high as 35 per cent of the extrarenal loss under basal
conditions.

Constituents of Plasma and Lymph. Plasma proteins are decreased in
concentration in guinea pigs and rats, but usually not in dogs, during the
first week after total-body X irradiation. The maximum decrease,
amounting to about 1 gram per cent, occurs around the fifth postirradia-
tion day in guinea pigs and rats (Kohn, 1950, 1951a; Hanschuldt and
Supplee, 1949). The plasma refractive index parallels early protein
change. After the initial decrease, protein concentration levels off or
may return toward the normal value. In the dog, plasma protein con-
centration remains unchanged or may decline slightly during the first
and second postexposure weeks (Prosser, Painter, Lisco, et al., 1947a;
Soberman et al., 1951). This may be followed by a rise above the control
level in nonsurvivors during the terminal period. Total protein of
lymph collected from the thoracic duct fluctuates during the first two
days after irradiation and then decreases slowly for several days (Brown
et al., 1950). Maintenance or increase of the plasma protein concentra-

PHYSIOLOGY OF RADIATION INJURY 973

tion in the presence of a rising plasma volume suggests an increased
breakdown of tissue that liberates protein, a failure of mechanisms of
proteolysis, or an increased synthesis of protein. Animals exposed to
small daily doses of X rays do not exhibit an initial decline in plasma pro-
tein, and in the terminal period total protein may rise, fall, or remain
unchanged (Prosser and Moore, 1946; Fink, 1946). Definite changes in
plasma protein have not been seen in irradiated patients (Frieden and
White, 1950).

The albumin-globulin ratio as determined by salt fractionation is
elevated for several days after irradiation of the guinea pig or rat (Kohn,
1950, 1951a). The early increase in the A/G ratio is thought to be due
to the presence in plasma of an ether-soluble factor that affects the
fractionation procedure (Volkin and Kohn, 1950). There is no charac-
teristic change in the electrophoretic pattern of plasma proteins in dogs
during the first postexposure week (Muntz et al'., 1949). Albumin is
only slightly diminished during this interval, but a marked decrease,
amounting to 50 per cent, is found during the second week. Several days
before death the a-globulin fraction increases sharply, especially the
a3 and a4 fractions, and may actually be doubled. The /3-globulin-
fibrinogen complex is also increased but not so markedly as that of the
a-globulin. Similar changes occur in dogs treated with daily doses of
X rays or injected with Sr89 and Pu239 (Prosser, Painter, Lisco, et al.,
1947). Differences in the electrophoretic pattern have been noted in
dogs and goats after X irradiation (Buchanan and Barron, 1947). In the
latter, increase in the jS-globulin-fibrinogen fraction predominates and
little change is seen in the a-globulin.

In contrast with the results observed in X-irradiated dogs, in neutron-
irradiated rabbits there is an initial decrease in the y-globulin fraction
which parallels the decline in leukocytes (Sanigar et al., 1947). This,
however, is followed by the increase in globulins and decrease in albumin
that is seen with X rays. Direct neutron irradiation of blood plasma
does not influence the electrophoretic pattern. The changes observed in
irradiated animals are nonspecific and related presumably to tissue
destruction, infection, and inanition. Similar patterns have been found
after thermal and other injuries and after the injection of adrenal cortical
hormones (Gjessing and Chanutin, 1947, 1950). It is not known whether
the diminution in albumin is due to impaired synthesis or to its loss from
the blood because of altered capillary permeability. Albuminuria is not
evident (Muntz et al, 1949).

Plasma nonprotein nitrogen rises by over 25 per cent in rats and guinea
pigs during the first few days after exposure to X rays (Kohn, 1950,
1951a). On the other hand, there is little change or a gradual decline in
the dog until the terminal period when a marked increase is manifest
(Prosser, Painter, Lisco, et al, 1947). A definite elevation of NPN and

974 RADIATION BIOLOGY

uric acid levels occurs in the lyrnph 4 hours after X irradiation and
coincides with the period of greatest lymphocyte destruction (Brown
et al., 1950). Part of the late rise in plasma nonprotein nitrogen in dogs
can be accounted for by the terminal anuria. Plasma urea nitrogen
follows closely the change in nonprotein nitrogen. There is little altera-
tion in nonprotein nitrogen with daily dosages of 10 to 50 r, although a
sharp fall may be noted terminally with the higher dosage (Prosser and
Moore, 1946). The reason for this difference in terminal behavior
between dogs irradiated acutely and chronically is not clear. After
neutron exposure (400 n in four divided doses) plasma NPN in the dog
falls on the third day and is decreased until death (Ross and Ely, 1947).

Plasma glucose is increased in the guinea pig and rat (Kohn, 1950,
1951a; Lourau and Lartigue, 1950a). The change roughly parallels that
of nonprotein nitrogen, and it has been suggested that the gluconeogenesis
necessary to maintain hyperglycemia during the period of radiation
anorexia is partly responsible for the rise in NPN (Kohn, 1950). Plasma
glucose is also increased in the rabbit and dog after neutron or X irradia-
tion (Ross and Ely, 1947; Prosser, Painter, Lisco, et al., 1947). In the
X-irradiated dog, blood sugar is increased for several days prior to the
onset of fever and normal levels are found terminally. This is appar-
ently not the case with neutron irradiation where hyperglycemia is
progressive until death. Glucose concentration of lymph is unaltered
during the first few days after X irradiation (Brown et al., 1950).

There is a variable pattern of change in the principal electrolytes of
plasma. Chloride rises in the rat and guinea pig after comparable dosage
(Kohn, 1950, 1951a). The increase occurs on the third day and normal
levels are evident by the second week. Blood cell chloride declines
roughly in proportion to the plasma increase; thus, the total chloride is
not altered appreciably. Plasma inorganic phosphorus rises slightly in
the rat and parallels the hyperglycemia and elevated NPN (Kohn, 1951a).
Serum sodium is also elevated in this species, while serum potassium may
rise slightly or remain within normal limits (Bowers and Scott, 1951a, b;
Bennett et al., 1949; Kohn, 1951a). Consistent changes in the plasma
levels of sodium, potassium, calcium, chloride, and phosphate have not
been observed in dogs following acute or chronic exposure to X rays
(Soberman et al., 1951; Prosser and Moore, 1946; Fink, 1946), and only
minimal effects have been seen following neutron irradiation (Ross and
Ely, 1947). There are few recent data relating to acid-base balance;
carbon dioxide combining power is said to be decreased in rats on the
fourth day after a total-body exposure to 600 r (Kohn, 1951a).

Increased amounts of lipids have been detected in several mammalian
species following total-body irradiation. Turbidity of plasma, presum-
ably a result of the large increase in total lipid, has been observed in the
rabbit and is believed to presage early death in this species (Rosenthal,

PHYSIOLOGY OF RADIATION INJURY 975

1949). Plasma turbidity is not apparent, however, in the irradiated rat,
although the total lipid content of plasma is elevated (Kohn, 1951a).
Plasma phospholipid may rise or remain unchanged (Kohn, 1951a;
Neve and Entenman, 1951), while plasma cholesterol is reported to
increase in guinea pigs, mice, rats, and dogs (Prosser, Painter, Lisco,
et al., 1947; Kohn, 1950, 1951a; Low-Beer, 1933). In the rat and guinea
pig cholesterol rises on the second or third day after acute irradiation and
returns to normal by the sixth day, the pattern resembling that of
chloride. In chronically irradiated dogs (50 r per day) cholesterol is
elevated only terminally. There are no significant changes in cholesterol
concentration with daily dosages of 10 r or less (Fink, 1946). Alteration
in the cholesterol content of serum and in the chylomicron count (mobile
visible plasma lipids) has been observed in patients receiving radiation
therapy. Several investigators have alluded to a relation between the
direction of cholesterol change and clinical radiation sickness; the latter
is not seen in patients whose serum cholesterol is elevated (Low-Beer,
1933; Hummel, 1930; Holmes, 1941). Reduction of the chylomicron
count also appears to be related temporally to radiation sickness and is
observed after irradiation of the abdomen or thorax but not of the
extremities (Setala and Ermala, 1948).

A moderate increase in plasma alkaline phosphatase occurs in rats
24 hours after irradiation with dosages of 200 to 600 r (Ludewig and
Chanutin, 1950a). Enzyme activity decreases on the second day in
animals exposed to the higher dosage. Phosphatase activity in the rat is
also depressed by multiple small dosages of radiation. There is little
change in alkaline and acid phosphatases of plasma after chronic X
irradiation of dogs (Prosser and Moore, 1946; Fink, 1946).

The sulfhydryl content of plasma is decreased in the rat about 5 days
after X irradiation with 750 r (Shacter, 1951); the significance of this
change is not understood. Similar effects have been observed after
nitrogen mustard poisoning and surgical trauma and appear to be related
chronologically to initiation of tissue regeneration. A fall in serum
ascorbic acid has also been reported in animals and man (Kretzschmar
and Ellis, 1947). The maximum decrease is seen about one week after
exposure.

Plasma histamine is increased in the irradiated rat and rabbit (Painter
et al, 1947; Weber and Steggerda, 1949). In the rat given 600 r, two
peaks of histamine elevation are seen, the first two hours after exposure
and the second at five days (Weber and Steggerda, 1949). There is some
reason to believe that histamine liberated from damaged cells and perhaps
newly formed from irradiated histidine can account for certain of the
radiation effects (Ellinger, 1951). Other toxic materials may be present
in the plasma of irradiated animals, but these have not been identified
(Venters and Painter, 1950).

976 RADIATION BIOLOGY

Aberrations in the plasma constituents are rarely related in specific
experiments to changes in water distribution, or to renal and gastro-
intestinal function, and this aspect of radiation injury deserves study.
All the changes that have been described appear to be nonspecific reac-
tions to the injury, and some of these are undoubtedly mediated by the
pituitary-adrenal system. Yet, differences are apparent between these
radiation effects and the alarm reaction (Kohn, 1950, 1951a, b). The
pattern of change is said to be similar in young and adult animals and
is about equal in the rat and guinea pig after comparable exposure,
despite the greater sensitivity of the latter to the lethal action of radiation.
The inconsistency of the plasma changes following median lethal irradia-
tion, not only among different species but also among animals of the
same species, suggests, moreover, that these effects are not critical
parameters of radiation injury and probably do not constitute an impor-
tant cause of death.

CARDIOVASCULAR SYSTEM

The erythematous reaction of skin is not only the most evident but
perhaps also the most extensively studied circulatory response to radia-
tion, since it plays an important role in determining the course of radia-
tion therapy. It is well known that a smaller dosage of low-energy
radiation is necessary to produce skin erythema than of high-energy
radiation and that the biologic effectiveness decreases with protraction or
fractionation of radiation (Shields Warren, 1943c; Schottelndreyer,
1949; Larkin, 1941, 1942). The severity of the cutaneous erythema
depends, moreover, on the size of the radiation field (Shields Warren,
1943c; Jolles, 1941, 1950; Jolles and Mitchell, 1947). Erythema has
been described following exposure to the different qualities of radiation,
and there is little difference in the reactions that are produced (Miescher,
1938; Shields Warren, 1943c; Larkin, 1941, 1942). The 7-to-X-ray
ratio of effectiveness for cutaneous erythema is about 1.3 (Mottram and
Gray, 1940).

Waves of erythema were originally defined by Miescher (1924) and
may be seen in human skin during the first day, the second to third
week, and at the end of the first month after irradiation. Telangiectasis
may appear after heavy irradiation. Similar patterns occur in rabbit
skin after local exposure to X rays. With 1000 r, erythema is seen on the
first, fifth, and tenth postirradiation days (Painter et at., 1947). The
early cutaneous effects have been related to cell injury, the later erythema
to alterations in the vascular bed (Shields Warren, 1943c). An increase
in the number of patent blood vessels and dilatation of existing capillaries
and lymphatics may be noted in irradiated skin (Painter et al., 1947;
Borak, 1942a, b, c; Pohle, 1926, 1927; Pendergrass et al, 1944). The
bluish cast that is sometimes observed in a hyperemic area after heavy

PHYSIOLOGY OF RADIATION INJURY 977

local irradiation is thought to be due to alternate regions of constriction
and dilatation in the same vessels (Borak, 1942a, b, c). Several mecha-
nisms have been proposed to account for the vascular reactions, including
a direct effect of radiation on blood vessel walls and the release of humoral
agents (H substances) in the irradiated area (Ellinger, 1951 ; Light, 1935).
Diffusible substances are postulated to account for the observation that
two radiation fields a distance apart show less injury than areas that are
closer together (Jolles, 1950).

Scratch tests of erythematous skin lead to a persistent vasoconstriction,
and wheal formation is not induced by histamine (Larkin, 1942). Skin
temperature is unchanged during the latent period between irradiation
and erythema, but, with the appearance of the latter, the temperature
may rise several degrees centigrade, suggestive of marked vasodilatation
(Larkin, 1942). It has been pointed out that the cutaneous vessels show
an increased sensitivity to dilator substances and a decreased responsive-
ness to constrictor stimuli during the period of skin erythema (Lazarew
and Lazarewa, 1926). Blanching of the hyperemic regions can occur
after injection of epinephrine, however (Larkin, 1942).

Direct in vivo observations of blood vessels in the frog's web or bat's
wing reveal that radiation is a nonspecific vascular damaging agent
(Painter et al., 1947; Smith, Svihla, and Patt, 1951). A similar conclu-
sion may be drawn from studies of the nail-fold area (Braasch and Nick-
son, 1948). Detailed studies of circulation have been made in the wing
of the bat, Myotis lucifugus, following total-body irradiation and following
local irradiation of a portion of the wing (Smith, Svihla, and Patt, 1951).
Of interest is the radioresistance of the hibernating bat.1 Circulatory
changes do not occur in the wing unless the total-body dosage with
250-kv X rays exceeds 10,000 r. With dosages over the range of 10,000
to 60,000 r, adherence of leukocytes to blood vessel walls, clumping of red
cells, and stagnation of blood are prominent. There are, however, no
consistent changes in vessel diameter or in venomotor activity following
a total-body exposure and hemorrhage is not apparent. On the other
hand, there is some evidence of an increase in capillary permeability.
After local irradiation of the wing, vascular reactions are confined to the
irradiated area, and the threshold for circulatory disturbance is in the
neighborhood of 50.000 r for 50-kv X rays. With the exception of red
cell clumping, the intravascular changes noted after local exposure, e.g.,
leukocyte sticking and clumping, platelet thrombi, and stagnation, appear
to be related to radiation dosage. Platelet thrombi and leukocyte clumps
have also been observed in tissues taken from patients and from animals
treated with radium and X rays, and it is presumed that these changes
lead to an impairment of blood flow (Pullinger, 1932). Hepatic blood

1 Although the bats were collected while in hibernation, observations were carried
out at room temperature.

978 RADIATION BIOLOGY

flow is decreased only by a small factor after heavy 13 irradiation of the
mouse liver with a Y90 colloid (20,000 rep in 3 days) (Dobson and Jones,
1952).

The influence of radiation on vascular permeability is largely undeter-
mined. From studies of relatively simple systems there is reason to
believe that cell permeability is not affected directly unless massive
dosages are employed. Although changes suggestive of altered capillary
permeability have been described, it is not known whether they are a
result of the action of radiation on the endothelium or of alterations in the
vascular bed. Intravenously injected dyes appear sooner in patches of
irradiated skin. When Evans blue dye is injected into rabbits immedi-
ately after local X irradiation, the exposed area turns blue earlier than the
nonexposed (Painter et al, 1947). A similar increased localization and
concentration of dye has been noted with intravenous trypan blue as
long as the injection is made within 1 hour of X irradiation (Rigdon and
Curl, 1943). An increased spread of intradermally injected Evans blue,
which is presumed to be indicative of an accelerated lymph flow, has also
been seen in the irradiated skin of rabbits during the first few hours
(Painter et al, 1947); yet, cutaneous lymph flow in man is apparently
unchanged following moderate amounts of radiation and may actually
decrease with larger dosages (Ane and Burch, 1941). There is little
change, moreover, in the output of lymph from the thoracic duct of the
cat during the first hours after total-body irradiation (Valentine et al,
1948), although an increased lymph flow has been observed in the dog
several days later (Bigelow et al., 1951).

Intravenous Evans blue appears in the irradiated and nonirradiated
regions at the same time during the late erythematous reaction of rabbit
skin, but this is not the case with fluorescein, which is seen within a few
seconds in the exposed area and within several minutes in the nonexposed
region (Painter et al., 1947). The reason for this difference is not obvious.
It has been suggested that the appearance of Evans blue may be masked
by erythema of the irradiated skin. The rapid appearance of fluorescein,
on the other hand, may be more a matter of hyperemia than of an increase
in permeability of the vascular endothelium.

Furth and his associates (Furth, Andrews, et al, 1951; Bigelow et al,
1951; Wish et al, 1952) have contributed several significant papers that
suggest that an important part is played by endothelial damage and
increased permeability after total-body irradiation. They have deter-
mined that labeled homologous and heterologous plasma, homologous
and heterologous erythrocytes, Evans blue, and colloidal radiogold, in
general, disappear faster from the circulation of X-rayed than of normal
mice and rabbits. Maximum alteration occurs during the second post-
exposure week and may be correlated with the period of greatest bleeding.
As contrasted with the above observations, a decreased disappearance of

PHYSIOLOGY OF RADIATION INJURY 979

circulating Evans blue has been described in dogs by other investigators
(Prosser, Painter, and Swift, 1947) ; yet plasma albumin, to which Evans
blue is bound, is said to be diminished at the time of greatest reduction in
dye disappearance. An increase in the rate of dye disappearance might
be expected if the decline in albumin were due to its leakage through
capillaries with increased permeability to colloids. The discrepancy in
dye disappearance is not easily resolved ; in the dog experiments determin-
ations were made on the same animals at different postirradiation times,
whereas with mice and rabbits, single determinations were made on
paired control and irradiated animals.

That augmented disappearance of various tagged substances in mice
and rabbits is indicative of increased permeability is perhaps strengthened
by the finding that macrophage function is not altered appreciably (Wish
et al., 1952; Barrow et al., 1949). Although colloidal gold leaves the
circulation more rapidly, the absolute amount that is retained in the
liver and spleen is essentially similar in both control and irradiated
animals. It is conceivable that the apparent increase in permeability
results, in part, from intravascular changes such as cell clumping and
platelet thrombi, which lead to stagnation and decrease in the effective
blood volume, although local anoxia and increased hydrostatic pressure
would favor leakage under these conditions. Since capillary obstruction
has been noted in in vivo preparations (Smith, Svihla, and Patt, 1951), it
is difficult to guess the extent to which heightened capillary permeability
is apparent or real and whether this is a direct or indirect result of the
action of radiation on the vascular endothelium. It should be pointed
out that there is no evidence for increased permeability of renal glomeruli
and, therefore, that altered permeability following irradiation is probably
not typical of capillary endothelium.

Vascular fragility is thought to be increased after irradiation since
hemorrhage is not always related to the thrombocytopenia and delayed
clotting, and mild trauma frequently results in showers of petechiae
(Field and Rekers, 1949a). Increased fragility is suggested further by
the rapid disappearance of labeled erythrocytes from the circulation and
by the presence of blood in the lymph of X-rayed rats and dogs several
days postexposure (Bigelow et al., 1951; Wish et al., 1952). Bleeding
may be minimized, moreover, by certain of the flavonoids although this
has not been verified universally (Rekers and Field, 1948; Field and
Rekers, 1949a; Kohn, Robinett, and Cupp, 1948). Rutin is reported to
antagonize hyaluronidase (Beiler and Martin, 1947), but the enzyme
probably does not play a role in altered fragility. Other hyaluronidase
inhibitors fail to influence radiation toxicity (Field and Rekers, 1949b),
and hyaluronidase itself is inactivated by irradiation in vitro (Schoenberg
et al., 1950). It is, of course, possible that the relatively small dosages
of radiation that are required to induce bleeding result in direct degrada-

980 RADIATION BIOLOGY

tion of hyaluronate. While the decline in platelets and appearance of
heparinoidlike substances are probably largely responsible for capillary
bleeding, it is likely that other factors also play a role in its development.

The typical acute reaction to radiation injury may embarrass the
circulation initially and terminally; the intermediate period, however,
appears to be relatively free of any gross circulatory disturbance. This
applies to penetrating radiation; unfortunately there are no data that
relate to circulation after superficial irradiation other than local blood
vessel effects. It is, of course, well known that these responses are
separated in time by periods of relative recovery. A shocklike state has
been seen in dogs following massive X irradiation of the whole body or of
the abdomen (Moon et al., 1940, 1941). This is characterized by progres-
sive hemoconcentration, dehydration, and hypotension, with death occur-
ring several days after exposure. The severity of this type of early
reaction to large radiation dosages often parallels the recognizable tissue
damage. Post-mortem visceral changes are described as those character-
istic of circulatory failure of the shock type, and the physiological dis-
turbances have been thought to be a consequence of acute toxicity result-
ing from absorption of the products of tissue necrosis, although, as in the
case of irreversible shock from other causes, this has been difficult to
confirm experimentally.

The shocklike syndrome is generally not observed after median lethal
irradiation. Early circulatory changes may appear in some species, how-
ever. A fall in blood pressure has been detected in rats and rabbits but
not in dogs during the first 24 hours after X irradiation (Prosser, Painter,
Lisco, et al., 1947; Painter et al., 1947; Weber and Steggerda, 1949;
Strauss and Rother, 1924). Hypotension is reported in the rabbit after a
total-body exposure as low as 50 r (Painter et al., 1947). A similar
early decrease in blood pressure has been noted in patients receiving
X-ray therapy (Bediirftig and Griissner, 1949). There is reason to
believe that part of the early hypotension is reflex, since vagotomy and
atropinization can reduce the blood pressure response (Strauss and
Rother, 1924; Painter et al., 1947). That the immediate hypotension is
neural in origin is suggested further by its absence in the spinal animal
(Montgomery and Warren, 1951). Repeated injections of epinephrine
are also effective in antagonizing initial hypotension in rabbits (Painter
et al., 1947). Blood pressure changes have been described after irradia-
tion of the hind legs, but the effects are not consistent. The direction of
early blood pressure change may depend upon the area that is exposed
(Toyoma, 1933b). A small increase in blood pressure, which is not
affected by atropine or nicotine, has been observed during irradiation of
the cardiac area. A rise also occurs during exposure of midbrain; when
irradiation includes the entire head of the animal, there is a decrease that
can be prevented by vagotomy.

PHYSIOLOGY OF RADIATION INJURY 981

Plasma histamine rises in the rat and rabbit and this is related in time
to the fall in blood pressure (Painter et al., 1947; Weber and Steggerda,
1949). The presence of depressor substances receives further support
from transfusion experiments (Painter et al., 1947). It is of interest that
vagotomy, atropine, or benadryl reduces the blood pressure response in
the rabbit by about half (Painter et al., 1947). It might be expected that
vagotomy or atropine on the one hand and the antihistamine benadryl,
on the other, would be additive in their effect on blood pressure. This is
not the case, which may imply that benadryl is acting as a parasympatho-
lytic agent or that histamine effects are decreased in the absence of
parasympathetic activity. The temporal relation between histamine
appearance and hypotension may, of course, be fortuitous, and other
factors could be responsible for the lowering of blood pressure (Mont-
gomery and Warren, 1951).

After the initial hypotension, arterial pressure recovers to a level
somewhat below normal for a few days and then declines gradually. In
the terminal period, blood pressure is frequently below 50 mm Hg
(Painter et al., 1947). Relatively few measurements have been made in
the dog. Blood pressure is said to be constant in this species until the
acute terminal period (several days before death), when it is reduced by
about 25 per cent. Similar changes occur in acutely and chronically
X-irradiated animals (Prosser, Painter, Lisco, et al., 1947; Prosser and
Moore, 1946).

Heart rate is unchanged in the dog, but it increases in the rabbit during
the first few hours (Prosser, Painter, Lisco, et al., 1947; Painter et al.,
1947). The latter probably represents a compensatory response to the
initial hypotension. During the terminal period preceding death, pulse
rate is increased in the dog but not always in the rabbit. Myocardial
hemorrhages and abnormal electrocardiographic findings, e.g., lowered
take-off level and sometimes reversal of the T wave, are described in dogs
dying after acute or chronic X irradiation (Prosser, Painter, Lisco, et al.,
1947; Prosser and Moore, 1946). Terminal myocardial damage has also
been seen in a small series of dogs following exposure to cyclotron neu-
trons (Ross and Ely, 1947).

While radiation may exert a direct local action on blood vessels, the
myocardium appears to be quite resistant (Leach and Sugiura, 1942) ;
changes that are observed after irradiation of the whole body are probably
consequences of autonomic and humoral influences initially, and of local
extravasation of blood and anoxia terminally (Prosser, Painter, Lisco,
et al., 1947). On the whole, the evidence for a generalized circulatory
disturbance after median lethal irradiation is not impressive, although
this seems to be a factor in relation to massive dosages. After smaller
amounts of radiation, early severe hypotension and death are seen in
some species, and failure of the circulation may develop during the last

982 RADIATION BIOLOGY

few hours or days of life. Little can be said about the efficiency of the
circulation to specific sites or the capacity of the cardiovascular system
to compensate for any excessive demands, such as repeated transfusions
of whole blood or plasma, that may be placed upon it. These considera-
tions deserve attention, for they pose a number of practical problems,
especially in the event of combined radiation and traumatic injuries.

GASTROINTESTINAL TRACT

Many of the manifestations of irradiation of the whole body are refer-
able to the gastrointestinal tract, which constitutes a sensitive locus for
radiation action. The nausea, vomiting, and anorexia of clinical radia-
tion sickness were first described by Walsh in 1897, and degeneration of
the intestinal mucosa of irradiated animals was reported by Krause and
Ziegler in 1906. Perhaps the most significant of the early observations
were those by Regaud, Nogier, and Lacassagne in 1912 and by Whipple
and his associates in 1919-1923. Regaud et al. (1912) pointed out that
the small intestine of the dog is more sensitive to direct X irradiation than
the stomach or colon and that the duodenum and jejunum are the most
sensitive regions of it. Hall and Whipple (1919) and Stafford L. Warren
and Whipple (1922a, b, c, d; 1923 a, b, c, d) described a toxic reaction
following heavy irradiation of the abdomen, which was most marked at
the time of greatest injury to the crypt epithelium and resembled intoxica-
tion resulting from intestinal obstruction or severe nonspecific intestinal
injury. The effects of X irradiation of the abdomen were also shown to
be more severe than those following exposure of other portions of the
body. Similar conclusions may be drawn from more recent investiga-
tions (Moon et al, 1940; Friedman, 1942; Shields Warren and Friedman,
1942; Bond et al, 1950; Quastler et al, 1951).

Early extensive gastrointestinal injury followed by death within
several days is seen after massive dosages of radiation delivered either
to the abdomen or to the whole body (Hall and Whipple, 1919; Moon
et al, 1941; Quastler et al, 1951). Intestinal injury is generally con-
sidered to account for the toxicity resulting from such exposure. It has
been found, for example, that the mean survival time in mice over the
dose range of 1000 to 12,000 r is three to four days and that early killing
occurs only if a large portion of the intestine is irradiated (Quastler et al,
1951). Exposure of the liver, kidney, spleen, and adrenal does not lead
to early death, at least in mice. This picture is probably different from
that observed after lower dosages, where several competing mechanisms
contribute to the lethal action. By exposing only the lower abdomen of
mice, thus irradiating a considerable portion of the small intestine while
sparing the liver and spleen, Chrom (1935) observed a reduction in the
enterogenous infection and greatly reduced toxicity with dosages of

PHYSIOLOGY OF RADIATION INJURY 983

550 to 1100 r. Effects on radiation morbidity and mortality have also
been demonstrated by Jacobson and his associates (Jacobson, Marks,
Robson, et al., 1949; Jacobson, Simmons, et al., 1950) with spleen shielding
of an otherwise totally irradiated mouse. Even in the median lethal
dosage range, however, mortality has been shown to be related in time
to a bacteremia of intestinal origin (Miller et al., 1950b), and it is perhaps
significant that maximal damage to the crypt epithelium precedes the
peak mortality of chronically irradiated mice (Bloom, 1950).

Qualitatively similar effects on the gastrointestinal tract have been
observed in a number of species after exposure to penetrating radiations
or to internally administered radioisotopes (Friedman, 1942; Desjardins,
1931a, b, c; Lawrence and Tennant, 1937; Pierce, 1948). The most
impressive changes occur in the epithelial cells, although the reactions
are not confined to them. Lesions have been noted during the first
hour after irradiation (Pierce, 1948; Tsuzuki, 1926; Friedman, 1945).
While it is generally agreed that the crypt epithelium is the most sensitive
site, destruction of the entire intestinal lining can occur with lethal dos-
ages, leaving fragmented crypt cells, denuded villi, edema, hemorrhage,
and ulcers in its wake. A marked accumulation of bacteria is often seen
on the surface of the intestinal mucosa. In contrast, nuclear injury in
the colon and in the surface epithelium of the stomach is usually slight,
although the gastric glands are rather easily damaged. The patho-
logical physiology does not always reflect morphological injury. The
occurrence of severe diarrhea, for example, may be unrelated temporally
to histological changes in the intestine (Shields Warren, MacMillan, and
Dixon, 1950a, b).

As might be anticipated, intestinal damage is influenced by the rate as
well as the intensity of exposure, injury of the crypt epithelium and the
production of ulcers being decreased with fractionation (Stafford L.
Warren and Whipple, 1923b; Engelstad, 1935, 1938). Indeed, some
radioresistance of the crypt epithelium may be acquired with suitably
spaced small exposures (Bloom, 1950). No doubt, much of the variance
in the early investigations can be attributed to differences in the physical
conditions of irradiation.

The causal relation between irradiation and morphological injury is
poorly understood. Nucleic acid, protein, and ash content of the crypt
epithelium is decreased soon after neutron and X irradiation (Ely and
Ross, 1948a, b; Ross and Ely, 1949a). Although the rates of synthesis
of DNA and RNA in rat intestine and in rabbit intestine are markedly
reduced by X rays, protein synthesis is relatively unaffected (Abrams,
1951). Alkaline phosphatase activity of the crypt cells is essentially
normal when assays are carried out at pH 7; phosphatase activity is
increased, however, at pH 9 (Ross and Ely, 1949b). It is not known
whether these chemical changes are the cause or effect of morphological

984 RADIATION BIOLOGY

injury and cell death. The same considerations may well apply to other
metabolic phenomena. Respiration of all segments of the small intestine
is inhibited several hours after exposure of the rat to 700 r (Barron et al.,
1947). Oxygen consumption is essentially normal at 24 hours, but is
again depressed on the third day. Anaerobic glycolysis is apparently
unaffected at all intervals.

A decrease in dry weight of the intestinal mucosa occurs during the
first two days after irradiation; this is not observed in nonirradiated
pair-fed controls (Ross and Ely, 1949a). In contrast, similar weight
losses have been detected in the gastrointestinal tract of X-rayed and
fasted rats at 4 days (Painter, 1948) . Although sodium space is increased
by some 50 per cent in both the irradiated and fasted animals, total water
content is not changed significantly. The increase in sodium space may
be related in part to an increase in extracellular fluid volume, and also to
an exchange of sodium for potassium in the intestinal cells (Painter and
Pullman, 1950). Radiosensitive tissues, in general, show an initial
decrease in sodium and potassium followed by an increase in the former
(Bowers and Scott, 1951a, b).

Anorexia and weight loss have been observed in a number of species and
are reasonably good indicators of the severity of radiation injury (Prosser,
Painter, Lisco, et ah, 1947; Ely and Ross, 1947). A transient diminution
in food intake has been noted by Smith, Tyree, Patt, and Bink (1951) in
the rat after only 50 r delivered to the whole body. After dosages of
250 to 10,000 r, food consumption drops to less than 10 per cent of normal
on the first day. Although the initial anorexia is rather similar over a
wide dosage range, recovery varies more or less directly with the magni-
tude of exposure, being more rapid after 250 r than after 500 r and non-
existent after 1000 r. With median lethal irradiation, food intake may
return toward normal after a few days, only to decrease again several
days before death. This is perhaps suggestive of some recovery in the
gastrointestinal tract during the intermediate period. Anorexia appar-
ently accounts for all or most of the weight loss that is seen after exposure
to penetrating radiation (Ely and Ross, 1947; Smith, Tyree, Patt, and
Bink, 1951). Observations made on a few rats after external 0 irradia-
tion reveal a decreased food intake for the first week or two, followed by an
increase above the normal that is maintained until death (Anderson,
1946). Body weight, however, falls progressively, perhaps as a result of
extensive fluid loss.

Anorexia, vomiting, and diarrhea are generally not observed in animals
after exposure of regions remote from the abdomen and are thought to be
a result of direct injury of the digestive tract. Symptoms of the same
general nature have been seen clinically after X irradiation of extra-
abdominal areas with dosages far below those employed experimentally.
The severity of the clinical reaction has been related to the size of the

PHYSIOLOGY OF RADIATION INJURY 985

capillary bed in the irradiated area (Jenkinson and Brown, 1944). Since
clinical irradiation is invariably confined to persons in ill health and since
the immediate clinical response has a large psychosomatic component, it
is not always easy to evaluate the results of such exposure. It is well
known that the clinical response is influenced by the nutritional status of
the person undergoing radiotherapy (Bean et al., 1944). The mechanisms
responsible for reactions in clinical radiation sickness and in the acute
radiation syndrome may be quite different.

The initial anorexia of whole-body irradiation need not be a conse-
quence of a direct action on the gastrointestinal tract, since exposure of
only the head of the rat results in a comparable early diminution in food
intake (Smith, Tyree, Patt, and Bink, 1951). This is not observed when
irradiation is confined to the extremities. Weight loss during the first
day or two is also rather similar in abdomen-shielded and abdomen-
exposed rats (Bond et al., 1950). These facts suggest that the early
anorexia may be neurogenic or humoral in origin. Delayed gastric
emptying (Ely and Ross, 1947; Mead et al., 1950), perhaps as a result of
increased tone of the pyloric sphincter, may be responsible for the
anorexia. Subsequent effects on food consumption, however, are prob-
ably related more directly to injury of the intestinal mucosa. When the
abdomen is shielded, late effects may possibly be attributed to damage of
oral and esophageal structures.

Intestinal motility and tonus are increased by irradiation (Hall and
Whipple, 1919; Martin and Rogers, 1923; Swann, 1924; Toyoma, 1933a;
Conard, 1951). This has been observed in intestinal loops and in the
intact animal. The increase in tonus and amplitude of contraction can
be detected during irradiation. With small dosages of X rays tonus
returns to normal a few minutes after the exposure is terminated. Large
dosages lead to a greater and more prolonged rise in tone, and a spastic
contraction, analogous to that seen after other forms of intestinal injury,
may be evident. Augmented contraction and hypertonicity are largely
prevented by parasympatholytic drugs and ganglionic blocking agents
(Conard, 1951). Vagotomy and body shielding afford only a slight
reduction of the intestinal response, which is considered to be a conse-
quence of direct action of radiation on cholinergic elements in the intes-
tinal tract. Increased synthesis of acetylcholine by brain (Torda and
Wolff, 1950) and a reduction in blood choline esterase (Barnard, 1948)
have been observed after exposure to X rays.

Irradiation of the stomach reduces gastric acidity and results in exten-
sive atrophy of the gastric glands (Miescher, 1923; Ivy et al., 1924; Ely
and Ross, 1947; Simon, 1949; Hedin et al., 1950; Douglas et al., 1950).
A transitory rise in acidity preceding the depression may occur after
exposure of the whole abdomen (Ivy et al., 1924). Secretory depression
has been observed with X rays, /3 rays, and neutrons and appears to be a

986 RADIATION BIOLOGY

local rather than a systemic effect of the radiation. Peptic activity of
the gastric juice is diminished but not to the same extent as the acidity
(Ivy et al, 1924; Ely and Ross, 1947). This is of interest, since histo-
logical observations reveal that the chief cells of the gastric mucosa are
more sensitive than the parietal cells, which are more intimately con-
cerned with acid secretion (Friedman, 1942; Hueper and de Carvajal-
Forero, 1944). Radiation has been employed clinically to reduce gastric
secretion in patients with ulcers; the advisability of such therapy may be
questioned, however, since it may itself lead to perforation and hemor-
rhage (Friedman and Warren, 1942; Brick, 1947; Ricketts et al, 1948).
In view of the extensive injury to the gastrointestinal tract following
irradiation of the abdomen, it is reasonable to assume that absorption
will be impaired. While decreased intestinal absorption has been
observed by several investigators (Martin and Rogers, 1923; Buchwald,
1931; Barron et al, 1947), others (Mead et al, 1950) believe that there is
no basic disturbance. The problem is complicated by changes in
emptying time of the stomach and in intestinal motility. Inhibition of
glucose absorption and diminished phosphorylation of fructose have been
observed as early as 4 hours after X irradiation (Barron et al, 1947).
The absorption of cream is also said to be diminished (Martin and
Rogers, 1923). Although an increase in fecal fat has been observed in
human beings treated with X rays (Dodds and Webster, 1924), similar
increases may be seen in animals on a fat-free diet (Mead et al, 1950).
There is little interference, moreover, with absorption of fat in irradiated
mice (Mead et al, 1950) and of vitamin A in irradiated rats (Bennett,
Bennett, et al, 1950) during the first postexposure week. There are
also indications that total-body irradiation does not disturb protein
absorption (Bennett et al, 1951). Enteral administration of protein
hydrolysates is apparently as efficacious as parenteral administration in
lowering the sensitivity of irradiated protein-depleted rats (Jennings,
unpublished observations, 1950). Mead and his associates (1950;
Bennett et al, 1951) believe that the mechanisms of absorption are
not impaired by irradiation and that the changes that have been noted
can be attributed in many instances to concomitant effects on motility.
The available data are largely restricted to the early period after
irradiation; little is known about gastrointestinal function in the inter-
mediate and terminal periods following acute exposure. Even less can
be said about the nutritional status of the irradiated organism and its
potential contribution to the radiation syndrome.

LIVER

It is assumed from morphological studies that the liver is relatively
resistant to radiation (Seldin, 1904; Smyth and Whipple, 1924; Pohle and

PHYSIOLOGY OF RADIATION INJURY 987

Bunting, 1932; Ely, Ross, and Gay, 1947; Rhoades, 1948a). This may
be attributed, in part, to the phenomenal regenerative capacity of liver
cells, and possibly to the low oxygen tension of the tissue. In an early
study, Seldin (1904) compared irradiated and shielded areas of the same
liver and was unable to detect any difference. The results of an extensive
investigation reported by Rhoades (1948a), in which the livers of rabbits,
rats, mice, and guinea pigs were examined after total-body X irradiation
with dosages of 25 to 1200 r confirm the radioresistance of liver epi-
thelium. Alterations, when observed, are considered to be secondary to
the general toxicity. When irradiation is accomplished with internal
emitters, histological change is also minor, with the notable exception of
plutonium, in which case hepatic injury is an outstanding feature of the
poisoning (Brues, 1948).

A nonspecific fatty infiltration is sometimes seen in the livers of irradi-
ated animals. The appearance of sudanophile fat in the mouse liver has
been attributed to the release of histamine-like tissue breakdown products
(Ellinger, 1945). Liver cholesterol has been shown to decrease and liver
glycogen to increase during the first two days after total-body irradiation
(North and Nims, 1949). These changes may reflect an increased
adrenal cortical activity. After local irradiation of the hepatic area of
the guinea pig, glycogen disappears somewhat more slowly from the liver
upon incubation (Ullmann, 1933). Inhibition of glycogen cleavage is
greater with 1200 r than with 600 r but is not well correlated with the
changes in total liver glycogen.

The possibility of metabolic disturbances in the liver that are not
manifested by morphological alterations is indicated by the decrease in
oxidative capacity of surviving liver slices obtained from animals exposed
to 7 and X rays or a rays from injected plutonium (Barron, 1946; DuBois,
unpublished observations, 1950). It is of interest that the oxidations
that are inhibited are those normally catalyzed by sulfhydryl enzymes.
However, it is noteworthy that other sulfhydryl enzymes in the liver are
not inhibited by large dosages of ionizing radiation. Thus there are no
significant changes in succinic dehydrogenase and adenosine triphos-
phatase when specific assays for these enzymes are made on livers taken
from mice exposed to y rays (DuBois, Cochran, and Doull, 1951a). The
activity and distribution of non-mercapto enzymes (catalase, alkaline
phosphatase, esterase, arginase, and rhodanase) in liver cells and in liver
connective tissue are unchanged following irradiation with 500 r (Ludewig
and Chanutin, 1950b). After lethal dosages, liver catalase activity is
decreased (Feinstein et al., 1950), while alkaline phosphatase is increased
(DuBois, unpublished observations, 1950).

Impairment of oxidative mechanisms, as well as possible changes in
phosphatase activity, may account for the decrease in acid-soluble
organic phosphorus, the increase in inorganic phosphorus, and the

988 RADIATION BIOLOGY

diminished turnover of phospholipid and desoxyribonucleic acid phos-
phorus in the livers of irradiated mice and rats (Hevesy, 1946, 1947;
Kelly and Jones, 1950; Thomson, unpublished observations, 1951). A
further indication of altered metabolic function as a result of irradiation
is the finding that citrate accumulates in the livers of irradiated male
rats after fluoroacetate treatment in contrast to the lack of effect of
fluoroacetate in the nonirradiated animal (DuBois, Cochran, and Doull,
1951a). This may be attributed to an effect of radiation on the testes,
since castration also leads to an increase in liver citrate that can be pre-
vented by testosterone (DuBois, Cochran, and Zerwic, 1951b). Citrate
accumulation with fluoroacetate occurs in the nonirradiated female.

Information relating to the effects of radiation on liver function is
nebulous and incomplete. Alterations in serum proteins produced by
radiation are not associated with significant changes in liver function as
indicated by the cephalin flocculation, colloidal gold, and thymol turbidity
tests (Schwartz et al.) 1948). Some decrease in the output of bile salts
has been seen when large dosages are delivered to the liver of the dog
(Smyth and Whipple, 1924). Terminal impairment of liver function
may be reflected by increased urinary excretion of urobilinogen and
coproporphyrin and the increased ratio of uric acid to allantoin (Schwartz
et al, 1948; Krizek et al., 1946; Miyazaki, 1937). Abnormal function
may also be indicated by the terminal rise in kynurenic acid excretion,
which is perhaps suggestive of some alteration in tryptophane metabolism
in the liver (Wattenberg and Schwartz, 1946a, b).

Another aspect of liver physiology is referable to its reticulo-endothelial
elements. Although there is evidence of a decrease in the capacity of
fixed liver phagocytes of irradiated mice to remove intravenously injected
bacteria (Chrom, 1935), the uptake of a gold colloid by the reticulo-
endothelial system is not impaired nor is there an appreciable difference in
the fate of heterologous erythrocytes in normal and X-rayed animals
(Wish et al, 1952; Barrow et al, 1949). There is, in fact, some evidence
of an increase in phagocytic activity after irradiation. Yet, injury of
liver phagocytes by radiation may be of some consequence since lethality
has been shown to be potentiated when rats are injected with minimal
lethal dosages of P32 and colloidal Au19S (Friedell and Christie, 1951).

KIDNEY

The kidney is another example of an organ that appears to be resistant
to radiation, at least as judged by morphological studies. According to
Shields Warren (1942), dosages in excess of several thousands of roentgens
are required for renal changes. Kidney damage is generally not apparent
in irradiated animals (Hall and Whipple, 1919; McQuarrie and Whipple,
1922; Impiombato, 1935; Ely et al, 1947; W. Bloom, 1948), and the usual

PHYSIOLOGY OF RADIATION INJURY 989

picture of intoxication follows heavy exposure even though the kidney is
shielded (McQuarrie and Whipple, 1922). An exception to the lack of
abnormalities in the kidney with moderate amounts of radiation is the
acute necrosis of the developing tubules and glomeruli of baby chicks
(W. Bloom, 1948; Steamer et al., 1951). The severe renal damage is
thought to predispose to early deaths in these animals (Steamer et al.,
1951).

There is some evidence of inhibition of kidney respiration and of oxida-
tion of substrates requiring sulfhydryl enzymes (Barron, 1946) ; this has
not been verified, however (DuBois, unpublished observations, 1950;
LeMay, 1951). Although extensive studies of glomerular and tubular
function have not been made, there is reason to believe that kidney
function is not impaired, except possibly in the agonal period following
acute irradiation of the whole body (Prosser, Painter, Lisco, et al., 1947).
In the dog, urinary excretion of water and nitrogen is not altered appreci-
ably, and the specific gravity of urine remains relatively constant.
Polyuria is seen in rats and rabbits during the first few postirradiation
days. Excretion of sodium, potassium, and chloride is increased in the
former on the first and fifth days (Edelmann, 1949). Increased urine
output is not always associated with polydipsia and may be a result, at
least in the rat, of an antidiuretic substance from the pituitary (Edelmann
and Eversole, 1950). Several days before death, water intake is generally
reduced in all species; urinary output is diminished less than water intake
(Prosser, Painter, Lisco, et at., 1947). Although an increased clearance of
phenol red has been observed in a few dogs during the second week
following median lethal dosages of X rays, a late diminished renal func-
tion is suggested by the marked terminal increase in blood nonprotein
nitrogen and in urea nitrogen. This is probably not a result of direct
radiation injury to the kidneys, but rather of the terminal circulatory
impairment and renal hemorrhage.

ENDOCRINES

Adrenal. It is recognized universally that the adrenal glands consti-
tute a buffer against a variety of traumatic conditions. The nonspecific-
ity of the adrenal response to stress and its role in the general adaptation
syndrome have been ably presented by Selye (1946) and elaborated upon
by others (Sayers, 1950). Ionizing radiations, in common with other
noxious stimuli, induce changes that are presumed to reflect an increased
demand for the adrenal hormones. In the absence of these secretions
susceptibility to irradiation is augmented. The functional response of
the adrenals does not appear to be a direct result of their irradiation.
Rather, it is mediated by the pituitary and closely resembles that seen
following a host of injuries. These considerations along with the appar-

990 RADIATION BIOLOGY

ent lack of specificity of radiation injury raise the question of the place
of the adrenal in the development of, and recovery from, the acute
radiation syndrome.

The suggestion that clinical radiation sickness reflects adrenal cortical
damage has been made by a number of investigators, who were either
impressed with the similarity between the symptoms of radiation sickness
and those of adrenal cortical insufficiency or with the efficacy of adrenal
preparations in alleviating the discomfort sometimes associated with
therapeutic irradiation (Narat, 1922; Hirsch, 1923; Thaddea, 1940;
Weichert, 1942; Ellinger, 1948). As early as 1924, however, Martin and
his associates were unable to detect functional changes in dogs from which
one adrenal was removed and the other irradiated with a dosage sufficient
to cause fibrotic changes. When the same dosage of X rays was applied
to an isolated loop of small intestine, cachexia and death resulted. Later
Fisher et al. (1928), employing a similar technique, observed signs of
adrenal insufficiency in a dog but only after heavy irradiation and a
latent period of about three months.

It appears, from morphological studies, that the adrenals are not
peculiarly radiosensitive (Frey, 1928; Desjardins, 1928; Engelstad, 1936;
Engelstad and Torgersen, 1937; Torgersen, 1940; Rhoades, 1948b).
Although degenerative changes have been seen after heavy local irradia-
tion (above 1500 r), dosages of 1000 r result only in minimal morpho-
logical alteration in the adrenal cortex and medulla (Engelstad, 1936;
Engelstad and Torgersen, 1937). Notwithstanding the apparent resist-
ance of the adrenals to structural change, functional responses may be
elicited with relatively low dosages. Loss of adrenal cortical lipids and
adrenal ascorbate occurs soon after irradiation, and urinary excretion of
the 17-ketosteroids may be increased several days later (Dougherty and
White, 1946; Patt et al, 1947; Nizet et al., 1949; North and Nims, 1949;
G. H. Lawrence, 1949). Lipid depletion has also been noted in some of
the victims of radiation exposure in Japan (Shields Warren, 1946). The
pattern of adrenal response in X-rayed rats consists of an initial reduction
in adrenal cholesterol, a normal or elevated cholesterol concentration
associated with adrenal hypertrophy several days later, and a marked
terminal depression (Patt et al., 1947). The adrenal response does not
occur in hypophysectomized animals and can be prevented, in part, by
suitable administration of adrenal cortical extract (Patt et al., 1948;
Swift et al., 1948). In contrast to the changes in adrenal lipids, oxidative
capacity of adrenal slices obtained from irradiated rats is not altered
appreciably (Barron, 1946).

The changes in adrenal lipids after irradiation probably reflect an
increased demand for cortical secretions, which may perhaps be satisfied
initially but not terminally. It is noteworthy that a single injection of
adrenocorticotrophic hormone decreases adrenal cholesterol in normal

PHYSIOLOGY OF RADIATION INJURY 991

animals, whereas cholesterol concentration increases after hormonal
stimulation of several days duration (Sayers et al., 1944). The rise in
adrenal cholesterol occurring several days after median lethal irradiation
may represent over-stimulation in excess of cortical hormone demand
(Patt et al., 1947). That the elevated cholesterol is not a result of
adrenal exhaustion is suggested by its absence with higher dosages.
Although adrenal lipids are usually depleted before death, it is not known
whether this is a cause or an effect of the terminal events. The possi-
bility exists that direct injury of the gland decreases its capacity to
respond to the stress of irradiation, since some protection has been
observed in rats when the adrenals are shielded during exposure (Craver,
1948; Edelmann, 1951a). Moderate degrees of stress (KC1 and hista-
mine) are well tolerated by the mouse after minimally lethal irradiation
(W. W. Smith, 1951).

There are a few indications of altered medullary activity. However,
the results are not particularly impressive. An initial discharge of
epinephrine may be anticipated with irradiation of large areas of the
body, but this has not been proved. Irradiation of the dog's adrenal
is said to increase the pressor action of blood collected from the adrenal
vein (Zunz and La Barre, 1927). On the other hand, significant changes
in adrenal catechols, which include epinephrine, have not been observed
after exposure of the rat adrenal to 100 or 1000 r (Raab and Soule, 1927).
Therapeutic irradiation apparently prevents increased output of medul-
lary hormone after exercise in patients with angina pectoris (Raab, 1941).
Reports of a salutary effect of adrenal irradiation in hypertension are
controversial (Torgersen, 1940).

The adrenals are implicated in some of the remote and indirect effects
of irradiation. Generalized involution of lymphoid structures has been
described following heavy local irradiation (Leblond and Segal, 1942;
Halberstaedter and Ickowicz, 1947). After adrenalectomy, the changes
are restricted to the irradiated area. The picture following total-body
exposure is somewhat obscure, although there is general agreement that
lymphoid involution after moderate dosages does not require the adrenal
or pituitary for its development (Dougherty and White, 1946; Patt et al.,
1948). Involution of lymphoid tissues and augmentation of antibody
titer subsequent to total-body exposure to 10 r have been attributed to
the adrenal (Dougherty and White, 1946; Dougherty et al, 1944); the
indirect action of a minimal lymphopenic dose and the "anamnestic
response" of antibody titer are not well substantiated, however (Crad-
dock and Lawrence, 1948; Marder et al., 1948).

The toxic reaction resulting from irradiation of large portions of the
body is potentiated in the absence of the adrenals or pituitary (Leblond
and Segal, 1942; Cronkite and Chapman, 1950; Edelmann, 1951b; Kaplan
et al., 1951; Patt et al., 1948). Susceptibility of adrenalectomized mice

992 RADIATION BIOLOGY

to the lethal action of X rays is especially striking with low dosages.
The degree of potentiation of radiation toxicity will, of course, depend
upon the nature of replacement therapy and the time intervening between
adrenalectomy and irradiation as well as the completeness of adrenal
extirpation. It is significant that survival of adrenalectomized mice is
similar to that of animals with intact adrenals when a constant dose of
adrenal cortical extract is given daily (Straube et al, 1949).

While there may be some rationale for employing adrenal corticoids to
reduce radiation mortality, for the most part the results of such endeavors
have been rather disappointing. Desoxycorticosterone has been shown
to prevent fatty infiltration of the liver and to improve survival in mice
(Ellinger, 1946, 1947). There are also some reports of its efficacy in
clinical radiation sickness (Weichert, 1942; Ellinger et al, 1949). These
observations have not been verified by others; a number of preparations,
including desoxycorticosterone, 11-dehydrocorticosterone, whole adrenal
cortical extract, and cortisone have been used experimentally without
success (Swift et al, 1948; Straube et al, 1949; Graham, Graham, and
Graffeo, 1950; Graham and Graham, 1950; W. W. Smith et al, 1950).
This is true also of the adrenocorticotrophic hormone (W. W. Smith et al,
1950). It is well to recall that negative results have been obtained with
adrenal extracts in other conditions of stress. It remains to be deter-
mined whether this obtains because of the difficulty in approximating
the quality and quantity of the naturally occurring internal secretions.

Thyroid. The normal thyroid is quite resistant to ionizing radiation in
contrast to the relative sensitivity of the hyperplastic gland. Heavy
irradiation, more than several thousand roentgens, is required to induce
structural changes in the thyroids of the guinea pig, rat, rabbit, and dog
(Shields Warren, 1943b; Bender, 1948). It has been found, for example,
that local exposure of the thyroid area of young or mature rats to 50 or
5000 r of X rays does not alter the histological appearance of the gland,
nor modify basal oxygen consumption or body weight (Bender, 1948).
Uptake of radioiodine is also unaffected by local irradiation with 1000 r
and is actually increased with 3000 and 6000 r (Hursh et al, 1949).
Increased uptake of I131 with the higher dosages occurs in the absence of
morphological changes and is regarded as an indirect result of irradiation.
The level of local X irradiation necessary to produce significant destruc-
tion of thyroid tissue in the rat is apparently greater than the highest
dosage that can be administered without a fatal outcome (Bender, 1948;
Hursh et al, 1949). Dosages in excess of 5000 r applied to the thyroid
area of the rat invariably result in death within about a week. The only
grossly abnormal feature is dehydration, which has been attributed to
failure to consume food and water for several days preceding death.
While death may be delayed by suitable administration of fluids, the
mechanism responsible for the lethal action is undetermined. It is

PHYSIOLOGY OF RADIATION INJURY 993

apparently not due to damage of the thyroid, esophagus, or pituitary,
nor is it related to changes in the blood. Complications have also been
observed clinically after excessive irradiation of the thyroid area (Lukens,
1948).

The avidity of the thyroid for iodine and the availability of radio-
iodine have greatly facilitated the study of thyroid sensitivity. The
results of such investigations reveal, in general, that relatively large
amounts of radioiodine as equivalent roentgens are required to embarrass
thyroid function (Findlay and Leblond, 1948; Feller et al, 1949; Skanse,
1948; Gorbman, 1949; Goldberg, Chaikoff, et al, 1950). Partial destruc-
tion of the thyroid with colloid degeneration has been seen in rats 6 days
after injection of approximately 70 /xc of I131, at which time about 20,000
rep had been delivered to the gland (Findlay and Leblond, 1948). Thy-
roid function as indicated by the chemical iodine content of the gland, its
retention of radioiodine, and the level of plasma iodine is not disturbed
after injection of 30 fxc of I131 in the rat (Feller et al, 1949). Weight of
the chick thyroid may be decreased 16 days after administration of only
10 /xc, but even this tracer dose is equivalent to approximately 15,000 rep
in terms of thyroid irradiation over the entire period (Skanse, 1948). In
the rat, large amounts of I131 may result in cytological changes in the
anterior pituitary that are identical with those seen after thyroid removal
(Goldberg and Chaikoff, 1950).

The thyroid apparently plays a negligible role in the total-body irradia-
tion syndrome. Radiation toxicity in the mammal is not altered appreci-
ably by thyroidectomy or thiouracil administration (Blount and Smith,
1949; Hempelmann et al, 1949; Haley et al, 1950). Toxicity may be
enhanced, however, when desiccated thyroid is given after irradiation
(W. W. Smith and Smith, 1951a). The potentiation of injury is prob-
ably related to elevated metabolism and is not specific for radiation
damage.

An increase in uptake of I131 by the thyroid has been noted after
irradiation of the whole body or the abdomen of rats (Evans et al, 1947).
The thyroid response occurs in the absence of the adrenals; it is not
observed in the newborn animal. While increased iodine uptake in the
adult animal is suggestive of an increase in thyroid function after irradia-
tion, the possibility exists that this may be related to anorexia and the
consequent decrease in dietary iodine. The fact that a definite increase in
the basal oxygen consumption of irradiated animals has not been demon-
strated argues against significant alteration of thyroid activity (Kirschner
et al, 1949; D. E. Smith, Tyree, Patt, and Jackson, 1951; W. W. Smith
and Smith, 1951b; Pratt et al, 1950; Patt, Swift, and Tyree, 1949).

Pituitary. It is generally agreed that the pituitary is only slightly
sensitive to X rays. The early literature has been reviewed by Shields
Warren (1943b), who concludes that the normal gland is radioresistant,

994 RADIATION BIOLOGY

although he is careful to point out that the response of the diseased gland
may he quite different. The relative resistance of the normal pituitary
to direct irradiation is also apparent from more recent studies. Single
dosages of 200 to 500 r applied to the pituitary region of adult female rats
do not lead to degenerative changes in the pituitary or in the ovaries,
adrenals, and thyroids (Kotz et al., 1941). A transient effect of X rays
on pituitary function is indicated by the reappearance of physiological
signs of heat when dosages ranging from 5 to 300 r are applied to the
pituitary of sexually mature female rats on the second day of their
estrous cycle (Freed et al., 1948). This reaction lasts for 6 to 8 hours and
is accompanied by a rather definite increase in pituitary weight and a
slight increase in uterine weight while ovarian weight is unchanged.
Comparable irradiation of immature female rats or ground squirrels does
not alter the development of their sexual systems (Freed et al., 1948;
Denniston, 1942). Although some increase in size and weight of the
pituitaries has been described in the infantile animal 11 to 14 weeks after
low-dose irradiation, there is no evidence of an increase in function
(Baidens et al., 1946). The early transient physiological effects of
pituitary irradiation are regarded by Freed et al. (1948) as secondary to
increased vascularity or altered permeability rather than as primary
stimulation of hypophyseal activity. While injurious effects are not
evident with small dosages of radiation, exposure of the pituitary region
to an air dose of 1000 r or more reduces the rate of growth of young rats
(Denniston, 1942). On the other hand, evidence of hypopituitarism is
not apparent in man with localized dosages of 8100 to 10,000 r (Kelly
etal., 1951).

The functional integrity of the pituitary during the first few days after
total-body irradiation is indicated by the typical pituitary-adrenal
cortical response to stress (Patt et al., 1947). Adrenal changes are not
seen in the irradiated hypophysectomized animal and there is some evi-
dence that pituitary ablation increases sensitivity to total-body irradia-
tion (Patt et al., 1948). It is not known whether the pituitary discharge,
which apparently takes place within several hours of exposure, is a
direct consequence of irradiation or of neural or humoral stimuli such as
epinephrine and tissue breakdown products. Many of these considera-
tions have already been discussed in connection with the effects of radia-
tion on the adrenal. It is worth recalling that the adrenocorticotrophic
hormone, in the one experimental study in which it was employed, did
not modify X-ray toxicity (W. W. Smith et al., 1950).

NERVOUS SYSTEM

The radioresistance of nervous tissue, indicated in the early experi-
ments of Kanoky (1907), has been confirmed in numerous investigations.

PHYSIOLOGY OF RADIATION INJURY 995

Gross dysfunction of the adult nervous system is not apparent following
total-body irradiation in the median lethal range (Prosser, Painter,
Lisco, et al., 1947), and there is no histological evidence of injury resulting
from such exposures (Snider, 1948). Although it is true that cytological
damage may occur following heavy irradiation, Shields Warren (1943a)
has pointed out that this can be largely indirect and perhaps attributed
to disturbance of the vascular system rather than to a direct effect of the
radiation on the cellular components of nervous tissue. This limiting
factor in the interpretation of radiation effects on the nervous system
has been emphasized by others (Campbell and Novick, 1949).

Early changes consisting of chromatolysis and vacuolization of the
ganglion cells have been observed following irradiation with several
thousand roentgens (Campbell et al., 1946; Novick, 1946). Changes in
the trigeminal ganglia of rabbits after local X irradiation of the head
with 3000 r can be separated temporally into three phases: an initially
severe chromatolysis immediately post-exposure, a recovery phase begin-
ning several hours later, and a second wave of chromatolysis beginning on
the third day and persisting for several weeks (Novick, 1946). The
clinical picture parallels the condition of the ganglion cells. Thus,
dyspnea, exopthalmos, somnolence, and postural abnormalities are evi-
dent upon termination of irradiation. These signs disappear rapidly
and are followed by a second phase of illness, characterized by emaciation
and excessive salivation during the delayed chromatolysis of the ganglion
cells. Delayed necrosis of brain cells, which appears several months
after exposure, has been described in rabbits and monkeys (Davidoff et al.,
1938; Russell et al., 1949). Localized necrosis in the central nervous
system of man, notably transverse myelitis, also may follow local irradia-
tion by 3000 r or more and may occur after months or years (Holmes and
Schulz, 1950). It is thought that vascular damage may be of importance
as an etiologic factor.

Local irradiation of the cerebral cortex of cats with small, high-energy
radon applicators results within a few hours in a sharp gradient of tissue
reactions. This technique has been used by Campbell and Novick
(1949) who have found that astrocytes are the most susceptible of the
various cells of the cerebral cortex; oligodendroglia and microglia are
highly radioresistant. Large dosages of X rays applied to one side of
the cerebrum of monkeys result in paralysis of the contralateral limbs
(Davidoff et al., 1938). Hemiplegia occurs rapidly with 4000 r and after
several months with 2000 to 3000 r. Massive irradiation of the mid-
thoracic spinal cord likewise results in a paralysis. Cerebellar lesions
may appear several months after irradiation of the cerebrum.

Irradiation of the whole body of the mouse with dosages in excess of
6000 r results in a hyperacute reaction that is characterized by motor
excitability (Quastler et al., 1951). Motor symptoms become more con-

996 RADIATION BIOLOGY

spicuous and death intervenes within a day or two after exposure to
12,000 r. These effects have been attributed to irradiation of the brain,
since they do not occur when the head is shielded. The application of
50,000 r to the entire body of guinea pigs, rabbits, and mice may lead to
death under the beam or within a few hours; hyperthermia, hyperesthesia,
intermittent seizures, and cyanosis may be noted in these animals
(Henshaw, 1944). Early death after massive irradiation occurs even
though the head is shielded (Quastler et al., 1951).

Peripheral nerve appears to be more resistant than brain and spinal
cord. The sciatic nerve of the rat is apparently unaffected by X-ray
dosages of 4000 to 10,000 r (Janzen and Warren, 1942). To accomplish
complete degeneration of the nerve, approximately 75,000 r of y radiation
is required. There is some evidence that daily exposures to 80 r may
interfere with regeneration of the hemisected sciatic, although there is
no appreciable effect on the Schwann cells with such" treatment (Gastaldi,

1949).

Synthesis of acetylcholine by brain is enhanced after sublethal X
irradiation (Torda and Wolff, 1950). This may be a result, in part, of
thymic involution and a consequent decrease in concentration of the
choline acetylase inhibitors ordinarily present in this tissue. The
capacity of peripheral nerve of hypophysectomized rats to maintain the
action potential of muscle during repetitive stimulation is partially
restored by low-dose irradiation (Torda and Wolff, 1950). Increased
synthesis of acetylcholine may bear some relation to the parasympatho-
mimetic effects that are evident during the initial phases of radiation
sickness. It is noteworthy that atropine minimizes certain early radia-
tion effects, including the hypotension in rabbits (Painter et al, 1947) and
the hypertonicity of intestinal loops (Conard, 1951); treatment with
atropine has also been shown to improve slightly the survival of irradiated
mice (Larkin, 1949).

The radioresistance of the adult nervous system stands in sharp con-
trast to the sensitivity of developing nervous tissue. The sensitivity of
developing neuroblasts in mouse and rat embryos during the latter two-
thirds of pregnancy has been reported by Hicks (1950). Irradiation of
the pregnant animal with 150 to 200 r results in extensive destruction of
the embryonic neuroblasts and severe malformations of the brain.
Extraneural lesions do not appear with this dosage. Susceptibility of the
embryonic nervous system to radiation injury has also been demon-
strated following irradiation of selected implantation sites in pregnant
rats without exposure of the mother (Wilson and Karr, 1950). The fac-
tors that act to influence responsiveness of tissue under various conditions
of growth and differentiation are unknown. It is not possible at present
to explain the relative sensitivities of developing and adult nerve cells
other than to implicate metabolic differences that exist between the two
(see also Chap. 13).

PHYSIOLOGY OF RADIATION INJURY 997

MISCELLANEOUS TISSUES AND ORGANS

The relative radio-sensitivities of various tissues and detailed histologic
information on radiation-induced changes within them are discussed by
Bloom and Bloom in Chap. 17. In the ensuing discussion, we shall men-
tion briefly certain physiologically important effects of ionizing radiation
on some of these tissues and organs.

Skin. Radiosensitivity of the skin constitutes an important factor in
radiation therapy, mainly owing to the fact that it often imposes the
chief limitation on deep therapy. Since erythema is the first visible sign
of radiation effects on skin and runs parallel to later and more serious
effects, much attention has been given to it in clinical practice. Its
vascular basis and usual course have been discussed already (p. 976).
Large dosages result after a few weeks in denudation of the epithelium
(moist epidermititis) (Regaud and Nogier, 1913). Treatment of the
entire surface of the bodies of mice by /3 rays from P32-containing plaques
reproduces this state over large areas where the dosage is calculated as
4000 rep, beginning in four weeks and continuing for some months there-
after. New epidermis is continually formed under the sloughs through-
out this period (Raper and Barnes, 1951b). These animals show a gradu-
ally increasing mortality during the years after exposure. With higher
dosages, survival becomes much shorter, and above 5000 rep most of the
deaths occur within three weeks.

It has been found in animals of different size (baby rats, mice, adult
rats, guinea pigs, and rabbits) that the lethal dose, expressed as the
45-day LD5o, increases with size from 2200 rep to 17,500 rep. These
dosages correspond to equivalent proportional volume dosages (gram-rep
per gram) for the several species, which are fairly comparable with the
respective lethal volume dosages of 7 rays (Raper, Zirkle, and Barnes,
1951). The reason for this correspondence is not clear; it is obvious that
external 0 irradiation and penetrating ionizing radiations act through
different mechanisms, since they are incompletely additive (Raper and
Barnes, 1951c) and since lethal external /? irradiation fails to produce
leukopenia (Raper and Barnes, 1951a). It seems probable that certain
of the mechanisms of death are the same as those following thermal burns,
although the changes in skin responsible for them are slower in developing.

Hair. Temporary epilation is produced in man by dosages of radiation
of the order of 375 to 500 roentgens (Pendergrass and Mahoney, 1948).
The basis for epilation is presumably inhibition of growth in the hair
follicles. Permanent epilation requires a considerably higher dosage.
Graying of hair is seen in mice following relatively low doses and is nearly
complete in certain areas at 1000 r (Chase, 1949). Although temporary
epilation is widely produced in the course of therapeutic irradiation,
graying is seen rarely after regrowth takes place, and it is therefore clear
that the human threshold for graying is considerably higher than that for

998 RADIATION BIOLOGY

some other species. One human case has recently been described in which
the equivalent of 390 r of soft X rays resulted in temporary epilation with
subsequent regrowth of normally pigmented hair (Hempelmann et al.,
1952). The basis of graying, which is permanent once it occurs, is prob-
ably destruction of melanoblasts; graying in mice is the characteristic
effect of radiation on the inactive (nongrowing) stage, while active
follicles respond by epilation. It has been suggested (Chase, 1949)
that the increased melanoblast population of follicles in the active stage
acts to protect these follicles against loss of potentiality to produce
pigment.

Eye. Superficial effects of irradiation on the cornea and conjunctiva
run parallel to effects on the skin, as regards dosage and the cyclical
nature of the response. It is of interest that a sensation of pain and
heat may be felt during irradiation with a few hundred roentgen equiv-
alents (Robbins et al., 1946). Many of the ocular lesions following total-
body irradiation are obviously the result of the systemic changes occur-
ring in the radiation syndrome (Wilder and Maynard, 1951).

Lenticular cataracts are among the most serious nonfatal consequences
of irradiation. Although this has long been known, it received particular
emphasis in 1948 when several cases were discovered in physicists who
had been exposed to cyclotron neutrons (Abelson and Kruger, 1949) and
when survivors of atomic bomb irradiation began to develop cataracts
several years after the exposure (Cogan et al., 1949). The threshold for
clinically serious cataract formation by X rays in adult animals is prob-
ably close to the acute total-body LD50, but is relatively much lower
following exposure to neutrons, so that this effect is seen more often in
survivors of total-body neutron exposure. Young animals are more
susceptible than adults, and the latent period is related inversely to dose
(Leinf elder and Kerr, 1936). Intermittent dosage with fast neutrons
results in a particularly high ratio of effectiveness between neutrons and
7 rays where cataracts are used as the criterion (Evans, 1948) . The radia-
tion cataract is seen typically in the posterior capsular area, but a cyto-
logical analysis of the course of cataract formation shows that an impor-
tant component in its pathogenesis is the destruction of epithelial cells
that later migrate to this area (Cogan and Donaldson, 1951).

Gonads. For a detailed account of radiation changes in the gonads the
reader is referred to Chap. 17 by Bloom and Bloom. The physiologic
responses resulting from irradiation of the testes and ovaries include
temporary or permanent sterility, certain endocrine responses, and, in
the case of the testis, evidence of impaired viability or diminished
numbers of sperm.

Evidence of temporary sterility is first seen in the mouse about three
weeks after a single irradiation (Glucksmann, 1947). Examination of
the time course of survival of various cell types indicates that spermato-

PHYSIOLOGY OF RADIATION INJURY 999

gonia are most sensitive, thus accounting for the delay between irradia-
tion and the diminution in sperm count (Eschenbrenner and Miller,
1950; Fogg and Cowing, 1951). Prior to the development of oligo-
spermia, however, matings by the irradiated male result in a smaller
than normal litter size, which may be explained by chromosome trans-
locations in the sperm cells (Brenneke, 1937; Snell, 1935). This decrease
in litter size and associated evidence of arrested segmentation of fertil-
ized ova (Parkes, 1948) appear in the dosage range between 250 and 500 r.
Local and total-body irradiation yield similar findings in the testes (Fogg
and Cowing, 1951). At lower dosages, recovery is essentially complete,
while no recovery occurs in mice subjected to testicular irradiation at
2000 r. After 5000 r the interstitial cells remain intact (Fogg and Cow-
ing, 1952a). The human being may be considerably more sensitive than
rodents in terms of temporary or permanent sterility (Glucksmann, 1947).
Sterility induced by 2000 r in mice and 4500 r in rats does not result in
feminizing changes in the submaxillary gland, which are a characteristic
consequence of castration (Fogg and Cowing, 1952b). " Castration cells "
do, however, appear in the pituitary after sterilizing irradiation (Liebow,
Warren, and DeCoursey, 1949).

The situation regarding physiological effects of irradiating the ovaries
seems to be somewhat more complicated. While temporary sterility
may be produced by 170 r to the human ovaries (Glucksmann, 1947;
Martin, 1950) and permanent sterility by less than twice this dose, it has
nevertheless been customary to treat sterility with similar dosages, and
pregnancies have been reported in 35 per cent of a large series of women,
believed to be sterile, after X irradiation of the ovaries with about 175 r
and of the pituitary with 225 r (Kaplan, 1949). The ovarian dosages
required to produce permanent amenorrhea vary within wide limits
(Liebow, Warren, and DeCoursey, 1949). In evaluating the effect of
radiation on the human ovary, it is well to remember that amenorrhea
does not necessarily imply inability to conceive. Mice show a temporary
reduction in the frequency of estrus after 200 r and temporary abolition
of estrus after 400 r (Bischoff et al., 1944). It would appear that second-
ary sex characters are more sensitive to gonadal irradiation in the female
than in the male (Bloom and Bloom, Chap. 17). Consideration of the
circumstances involved in the production of ovarian tumors in mice after
local X irradiation with 200 r suggests that this dosage, if delivered to all
of the ovarian tissue, results in a gonadotrophs response by the pituitary
(Lick, Kirschbaum, and Mixer, 1949).

Bone and Cartilage. Responses of these tissues have been described
thoroughly by Bloom and Bloom (Chap. 17). Retardation of bone
growth following moderate irradiation of the growing areas is a matter of
some concern to radiotherapists treating younger individuals, since
recovery is likely to be incomplete following 1000 or 2000 r or more.

1000 RADIATION BIOLOGY

Long-continued irradiation of adult bones by radium deposited in the
skeleton results in the appearance of areas of rarefaction and aseptic
bone necrosis, appearing late and progressing during the course of at
least 20 to 25 years. Necrosis is especially likely to occur in the jaw,
and the bone changes are precancerous (Martland, 1931).

Although cartilage is not remarkably radiosensitive, its recovery after
irradiation to the point where necrosis occurs is very poor, perhaps owing
to vascular damage (Kaplan, 1949). Consequently, therapeutic irradia-
tion of cartilaginous areas, notably the ear and trachea, must be carried
out with great caution.

METABOLISM AND TISSUE BREAKDOWN

Metabolic changes may result from the direct action of radiation on
enzyme systems and other biochemical mechanisms, altered gastro-
intestinal, hormonal, and renal function, and the products of tissue
breakdown. Certain of the metabolic consequences of exposure to
X rays were described in 1907 by Edsall and Pemberton, who interpreted
clinical radiation sickness as a toxic reaction to tissue breakdown. Intox-
ication was attributed to the inability of the organism to metabolize and
excrete the products of cellular disintegration. Subsequently, Doub
et al. (1925) observed a hyperphosphatemia that was related to radiation
dosage, and noted, in addition, that irradiation of large tumor masses
resulted in an early alkalosis and elevated blood levels of nonprotein
nitrogen, uric acid, and guanidine. Arguments in favor of the tissue
toxin hypothesis have been advanced by a number of investigators since
the early work of Edsall and Pemberton (Hall and Whipple, 1919;
Stafford L. Warren and Whipple, 1923d; Rolleston, 1930; Forfota and
Karady, 1937; Moon et al., 1941). It is quite possible that tissue break-
down products contribute to the early shock reactions with massive
irradiation. Their role with smaller dosages is not well defined, although
there is some evidence that circulating factors may be involved in the
initial toxicity resulting from such exposures (Barnes and Furth, 1943;
Painter et al., 1947 ; Weber and Steggerda, 1949 ; Ellinger, 1951). Toxins
may, of course, be expected in the presence of the bacteremia that occurs

later.

Metabolic changes attributable to protein breakdown are also apparent
from some of the more recent studies with deep roentgen therapy in man
and with experimental irradiation of animals (Goldman, 1943; Robertson,
1943; Prosser et al., 1947a, b). Urinary nitrogen is increased during the
first hours after irradiation, and specific organ proteinases have been
detected in the urine of the dog (Oster and Salter, 1938; Abderhalden,
1939). The subsequent excretion of nitrogen is usually maintained
within normal limits until shortly before death. Since intake is reduced

PHYSIOLOGY OF RADIATION INJURY 1001

there is a negative nitrogen balance (Prosser, Painter, Lisco, et al., 1947;
Prosser, Painter, and Swift, 1947). These findings are suggestive of a
shift in favor of catabolism.

Organ weights provide a reasonable index of tissue breakdown and
repair after irradiation. The most pronounced changes are seen in the
lymphoid organs and testes (Carter, Harris, and Brennan, 1950; Eschen-
brenner and Miller, 1950). The weight of the gastrointestinal tract is
also decreased (Painter, 1948; Ross and Ely, 1949a) as is the volume of
available marrow (Brecher et al., 1948). Muscle and kidney, on the
other hand, undergo little change, while the adrenals are increased in
size (Patt et al., 1947). Changes in thymus, spleen, and testis are useful
as biological dosimeters especially after low dosages; the differences in
organ weights are relatively slight, however, over the narrow range from
just lethal to completely lethal.

The finding that weight loss of the irradiated dog cannot be accounted
for by decreased food intake (Prosser, Painter, and Swift, 1947) is sug-
gestive of an increase in catabolism. In the rat, however, change in
body weight appears to be related directly to anorexia (Ely and Ross,
1947; D. E. Smith, Tyree, Patt, and Bink, 1951). Identical weight
losses have been observed, moreover, in starved and in starved-irradiated
rats (D. E. Smith, Tyree, Patt, and Jackson, 1951). Since lethal irradia-
tion does not influence the water content of muscle or the entire carcass of
the starved rat, it is apparent that water retention cannot be responsible
for the failure to find a greater depression of body weight in the starved-
irradiated animal. Extracellular water may be increased after lethal
irradiation, but the changes in total extracellular space are similar to
those seen in starvation (Painter, 1948). Tissue breakdown associated
with irradiation, as with starvation, does not result invariably in elevated
metabolism. Although direct evidence of an increase in the metabolic
rate of irradiated rats has been presented (Kirschner et al., 1949), changes
in oxygen consumption do not coincide with weight loss. Other investi-
gators have been unable to verify the increase in metabolic rate in mice,
rats, guinea pigs, or frogs (D. E. Smith, Tyree, Patt, and Jackson, 1951;
W. W. Smith and Smith, 1951b; Pratt et al, 1950; Patt, Swift, and
Tyree, 1949) . Measurements of oxygen consumption have not been made
during the terminal febrile period in dogs.

Oxygen utilization may actually be depressed, at least in certain tissues,
since the irradiated mouse is said to be more resistant to progressive
asphyxia than the nonirradiated (W. W. Smith and Smith, 1951b). It is
of interest that the respiratory rate of tissues obtained from irradiated
animals or of tissues irradiated in vitro is either unchanged or decreased
(Wels, 1924; Crabtree, 1935; Goldfeder and Fershing, 1938; Barron,
1946; Barron et al., 1947; DuBois, unpublished observations, 1950). An
exception to this is the brief increase in respiration that has been seen in

1002 RADIATION BIOLOGY

rabbit marrow homogenates immediately after X irradiation (Altman
et al, 1951) and in fowl erythrocytes after massive exposure (Frankenthal
and Back, 1944). While depressed tissue respiration can perhaps be
attributed to specific inactivation of oxidative enzymes by the radiation,
gross metabolic changes seem to be the result and not the cause of growth
inhibition. In fact, effects on growth may occur at dosages considerably
below those required to influence metabolism (Hubert, 1929; Fenn and
Latchford, 1931-1932 ; Packard, 1933) . The absence of profound changes
in the metabolic rate of the whole animal in the presence of possible
alteration in endogenous respiration of radiosensitive tissues is not
unreasonable, since tissues that account for most of the oxygen consump-
tion are notably radioresistant. Tissue breakdown products resulting
from irradiation apparently do not result in an appreciable over-all
increase in metabolism, perhaps because of their rapid excretion or slow
release, the associated inanition, or other as yet unknown factors.

IMMUNITY AND INFECTION

It is well known that total-body irradiation decreases resistance to
infection and interferes with immune reactions. The action of ionizing
radiations on barriers to infection attracted early attention when it was
noted that irradiation resulted in a severe depression of leukocytes
(Heineke, 1903), inhibition of antibody formation (Benjamin and Sluka,
1908; Hektoen, 1915), and a terminal bacteremia of intestinal origin
(Stafford L. Warren and Whipple, 1923a; Mottram and Kingsbury, 1924).
The development of enterogenous infection in X-irradiated mice, includ-
ing some histological evidence of local bacterial invasion of injured
intestinal areas, was described subsequently by Chrom (1935), who noted
also that bacteremia could be minimized by shielding the liver and spleen
during irradiation. The complication of bacteremia was also observed
after neutron irradiation (Lawrence and Tennant, 1937). These findings
take on added significance in view of the efficacy of antibiotic therapy in
certain species (Miller et al, 1950b; Hammond and Miller, 1950; Howland
et al, 1950; Furth, Coulter, and Howland, 1951; Koletsky and Christie,
1950; Gustafson and Koletsky, 1951) and the effect of spleen shielding
on hematopoietic recovery and radiation mortality (Jacobson, Marks,
and Lorenz, 1949; Jacobson, Simmons, et al, 1950).

The problem of radiation-induced infection has been the subject of
intensive investigation in recent years, and it now seems clear that
enterogenous invasion can be a substantial factor in radiation morbidity
and mortality. The cause of the bacteremia still remains a moot ques-
tion, however. Miller and co-workers (1950a) have observed that bac-
teremia in irradiated mice reaches its apogee during the period of greatest
mortality. A somewhat lower incidence of positive blood cultures has

PHYSIOLOGY OF RADIATION INJURY 1003

been seen in the dog, but even in this species infection appears to be
related to mortality in animals surviving the first two weeks of irradiation
(Bennett, Rekers, and Howland, 1950). Total-body X irradiation of
guinea pigs has also been shown to result in a generalized tissue invasion
by cholera vibrio from the original focus of infection in the bowel (Bur-
rows et al., 1950a). The intensity of enteric infection appears to be
related to a decrease in coproantibody titer. It has been suggested by
Burrows and his associates (1950a) that a similar decrease in titer may
account for bacteremia in irradiated animals if natural immunity to
intestinal organisms is related to the coproantibody response.

Mortality of irradiated animals may be reduced with suitable anti-
biotic therapy. Streptomycin and terramycin afford the most effective
protection to mice (Miller et al., 1950b; Hammond and Miller, 1950).
The incidence of diarrhea in rats is greatly diminished by terramycin or
aureomycin; survival may also be prolonged (Howland et al., 1950;
Furth, Coulter, and Howland, 1951). Mortality from internal radiation
(P32) is decreased in this species when streptomycin and penicillin are
given in combination (Koletsky and Christie, 1950). Definite effects on
survival have not been observed in irradiated dogs with aureomycin
(Furth and Coulter, 1950; Allen et al, 1951). It is of interest that
prophylaxis with terramycin for 2 to 3 days prior to exposure reduces
radiation mortality in rats, presumably by altering the intestinal flora
and consequent bacteremia (Gustafson and Koletsky, 1951). In general,
antibiotic therapy leads to a reduction in the number of positive blood
cultures; pancytopenia is usually not altered, however, although gastro-
intestinal hemorrhage and ulceration may be diminished.

Numerous reports of the effects of radiation on immunity have
appeared since the detailed observations by Hektoen in 1915 of altered
antibody formation following exposure to X rays. Decreased resistance
of irradiated animals to infection had actually been demonstrated some
years previously by Lawen (1909). Although there are some indications
of a protective or beneficial action with small dosages of radiation (Glenn,
1946), the evidence, for the most part, points to an impairment of
immunological responses and to an increased susceptibility to infection
as a result of irradiation. X irradiation decreases the Shwartzman
phenomenon of local tissue reactivity to intradermally injected bacterial
endotoxins (Becker, 1948) and also diminishes the renal lesions that occur
in rabbits after injection of bovine serum y- globulin (Schwab et al., 1950).
These effects may be related to suppression of antibody formation. The
extensive literature in this field has been the subject of recent reviews by
Taliaferro and Taliaferro (1951) and Craddock and Lawrence (1948).

A marked reduction in the capacity to form antibodies has been demon-
strated when irradiation is accomplished shortly before or after immuniza-
tion (Hektoen, 1915; Murphy and Sturm, 1925; Craddock and Lawrence,

1004 RADIATION BIOLOGY

1948; Clemmesen and Andersen, 1948; Schwab et al, 1950; Burrows et al.,
1950b; Kohn, 1951c). Some depression of antibody titer has been noted
when immunization precedes irradiation, but a given dose of X rays
becomes progressively less effective as the time of its administration is
delayed after the injection of antigen (Kohn, 1951c). The excretion of
fecal and urinary antibodies may be inhibited under these conditions
even though a deleterious effect on the serum titer is not apparent (Bur-
rows et al., 1950b). A transitory increase in the excretion of fecal and
urinary antibody has been observed in the guinea pig when immunization
with cholera O vaccine is begun 1 day, but not 3 days, after irradiation
(Burrows et al., 1950b). The apparently contradictory findings of
beneficial and detrimental effects of irradiation on infection conceivably
may be explained on this basis. Sublethal irradiation does not appear
to alter the rate of destruction of passively transferred homologous or
heterologous antisera (Hollingsworth, 1950; Borowskaja, 1946). The
capacity of the irradiated rabbit to produce antibodies to injected antigen
is largely retained if the spleen or appendix is shielded during exposure
(Jacobson, Robson, and Marks, 1950). It is not known whether the
shielded lymphatic tissue initiates antibody formation or makes it possible
for the process to be initiated elsewhere.

Augmentation of antibody titer has been observed in immunized mice
after irradiation with 10 to 100 r (Dougherty et al., 1944). The "anam-
nestic reaction" does not occur in adrenalectomized mice and is thought
to be related to adrenal cortical stimulation, which results in lymphocyte
dissolution and release of 7-globulin. The anamnestic response of anti-
body titer has not been seen in rabbits after comparable low-dose irradia-
tion (Craddock and Lawrence, 1948). Local irradiation of the hind legs
of sensitized rabbits also fails to increase the amount of circulating anti-
bodies (Lecomte and Fischer, 1949). There are other indications that
the lymphocytes do not serve as a store of antibodies, since fairly large
dosages of X radiation do not result in an appreciable change in the
antibody concentration of lymph, despite the widespread destruction of
lymphatic tissue (Craddock et al., 1949). There is no clear indication
moreover of a relation between the height of antibody titer in serum and
lymph. The role of the adrenal cortex in the production and release of
antibody and 7-globulin has been questioned by others (Eisen et al.,
1947;Fischele£aZ., 1949).

NONSPECIFIC PHYSIOLOGICAL STRESSES

In response to stress, there are evoked compensatory or buffer mecha-
nisms whose chief function is to maintain vital processes by minimizing
injury and promoting repair. The pituitary-adrenal and reticulo-
endothelial systems are concerned intimately with these phenomena.

PHYSIOLOGY OF RADIATION INJURY 1005

Obviously, the extent to which adaptive and defensive mechanisms are
successful will depend, for the most part, upon the magnitude of the
stress. The nonspecific potentiation of one stress by another may be
interpreted on this basis. These considerations apply to the high-energy
radiations, which, because of their distribution, usually result in* fairly
extensive injury. It has been shown, for example, that hypophysectomy
(Patt et al., 1948), adrenal insufficiency (Cronkite and Chapman, 1950;
Edelmann, 1951b; Kaplan et al., 1951), infection (Taliaferro el al., 1945;
Shechmeister and Bond, 1951 ; Shechmeister el al., 1950), trauma (Brooks,
1951), exhaustive exercise (Stapleton and Curtis, 1946; Kimeldorf
et al., 1950), and nutritional deficiencies (Johnson et al., 1946; Jennings,
1949) enhance the susceptibility to irradiation and, conversely, that
irradiation increases the toxicity resulting from many of these conditions.
Moderate degrees of stress, however, may be well tolerated by minimally
irradiated animals (W. W. Smith, 1951; W. W. Smith and Smith, 1951b;
F. Smith and Smith, 1951). Of interest from the point of view of war
disaster, is the recent finding by Brooks (1951) that 100 r total-body X
irradiation, which by itself is nonlethal, increases the mortality of dogs
from 14 to 75 per cent following a standardized surface burn.

ACUTE RADIATION SYNDROME

The symptom complex appearing within several weeks after irradiation
of large areas of the body and the way it develops may be considered
pathognomonic of radiation exposure (Shields Warren and Bowers,
1950; Howland and Warren, 1947; Jacobson et al., 1949d; Painter and
Brues, 1949; Hempelmann, 1950; Shields Warren and Brues, 1950;
Bowers, 1951; Cronkite, 1951; Hempelmann et al., 1952). The relatively
mild discomfort of clinical radiation sickness, which is sometimes seen
after therapeutic exposure, should be distinguished from the more serious
and complex events of the acute radiation syndrome, of which it is a part.
It is well to recall that chronic effects, e.g., anemia, neoplasia, lenticular
opacities, and "premature aging"2 may appear in survivors of the acute
injury, or after protracted or repeated irradiation. The more chronic
sequelae do not, however, constitute as discrete an entity as the acute
syndrome. The pattern in man is, in general, similar to that observed
experimentally. Our knowledge of acute radiation injury in man is
based largely upon studies of the Japanese bomb casualties (Howland
and Warren, 1947; Liebow et al., 1949) and of persons injured by acci-
dental nuclear reactions (Hempelmann et al., 1952). Data relating to
the Japanese victims are necessarily incomplete and complicated by the

2 Shortening of life span is generally thought of as if it involved an acceleration of the
aging process. In fact, it rests almost entirely on statistical data and so is analogous
to semilethal effects in Drosophila (Brues and Sacher, 1952).

1006 RADIATION BIOLOGY

effects of heat and blast and the lack of precise information regarding
conditions of exposure. The accident cases, on the other hand, represent
uncomplicated radiation injury.

The disturbance consequent to irradiation assumes a distinctive time
course, which depends not only upon the dosage, duration, and manner
of irradiation but also upon the temporal relation between injury and
recovery of particular cells and their rate of utilization by the organism.
The initial damage leads to a complex chain of events, some of which
are a direct result of the primary injury while others are secondary to it.
Alteration in the cellular constituents of peripheral blood is an example of
the former; in fluid balance, of the latter. All the changes are dis-
tinguished by their lack of specificity.

Supralethal irradiation is followed by a shocklike reaction and death
within a few days. Severe intestinal damage and central nervous system
disturbances are prominent sequelae of massive irradiation. After
median lethal irradiation, there is an initially mild disturbance, a brief
period of apparent respite, and a final phase of progressive injury with
death or recovery. The waves of mortality that occur after moderate
irradiation suggest that multiple factors may be involved. These have
not been defined satisfactorily. It is apparent, however, that leukopenia,
septicemia, hemorrhage, and gastrointestinal damage constitute the most
important insults to the irradiated organism.

Medical management of the radiation syndrome may be resolved into
three basic components: (1) correction of the panhematopenia, (2) pre-
vention or treatment of infection, and (3) maintenance of adequate nutri-
tion and fluid balance. Although irreversible tissue injuries occur soon
after intensive irradiation, there are indications that it may be possible to
prevent effectively certain of the radiation changes and to facilitate the
regenerative capacity of critical tissues. It is noteworthy, for example,
that morbidity and mortality are influenced by procedures that promote
recovery of blood-forming tissues or prevent bacteremia, e.g., spleen and
marrow transplants, and antibiotics. Data pertaining to biological pro-
tection against the acute syndrome by means of organ shielding are most
impressive and should encourage further attempts to influence tissue
recovery with humoral agents (cf. Chap. 16). From a practical point of
view, it is imperative to establish the role of blood transfusion, hemato-
poietic and antihemorrhagic factors, and parenteral feeding. Early
venesection and blood replacement have been shown to reduce radiation
mortality in dogs; periodic blood transfusions, on the other hand, mini-
mize anemia but apparently do not influence other radiation sequelae or
mortality.

Specific chemical protection against many of the effects of ionizing
radiation is of considerable theoretical importance. Elucidation of these
protective mechanisms may reveal the nature of the early chemical

PHYSIOLOGY OF RADIATION INJURY

1007

1008 RADIATION BIOLOGY

effects. The products of irradiated water, mainly oxidants, are probably
responsible for a number of radiobiological responses; oxygen deprivation
during exposure and certain reductants such as cysteine have been shown
to diminish acute toxicity. Organic molecules are believed to be altered,
e.g., oxidized, reduced, depolymerized, or denatured, by direct ionization
and excitation as well as by interaction with the products of irradiated
water. While this is in accord with effects on simple systems, the nature
of the biochemical targets is poorly understood. It is assumed that the
early events involve molecules that are concerned with regulatory
mechanisms, e.g., enzymes or genes, as well as molecules whose integrity
ensures the structural stability of the nuclear material and the ability of
the cell to divide normally. Cytological damage, the earliest recognizable
effect, apparently leads to deficiencies that are manifest first in areas of
rapid cell turnover, the most critical of which are the hematopoietic and
intestinal tissues. The essential features of the acute radiation syndrome
presented diagrammatically in Fig. 15-1, can be largely explained on this
basis.

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1010 RADIATION BIOLOGY

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PHYSIOLOGY OF RADIATION INJURY 1011

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1012 RADIATION BIOLOGY

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PHYSIOLOGY OF RADIATION INJURY 1013

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1014 RADIATION BIOLOGY

-, and D. M. Gay (1947) Changes produced in testes, spleen, bone

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— and (1949b) Studies of the effects of flavonoids on roentgen irradia-
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3: 245-247.

PHYSIOLOGY OF RADIATION INJURY 1015

Forfota, E., and S. Karady (1937) Tiber die biologische Allgemeinwirkung der

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1016 RADIATION BIOLOGY

Goldman, D. (1943) Metabolic changes occurring as the result of deep roentgen

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med. Wochschr., 50: 2090-2092.
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J. Infectious Diseases, 17: 415-422.
Hempelmann, L. H. (1950) The acute radiation syndrome. USAEC Report

AECU-379. *
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of nine cases and a review of the problem. Ann. Internal Med., 36: 279-510.
T. T. Trujillo, and N. P. Knowlton, Jr. (1949) The effect of lethal doses of

X rays on chilled and thyroidectomized animals. USAEC Report AECU-239;*

Nuclear Sci. Abstr., 3: 46.
Hennessy, T. G, and R. L. Huff (1950) Depression of tracer ion uptake curve in rat

erythrocytes following total-body X irradiation. Proc. Soc. Exptl. Biol. Med.,

73: 436-439.
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changes brought about with single massive doses of radiation. J. Natl. Cancer

Inst., 4: 503-512.

PHYSIOLOGY OF RADIATION INJURY 1017

Hevesy, G. (1946) Effect of X-rays on the rate of turnover of phosphatides. Nature,

158: 268.
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flow in rats. Radiology, 37: 203-204.
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boplastinemia following total-body irradiation. Proc. Soc. Exptl. Biol. Med.,

70: 553-556.
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the Japanese. USAEC Report MDDC-1301.*
Hubert, R. (1929) Der Einfluss der Rontgenstrahlen auf die energieliefernden Reak-

tionen des wachsenden Gewebes. Arch. ges. Physiol. (Pfliigers), 223: 333-350.
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of the gastric region with small doses of roentgen rays upon the stomach and

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genkater. Strahlentherapie, 38: 308-314.
Hursh, J. B., J. B. Mohney, and P. A. Van Valkenburg (1949) Effect of X-irradiation

on thyroid function in rats. USAEC Report AECU-615.*
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487-503.
Ingram, M., and W. B. Mason (1950) The effects of acute exposure to roentgen radia-
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neutrons and X rays. II. Hematological effects produced in the rabbit by fast

neutrons. USAEC Report MDDC-1372.*
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X rays on rabbits. II. The hematological effects of total-body X irradiation in

the rabbit. USAEC Report MDDC-1174,*

1018 RADIATION BIOLOGY

-, M. J. Robson, and E. K. Marks (1950) The effect of X radiation on anti-

body formation. Proc. Soc. Exptl. Biol. Med., 75: 145-152.
— , E. L. Simmons, and M. H. Block (1949) The effect of splenectomy on the

toxicity of Sr89 to the hematopoietic system of mice. J. Lab. Clin. Med., 34:

1640-1655.
— , , E. K. Marks, and J. H. Eldredge (1951) Recovery from radiation

injury. Science, 113: 510-511.
— , , , E. O. Gaston, M. J. Robson, and J. H. Eldredge (1951)

Further studies on recovery from radiation injury. J. Lab. Clin. Med., 37:

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Janzen, A., and S. Warren (1942) Effect of roentgen rays on the peripheral nerve
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Jenkinson, E. L., and W. H. Brown (1944) Irradiation sickness: hypothesis concern-
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Jennings, F. L. (1949) Effect of protein depletion upon susceptibility of rats to total-
body irradiation. Proc. Soc. Exptl. Biol. Med., 72: 487-491.

Johnson, C. G., C. F. Vilter, and T. D. Spies (1946) Irradiation sickness in rats.
Am. J. Roentgenol. Radium Therapy, 56: 631-639.

Jolles, B. (1941) X-ray skin reactions and the protective role of normal tissues.
Brit. J. Radiology, 14: 110-112.

(1950) The reciprocal vicinity effect of irradiated tissues on "diffusible

substance" in irradiated tissues. Brit. J. Radiology, 23: 18-23.

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malignancy; Report of three cases receiving 8,000-10,000 r tissue dose to the

pituitary gland. J. Natl. Cancer Inst., 11: 967-984.
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Kelsall, M. A., and E. D. Crabb (1952) Increased mast cells in the thymus of X-irra-

diated hamsters. Science, 115: 123-125.
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the lethality of roentgen rays for rats. Science, 112: 175-176.
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syndrome. Am. J. Physiol., 162: 703-708.
(1951a) Changes in composition of blood plasma of the rat during acute

radiation syndrome, and their partial mitigation by dibenamine and cortin.

Am. J. Physiol., 165: 27-42.

PHYSIOLOGY OF RADIATION INJURY 1019

— (1951b) Effect of immaturity, hypophysectomy, and adrenalectomy upon
changes in blood plasma of rat during acute radiation syndrome. Am. J. Physiol.,
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— (1951c) Effect of X rays upon hemolysin production in the rat. J. Immunol.,

66: 525-533.

— and P. Robinett (1948) The effect of heparin upon the LD-50% of the
X-irradiated rat. USAEC Report ORNL-1 1 6. *

-, and M. N. Cupp (1948) The effects of rutin upon the response

of the rat to total body X irradiation. USAEC Report AECD-2176.:

Koletsky, S., and J. H. Christie (1950) Effect of antibiotics on mortality from
internal radiation. Proc. Soc. Exptl. Biol. Med., 75: 363-366.

Kotz, J., J. F. Elward, and E. Parker (1941) Non-harmful effects of irradiation of
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Krause, P., and K. Ziegler (1906) Experimentelle Untersuchungen ueber die Ein-
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Krizek, H., I. Silverbach, and S. Schwartz (1946) The effect of lethal dose of total-
body X ray on uric acid and allantoin excretion in dogs. USAEC Report
MDDC-1674.*

Larkin, J. C. (1941) An erythematous skin reaction produced by an alpha-particle
beam. Case report. Am. J. Roentgenol. Radium Therapy, 45: 109.

(1942) Some physiological changes in the skin produced by neutrons. Am.

J. Roentgenol. Radium Therapy, 47: 733-739.

(1949) Effect of atropine on acute irradiation sickness in mice. Am. J.

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the whole body. J. Exptl. Med., 66: 667-688.
Lawrence, J. S., A. H. Dowdy, and W. N. Valentine (1948) Effects of radiation on

hemopoiesis. Radiology, 51: 400-413.
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Lazarew, N. W., and A. Lazarewa (1926) tlber die funktionellen Veranderungen der

Blutgefasse nach Rontgenbestrahlung. Strahlentherapie, 23: 41-78.
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heart of adult rats. Am. J. Roentgenol. Radium Therapy, 48: 81-87.
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effects of roentgen rays upon the organs of normal and adrenalectomized rats.

Am. J. Roentgenol. Radium Therapy, 47: 302-306.
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Leinfelder, P. S., and H. D. Kerr (1936) Roentgen-ray cataract: An experimental,

clinical, and microscopic study. Am. J. Ophthalmol., 19: 739-756.

1020 RADIATION BIOLOGY

LeMay, M. (1951) Effect of X-radiation upon succinoxidase of the rat kidney.
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Lick, L., A. Kirschbaum, and H. Mixer (1949) Mechanisms of induction of ovarian
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Liebow, A. A., S. Warren, and E. DeCoursey (1949) Pathology of atomic bomb
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Liechti, A., and W. Wilbrandt (1941) Radiation hemolysis. I. Hemolysis by
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Light, A. E. (1935) A histologic study of the effects of X-rays on frog skin. Radi-
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Lorenz, E., C. C. Congdon, and D. Uphoff (1952) Modifications of acute radiation
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, W. E. Heston, L. O. Jacobson, A. B. Eschenbrenner, M. Shimkin, M. K.

Deringer, and J. Doninger (1946) Biologic effects of long-continued whole-body
irradiation with gamma rays on mice, guinea pigs, and rabbits. III. Biological
effect of whole body irradiation with gamma rays: effects on life span, weight,
blood picture, and carcinogenesis, and the role of intensity of the radiation.
USAEC Report MDDC-655.*

, D. Uphoff, T. R. Reid, and E. Shelton (1951) Modification of irradiation

injury in mice and guinea pigs by bone-marrow injections. J. Natl. Cancer

Inst., 12: 197-201.
Lourau, M., and O. Lartigue (1950a) Modifications of glycemia under the influence of

total X-ray radiation. Compt. rend., 230: 1426-1428.
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produits par une irradiation unique de tout le corps (rayons X). Experientia,

6: 25-26.
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Roentgenstrahlenwirkung. Strahlentherapie, 46: 469-516.
Ludewig, S., and A. Chanutin (1950a) Effect of X-ray irradiation on alkaline phos-
phatase of plasma and tissues of rats. Am. J. Physiol., 163: 648-654.

and (1950b) Distribution of enzymes in the livers of control and

X-irradiated rats. Arch. Biochem., 29: 441-445.

Lukens, R. M. (1948) Complications following irradiation of the thyroid gland.

Ann. Otol. Rhinol. and Laryngol., 57: 633-642.
Lutwak-Mann, C, and F. W. Gunz (1949) Effect of body exposure to X rays upon

bone marrow. Biochem. J., Vol. 44, Proc. iii, No. 2.
Macht, S. H., and H. Perlberg, Jr. (1950) Use of anticoagulant (dicumarol) in

preventing postirradiation tissue changes in the human lung; preliminary report.

Am. J. Roentgenol. Radium Therapy, 63: 335-341.
Marder, S. N., H. S. Kaplan, and E. Lorenz (1948) Lack of effect of adrenalectomy

on the lymphopenic response of C57 black mice to total-body irradiation at a

low dosage level. USAEC Report ANL-4227,* pp. 21-27.
Martin, C. L. (1950) Radiation therapy in diseases of the female genital organs, in

E. A. Pohle, Clinical radiation therapy. Lea & Febiger, Philadelphia, pp.

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-, and N. F. Fisher (1924) The effect of roentgen rays on the adrenal

gland. Am. J. Roentgenol. Radium Therapy, 12: 466-471.
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tivity. Arch. Path. Lab. Med., 2: 465-472.
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Cancer, 15: 2435-2516.

PHYSIOLOGY OF RADIATION INJURY 1021

McQuarrie, I., and G. H. Whipple (1922) A study of renal function in roentgen-ray

intoxication. Resistance of renal epithelium to direct radiation. J. Exptl. Med.,

35: 225-242.
Mead, J. F., A. B. Decker, and L. R. Bennett (1950) The effect of X-irradiation

upon fat absorption in the mouse. USAEC Report UCLA-90;* J. Nutrition,

43: 485-500 (1951).
Miescher, G. (1923) tjber den Einfluss der Rontgenstrahlen auf die Sekretion des

Magens. Strahlentherapie, 15: 252-272.

(1924) Das Rontgenerythem. Strahlentherapie, 16: 333-371.

(1938) Vergleichende Untersuchungen zur Frage der Spezifitaet der Roent-

genwirkung. Licht-, Waerm-, Roentgen-, und Thorium-X-Reaktion. Strahlen-
therapie, 61: 4-37.

Miller, C. P., C. W. Hammond, and M. Tompkins (1950a) The incidence of bac-
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) ( and (1950b) Reduction of mortality from X radiation by

treatment with antibiotics. Science, 111:719-720.

Miyazaki, K. (1937) Effect of X ray (50-100 r) on urinary excretion of creatine,
creatinine, uric acid, and allantoin. Sei-i-Kei Med. J., 56: 2085-2104; Chem.
Abstr., 32: 5011, 1938.

Montgomery, P. O., and S. Warren (1951) Mechanism of acute hypotension follow-
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Moon, V. H., K. Kornblum, and D. R. Morgan (1940) Pathology of irradiation
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305-306.
; ; and (1941) The nature and pathology of radiation sickness.

J. Am. Med. Assoc, 116: 489-493.
Mottram, J. C, and L. H. Gray (1940) The relative response of the skin of mice to

X radiation and gamma radiation. Brit. J. Radiology, 13: 31-34.
and A. N. Kingsbury (1924) Some researches into the action of radium and

X rays correlating the production of intestinal changes, thrombopoenia, and

bacterial invasion. Brit. J. Exptl. Path., 5: 220-226.
Muntz, J. A., E. S. G. Barron, and C. L. Prosser (1949) Studies on the mechanism of

action of ionizing radiations. III. The plasma protein of dogs after X-ray

irradiation. Arch. Biochem., 23: 434-445.
Murphy, J. B., and E. Sturm (1925) A comparison of the effects of X-ray and dry

heat on antibody formation. J. Exptl. Med., 41: 245-255.
Murray, R., M. Pierce, and L. O. Jacobson'(1948) The histological effects of X-rays

on chickens with special reference to the peripheral blood and hemopoietic

organs. USAEC Report AECD-2303.*
Narat, J. K. (1922) Treatment after irradiation with roentgen ray. J. Am. Med.

Assoc, 79: 1681-1684.
Neve, R. A., and C. Entenman (1951) Changes in the plasma phospholipids in rats

following whole body X-irradiation. Nav. Radiol. Def. Lab. Report AD-

307(B).*
Nizet, E., C. Heusghem, and A. Herve (1949) Suprarenal cortex reactions following

the administration of X-rays from a distance in the rabbit. Compt. rend. soc.

biol., 143: 876-877.
North, N., and L. F. Nims (1949) Time-dose study of biochemical responses of rats

to X-radiation. Federation Proc. 8: 119-120.
Novick, R. (1946) Time course of changes in sensory cells of trigeminal ganglion of

the rabbit following irradiation. Proc. Soc Exptl. Biol. Med., 61: 355-358.
Oster, R. H., and W. T. Salter (1938) Immunization against neoplasm. Its effect

on the nitrogen metabolism of the host. Am. J. Cancer, 32: 422-433.

1022 RADIATION BIOLOGY

Packard, C. (1933) Biologic effects of roentgen rays and radium, in Science of
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Painter, E. E. (1948) Alterations in the sodium space of the gastrointestinal tract
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and A. M. Brues (1949) The radiation syndrome. New Engl. J. Med.,

240: 871-876.

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rabbits exposed to single doses of X rays. USAEC Report MDDC-761. *

and E. W. Pullman (1950) Water and electrolyte changes in rat intestine

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Parkes, A. S. (1948) Effects on early embryonic development of irradiation of

spermatozoa. Brit. J. Radiology, Suppl. 1, pp. 117 120.
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blood of the irradiated rat. Blood, 5: 758-763.
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of estrogens on the acute X-irradiation syndrome. Am. J. Physiol., 159:

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■ , M. N. Swift, and E. B. Tyree (1949) Oxygen consumption and water balance

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X-'radiation. Am. J. Physiol., 150: 480-487.
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sectomized rat. Science, 108: 475-476.
Pendergrass, E. P., P. J. Hodes, and J. Q. Griffith (1944) Effect of roentgen rays
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and J. F. Mahoney (1948) A consideration of the roentgen therapy in

producing temporary depilation for Tinea capitis: a new method. Radiology,
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Penick, G. D., E. P. Cronkite, I. D. Godwin, and K. M. Brinkhaus (1951) Plasma

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capillary changes after exposure to unfiltered radiation. Radiology, 6: 236-

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(1927) Studies of the roentgen erythema of the human skin. II. Skin

capillary changes after exposure to filtered roentgen rays and to ultraviolet radia-
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and C. H. Bunting (1932) Studies of the effect of roentgen rays on the liver.

Histological changes in liver of rats following exposure to single graded doses of

filtered roentgen rays. Acta Radiol., 13: 117-124.
Pratt, A. W., B. E. Burr, M. Eden, and E. Lorenz (1950) Preliminary report:

Mass spectrometer analysis of the respired air of rats. USAEC Report ANL-

4531,* pp. 40-46.
Prosser, C. L., and M. C. Moore (1946) The clinical physiology of dogs exposed to

daily total-body doses of X rays. USAEC Report MDDC-1271.*

PHYSIOLOGY OF RADIATION INJURY 1023

— , E. E. Painter, H. Lisco, A. M. Brues, L. O. Jacobson, and M. N. Swift (1947)
The clinical sequence of physiological effects of ionizing radiation in animals.
Radiology, 49: 299-313.
— , - — and M. N. Swift (1947) The clinical physiology of dogs exposed to

single total-body doses of X-rays. USAEC Report MDDC-1272. *
Pullinger, B. D. (1932) Causes of cell death in irradiated human tissue. J. Path.

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Quastler, H., E. F. Lanzl, M. E. Keller, and J. W. Osborne (1951) Acute intestinal

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radiation, R. E. Zirkle, ed. McGraw-Hill Book Company, Inc., New York,

National Nuclear Energy Series, Div. IV, Vol. 22E, Chap. 10.

— and - — (1951b) Gross effects of total-surface beta irradiation, in Biological
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McGraw-Hill Book Company, Inc., New York, National Nuclear Energy Series,
Div. IV, Vol. 22E, Chap. 7.

— , R. E. Zirkle, and K. K. Barnes (1951) Comparative lethal effects of external

beta irradiation, in Biological effects of external beta radiation, R. E. Zirkle, ed.

McGraw-Hill Book Company, Inc., New York, National Nuclear Energy Series,

Div. IV, Vol. 22E, Chap. 3.
Regaud, C, and T. Nogier (1913) Les effets produits sur la peau par les hautes doses

de rayons X selectionnes par la filtration a travers 3 a 4 millimetres d'aluminium.

Arch, d'electric med., 21: 49-97.
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etendues de l'abdomen et sur les lesions du tube digestif determinees par les

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1024 RADIATION BIOLOGY

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PHYSIOLOGY OF RADIATION INJURY 1025

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1026 RADIATION BIOLOGY

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PHYSIOLOGY OF RADIATION INJURY 1027

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1028 RADIATION BIOLOGY

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Manuscript received by the editor June 20, 1952

CHAPTER 16

The Hematologic Effects of Ionizing Radiation

Leon 0. Jacobson

Professor of Medicine, Department of Medicine, The University of Chicago,

Chicago; Scientific Director of the Argonne Cancer Research Hospital of the

University of Chicago; Division of Biological and Medical Research, Argonne

National Laboratory, Lemont, Illinois

Introduction. Comparative effects of ionizing radiations: Single exposures to X rays —
Single exposures to fast neutrons — Comparison of effects produced by cyclotron fast neutrons
and X rays — Factors concerning coagulation following single exposures. Hematologic
effects of chronic total-body exposure to external radiations: Chronic exposure to X rays
and fast neutrons — Chronic exposure to repeated small doses of fast neutrons — Chronic
exposure to small daily doses of fast neutrons — Chronic exposure to gamma rays (radium)
— Pancytopenia induced by chronic exposure to gamma radiation. Hematologic effects
of exposure to externally originating radiation (radioelements) : Phosphorus (P32) —
Sodium (Na,2i)— Barium-Lanthanum (Ba-La140)— Yttrium (Y91)— Strontium (Sr89)—
Radium— Plutonium (Pu239)— Gold (Au189)— Gallium (Ga72). Morphologic changes in
peripheral-blood cells produced by ionizing radiations: Direct vs. indirect effect of irradia-
tion on blood formation — "Stim ulation" of blood-forming tissue by radiation. Measures
modifying destructive effects of irradiation or affecting recovery: Prophylactic measures —
"Therapeutic" measures — Effects of combined prophylactic and therapeutic measures —
Antibody-formation studies — Comment. References.

INTRODUCTION

As a result of Heineke's (1903) pioneering research almost fifty years
ago it has been recognized that the blood-forming tissues are among the
most sensitive to ionizing radiations. After total-body exposure, mani-
festations of injury to the mammalian blood-forming tissues (such as
lymphopenia) may appear in the peripheral blood even in the absence of
demonstrable histopathologic change in the bone marrow or lymphatic
tissues. The reviews of Warren (1943), Warren and Whipple (1922), and
others (Minot and Spurling, 1924; Selling and Osgood, 1938) have
covered rather adequately the significant literature up to 1942. Few
observations have appeared in the past two decades that were not
recorded previously in the classic descriptions of the effects of X rays on
the blood and blood-forming tissues described in the earlier literature by
Lacassagne and Levadan (1924), Russ et at. (1919), Heineke (1904),

1029

1030 RADIATION BIOLOGY

Desjardins (1932), Aubertin and Beaujard (1905), Clarkson et al. (1938),
Mottram and Russ (1921), Czepa (1923-1924), and Linser and Helber
(1905). Because of the fact that dosage measurement was neither
accurate nor accurately reproducible and because of other technical
difficulties, much of the early research has been repeated in order to cor-
relate dose with effect. In addition, new types of radiation have become
available which have made comparative studies on the biological effects
of various radiations desirable. This chapter is based largely on studies
conducted by various investigators during the past ten years.

COMPARATIVE EFFECTS OF IONIZING RADIATIONS

In considering the irradiation of the whole body with penetrating radia-
tions of external origin, those of significance in so far as direct effects on
the blood and blood-forming tissue are concerned are X rays, y rays, fast
neutrons, and slow neutrons. The 0 rays and a particles that emanate
from radioisotopes can, in general, be disregarded because penetration in
tissue is only a few millimeters for /3 rays and a fraction of a millimeter for
a particles. Studies on mice and rabbits by Raper and Barnes (1951),
Raper, Zirkle, and Barnes (1951), Raper, Henshaw, and Snider (1951),
and others have demonstrated that, although /3 rays may penetrate the
skin, the hematologic effects, if any, are minimal and are secondary to
other pathologic changes such as ulceration of the skin. No studies are
yet available on the hematologic effect of exposure to /3 rays from a
betatron source.

Studies on the effect of acute exposure of animals to a slow-neutron
flux have been carried out by Zirkle (1945) and by Zirkle et al. (1947).
This slow-neutron source was contaminated with fast neutrons, and the
effect on the hematopoietic system, as reported by Raper, Henshaw, and
Snider (1951), is therefore difficult to interpret. According to Zirkle
(1947, 1950), however, it is not the slow neutron that produces biologic
effects but rather the 7 rays and fast atomic nuclei emitted upon nuclear
capture of the slow neutrons. It is likely, therefore, that exposure of
experimental animals to a pure slow-neutron flux would produce hemato-
logic effects comparable to those induced by 7 rays plus fast neutrons.
Studies using a 7-ray source for single total-body exposures delivered in a
few minutes, as reported by Henshaw et al. (1946) and by Henshaw,
Riley, and Stapleton (1947), indicate that the effect on the blood and
blood-forming tissue is qualitatively the same as that produced by X rays,
and therefore no special category of discussion is devoted to this subject.

Data on single exposures of rabbits to fast neutrons in about the 30-day
LD50 range (Jacobson and Marks, 1947) indicate that the degree of
depression of the formed elements in the circulating blood and the time
required for recovery are largely comparable to those observed after a

HEMATOLOGIC EFFECTS OF RADIATION 1031

30-day LD50 exposure of rabbits to X rays. This information is fairly
complete for dosage ranges up to the LD5o, but more complete data are
needed for higher dosages.

Correlation between the effects of these penetrating radiations on
blood-forming tissue and various hematologic constituents of the periph-
eral blood is generally good when the effects of dosages in the LD5o range
for any particular species are being considered. Major deviations, how-
ever, are recognized. With doses of 100 r or less there is a lack of correla-
tion because the changes that occur in the lymphatic tissue are difficult
to quantitate histologically. The lymphocytes in the peripheral blood
may be drastically reduced, although only equivocal changes are seen in
the lymphatic tissues.

SINGLE EXPOSURE TO X RAYS

Studies of the effects of a single total-body exposure to X rays indicate
that the rabbit (Hagen et al., 1944; Jacobson et al, 1947), rat (Steamer
et al., 1947a, b; Lawrence and Lawrence, 1936), monkey (Suter, 1947:
Ingram and Mason, 1950a), mouse (Henshaw, 1943-1944a; Bloom and
Bloom, 1947), chick (Murray et al, 1948), goat (Swift et al., 1946), pig
(Cronkite et al., 1949), guinea pig (Henshaw, 1943-1944b; Haley and
Harris, 1949; Lorenz, 1951; Brues and Rietz, 1948), and dog (Prosser
et al., 1947; Allen et al., 1948) show, in order, an increasing sensitivity to
radiation as measured by changes in the blood and blood-forming tissues.
As compared with the other species, it should be noted that the data on
the hematologic effects in monkeys are conspicuously inadequate.

Effects on Leukocytes. The term "leukocyte" is applied to all the
white cells in the peripheral blood. In the species discussed in this
chapter the polymorphonuclear cells of the granulocyte series and the
lymphocytes are the predominant forms. These constituents are dis-
cussed individually.

Aubertin and Beaujard (1908) first described the appearance of a
leukocytosis prior to leukopenia after total-body X irradiation. Actu-
ally, during the first 24 hours after exposure the total leukocyte count of
the peripheral blood reflects the rapidly changing status that may be
described briefly as follows: With doses of 500 r an initial significant
reduction occurs in the number of circulating leukocytes in the rabbit.
Since the number of heterophils remains constant during this period, the
reduction can be attributed to lymphocyte reduction which has been
initiated and continues. On the other hand, leukocytosis is apparent at
8 hours and again at about 24 hours. This leukocytosis, which is dis-
cussed in detail later, results entirely from an increased number of
heterophils.

Effects on Lymphocytes. With the possible exception of erythrocyte
iron uptake, the most sensitive indicator of acute radiation effect that can

1032

RADIATION BIOLOGY

be recognized by present methods in any of the various species studied is
a reduction in the number of lymphocytes in the peripheral blood. In
general, this effect is not significant for doses below 25 r. The pattern
of acute radiation response of the rabbit lymphocyte following dosages
ranging from 25-800 r is shown in Fig. 16-1. The response of the
lymphocytes of the circulating blood of dogs, mice, monkeys, rats, and
guinea pigs is fairly comparable to that of rabbits. According to Valen-
tine et al. (1947), cats exposed to 200 r respond with a lesser lymphopenia
than do the species listed. Lymphocyte values in rabbits fall below

8000

CO
3hr

CONTROLS
300 r

6 8 20 40 60

TIME AFTER EXPOSURE, days

a m 25 r o a 50 r o 1

« o 500 r o

140

100 r
700 r

200 r

600r « » 700r « e 800 r

Fig. 16-1. Effect of single doses of total-body roentgen irradiation on the lymphocyte
values of the peripheral blood of rabbits.

control values after 25 r and return to normal within 24-48 hours. With
doses of 50 and 100 r and above, a reduction occurs within 15 minutes, and
a maximum depression is reached by 24-48 hours. With dosages of
300 r and above, lymphocyte values approach the same minimum, and the
time required for recovery is a function of dose. Table 16-1 gives the
approximate mean time of recovery of lymphocyte values to normal
limits after dosages from 25-800 r. These data are in agreement with
those of Hayer (1934), Taylor et al. (1919), Thomas et al. (1919), Clarkson
et al. (1938), Linser and Helber (1905), Siegel (1920), Russ et al. (1921),
Russ (1921), and others.

In all species the lymphocyte values of the peripheral blood are gen-
erally the last to return to normal levels. After a dose of one-half the

HEMATOLOGIC EFFECTS OF RADIATION

1033

LD50 these values may not reach the normal range for 30-90 days.
According to Bloom (1947), lymphatic tissue and lymphocyte production
after exposures in this range are qualitatively normal as judged by
histologic methods at 20-30 days. This discrepancy between the findings
in the peripheral blood and in the lymphatic tissue is understandable when
it is realized that histologic findings are admittedly qualitative.

Table 16-1. Effect of Total-body X Irradiation and Exposure to Fast Neu-
trons on the Leukocyte Values of the Peripheral Blood of Rabbits

Dose

Degree of depression,
per cent

Time of maximum
depression, hours

Time required before

normal limits attained,

days

Lympho-
cytes

Hetero-
phils

Lympho-
cytes

Hetero-
phils

Lympho-
cytes

Hetero-
phils

5 r
10 r

25 r
50 r

100 r
300 r
500 r
600 r
700 r
800 r

9n

26 n
55 n
68 n
76 n
89 n
97 n

106 n
128 n

0
0
25
25
50
74
90
90
90
90

40

65
86
94
92
93
95
97
95

0
0
0
0

50

75
80
90

0
50

77
77
85
87
83
92
92

24
48
48
24
48
48
48
72

48
72
48
48
48
48
72
48
48

72
96
96
96

0
96
96
96
96
96
96
96
96

48 (hours)
16
36
50
50
50

50

5

a
a
a
• a
a
a
a
a

9
9

23

0
5

7

9

9

16

15

16

0 Recovery not complete during 37 days of observation.

Monocytes of the peripheral blood initially follow a pattern of response
similar to that of the lymphocytes but characteristically return to normal
values or show an absolute increase between the fourth and sixth days
after exposure of the animal to 100 r or above. No change of significance
has been observed in the number of plasma cells in the peripheral blood.

The polymorphonuclear leukocyte, referred to as a heterophil in the
rabbit and a neutrophil in man, follows a somewhat different pattern than
does the lymphocyte after total-body irradiation. The most complete
data have been derived from observations on rabbits. The sensitivity of
the heterophil in mice, rats, rabbits, monkeys, and swine is, in general,

1034

RADIATION BIOLOGY

comparable. Although individual animals in all these species may have
a reduction in the circulating heterophils after exposure to 100 r, a
significant reduction in well- controlled experiments occurs only after
exposure to 200 r or more. The relatively great sensitivity of the blood-
forming tissue of the guinea pig, cat, and dog, however, is reflected in a
significant heterophil reduction even after exposure to dosages of 200 r
or less. Aubertin and Beaujard (1908) reported an initial rise in the
polymorphs following exposure to dosages in the LD50 range, with a
return to about normal in 24 hours and a maximum depression by 2-4
days. This was confirmed by Jacobson et al. (1947), who showed, in
addition, that actually two separate elevations occur in the rabbit in the

12,000

12 16 20 24 28 32
TIME AFTER EXPOSURE, hours
-* CONTROLS o o iQOr a * 500 r

36 40

Fig. 16-2. Effect of single doses of total-bod y roentgen irradiation on the heterophil
values of the peripheral blood of rabbits.

first 24 hours after exposure (Fig. 16-2). A heterophil rise in the first 24
hours is characteristic of all species studied, but this rise may be entirely
masked in species in which lymphocytes constitute the larger percentage
of the circulating leukocytes. That the initial rise in granulocytes is
followed by a reduction after exposure of rabbits to dosages of 500 r or
greater is shown in Fig. 16-3. Recovery is usually complete in 12-21
days even with dosages of 800 r. An "abortive rise" in the heterophil
value is characteristically seen after dosages of 500-800 r. This tem-
porary elevation appears between the fourth and eleventh day. A similar
elevation is observed in lymphocyte values during the same postirradia-
tion period (Fig. 16-1).

The mechanism of the leukocytosis within the first 24 hours after
exposure is not well understood. Isaacs (1934) and Wuensche (1938)
described a hastening of maturation of granulocyte precursors in the bone
marrow and have suggested that the release of these cells into the circula-

HEMATOLOGIC EFFECTS OF RADIATION

1035

tion accounts for leukocytosis. In view of the fact that two separate
peaks occur in this period, Jacobson et al. (1949) suggested that the first
peak might be accounted for by hastening of maturation in the bone
marrow and the second peak by a mobilization phenomenon in response
to widespread tissue injury by irradiation. Histologic studies by Bloom
(1947) have shown that the granulocyte peaks, described in the peripheral
blood, are seen especially in the lymphatic tissues; an "invasion" of
these tissues with granulocytes occurs concomitantly with the peaks
observed in the peripheral blood.

10,000
8000

6000

200

CO
3hr

J — 1 L

J L

I

8

_L

20 40 60 80 100 120 140

CONTROLS

TIME AFTER EXPOSURE, days

— »25r o- a 50 r y ylOOr

— o 500 r o a 600 r e— — e 700 r

200 r

800 r

o- -o 300 r o-

Fig. 16-3. Effect of single doses of total-body X irradiation on the heterophil values
of the peripheral blood of rabbits.

Detailed studies on the effect of irradiation on the circulating eosino-
phils are not available. However, Aubertin and Beaujard (1908)
reported an increase in animals following irradiation, and Lawrence and
associates (1949) have reported an initial increase in dogs after an LD50
exposure.

The " Abortive Rise." An elevation in the lymphocyte, heterophil, and
reticulocyte values occurs in the rabbit after exposure to dosages of 300 r
and above (Figs. 16-1, 16-3). This elevation, first described by Jacobson
et al. (1947) and Jacobson, Marks, and Lorenz (1949), has been observed
by others in cats (Valentine and Pearce, 1952; Adams and Lawrence,
1947), dogs (Allen, 1947-1948), pigs (Cronkite et al, 1949), rats (Cohn.
1952; Suter, 1947), and guinea pigs (Lorenz, Uphoff, and Sutton, 1949).
The significance of this elevation, which appears generally between the
fourth and eleventh postirradiation day, is not known. It has been sug-

1036

RADIATION BIOLOGY

gested by Jacobson, Marks, and Lorenz (1949) that it may represent a
multiplication of cells that were injured at the time of irradiation and
that died after a limited number of divisions. Cells of the granulocyte
series, which are grossly abnormal morphologically, are found in the
peripheral blood and in the bone marrow (Bloom, 1948; Bloom and
Jacobson, 1948) during this period. The temporary increase in these
various cell types in the peripheral blood parallels temporary waves of

— •-

_L

J_

_L

_L

6 8

10 12 14 16 18 20 22 24 26 28 30 32 34

TIME AFTER X RAY, doys

8 6 4 2 0 2
PHENYLHYDRAZINE
ADMINISTRATION

©— © PHENYLHYDRAZINE AND 800 r OF TOTAL-BODY X IRRADIATION

• • CONTROLS. o— • PHENYLHYDRAZINE ONLY

o— — o 800 r OF TOTAL-BODY
Fig. 16-4. Effect of a single dose of 800 r of total-body roentgen irradiation on the
reticulocyte values in the peripheral blood of normal rabbits and rabbits with a
phenylhydrazine-induced anemia. {Originally published in Science, 107: 249, 1948.)

regeneration observed histologically by Bloom and Bloom (1947) and
Bloom (1948) in the bone marrow especially after radiation injury.

Radiation-induced Anemia. Red-cell precursors (erythroblasts) have
been described by Bloom and Bloom (1947) as being the most sensitive
to irradiation injury of all the cellular elements of mammalian bone
marrow. This conclusion is based on histologic evidence of the degree of
destruction of the erythroblasts in comparison with that of other cell
types such as granulocyte precursors and megakaryocytes. These
observations were made on mice, rats, chickens, and rabbits exposed to
30-day LD&0 dosages and those well below this range. In fact it has been
suggested by Bloom and Bloom (1947) that the sensitivity of erythro-
blasts is comparable to that of the lymphocyte. It must be pointed out,
however, that Bloom compared only the immediate morphologic response

HEMATOLOGIC EFFECTS OF RADIATION

1037

of cells to irradiation. Such observations should not be construed to
imply that sensitivity and rapidity of regeneration or functional reconsti-
tution are synonymous. Rates of regeneration from an atrophic hemato-
poietic tissue of various cell types differ markedly.

The erythrocyte, hemoglobin, or hematocrit values of the peripheral
blood do not reflect this erythroblast sensitivity in animals such as
rabbits, rats, and mice unless the total-body exposure is above 300 r. On
the other hand, reticulocytes, which are the immediate precursors of the
erythrocytes, are significantly reduced in these species after total-body
irradiation at dosages of 100 r and above. The findings of Bloom on the
relative sensitivity of the erythroblast have been confirmed by Hennesey
and Huff (1950). These authors studied the depression of tracer iron
uptake (Fe59) by rat erythrocytes following exposure to various dosages.
By this method they found that a significant inhibition of erythropoiesis
was apparent 24 hours after exposures as low as 5 and 25 r. On the basis
of available data from peripheral-blood studies, guinea pigs have the most
sensitive erythropoietic tissue of all the common laboratory animals.
Lorenz (1951) has shown that a severe anemia develops in guinea pigs
after exposure to 200 r, which is one-half the 30-day LD50 for this species.
Dogs are more nearly comparable to the guinea pigs in this respect.
Rabbits, which are the most resistant of the common laboratory animals
to irradiation (LD50, 800 r), develop a less severe anemia after an LD5o
exposure than do dogs, cats, guinea pigs, or mice. This is apparent in
material presented by Valentine and Pearce (1952) (see Table 16-2).

Table 16-2. Effect of Total-body X Irradiation on the Erythrocyte Values

of the Peripheral Blood

Species

Dosage,
r

Time of maximum
reduction, days

Per cent
reduction

Per cent recovery
at 26 days

Reference"

Guinea pigs .

220

14

60

77.7

310

14

71.1

55.6

420

14

62.2

82.3

1

Rats

300

12

18

95

500

18

45

70

2

550

250 (hours)

81.8

100 (41 days)

3

600

14

32.2

67.7

4

700

18

72

60.0

1

Rabbits. . . .

500

17

19.3

85.5

5

800

15

27.5

86.2

Dogs

200
300

20
23

42.5
50 0

^j}24-35days

6

350

21

32.25

79.04

7

a References: (1) Lorenz, 1951; (2) Suter, 1947; (3) Lawrence, Dowdy, and Valen-
tine, 1948; (4) Bennett, Hanson, and Dowdy, 1951; (5) Jacobson et al., 1947; (6)
Prosser, Painter, and Swift, 1946; (7) Rekers, 1949.

1038

RADIATION BIOLOGY

Determination of the erythrocytes of the peripheral blood and their
hemoglobin content by standard techniques is a rather crude means of
measuring the effect of irradiation, especially during the first two weeks
after exposure to X-ray dosages in the LD5o range or above. Even after
exposure to dosages well above the lethal range it is common experience
to find that no significant reduction in the erythrocyte, hemoglobin, or
hematocrit values occurs prior to the sixth day. Since, after exposure to
a dosage of this magnitude, destruction of the bone marrow occurs rapidly
(within the first 24 hours) and thus delivery of erythrocytes ceases, a
steady decline of these hematologic values would be expected to begin

e

E

v.
u>

aj
o

o

>■
o
o

X

(-
>-
cr

6 8 10 12 20

TIME AFTER X RAY, days

-•CONTROL

O -O OPERATED CONTROLS

A A 600 r WITH SPLEEN EXTERIORIZED

® ©1025 r WITH SPLEEN EXTERIORIZED

Fig. 16-5. Comparative effect of 600 and 1025 r of total-body roentgen irradiation on
the erythrocyte values of CFi female mice.

simultaneously with exposure and continue until such time as regenera-
tion and delivery of erythrocytes equaled or exceeded these effete cells.
As suggested by Furth et al. (1951) and Jacobson, Simmons, Marks, et al.
(1950), the destruction of hematopoietic tissue and thus the cessation of
erythropoiesis that occurs in dogs, rabbits, rats, pigs, guinea pigs, and
mice after exposure to dosages in the LD50 range may be sufficient to
account for the anemia observed. However, the degree and rapidity
with which anemia develops above the LD5o range suggests that addi-
tional factors are involved. The speed and severity with which anemia
appears in mice exposed to 1025 r as compared with that following 600 r
exemplifies this problem (Fig. 16-5). Histopathologic investigations
show that erythropoiesis ceases within 24 hours after either level of
exposure. The anemia would be expected to be qualitatively the same

HEMATOLOGIC EFFECTS OF RADIATION 1039

at about 9 days if a cessation of erythropoiesis was the only aspect
involved. Gross hemorrhage in mice is not a factor under these condi-
tions. Microscopic hemorrhage and extensive erythrophagocytosis are
constant histopathologic findings after exposure to 1025 r. Both proc-
esses culminate in the eventual hemolysis of the involved red cells. (As
described later, gross hemorrhage may occasionally be a major factor in
the production of an anemia in dogs, pigs, and guinea pigs.) Schwartz
et al. (1947) and Lawrence, Dowdy, and Valentine (1948), and others have
studied the role of red-cell hemolysis in postirradiation anemia and have
found evidence of an increased excretion of the breakdown products of
hemoglobin. The increased pigment excretion observed in dogs after
exposure to an LD5o dose is not of sufficient magnitude to warrant the
conclusion that increased erythrocyte hemolysis plays a role in the
anemia since an indeterminate number of hemoglobin-containing erythro-
cyte precursors are destroyed in situ by irradiation. This may account
for the increased pigment excretion. Davis et al. (1950) restudied this
problem and found an increase in bile-pigment excretion in dogs exposed
to 150-250 r during the first and second weeks after irradiation. No
increased erythrocyte fragility (thermal or mechanical) was apparent,
however, in these animals. Davis and his co-workers considered the
possibility that increased pigment excretion might be based on destruc-
tion of the erythrocyte precursors. Prosser et al. (1947) recognized that
the changes occurring in circulatory dynamics were masking actual
changes in red-cell and hemoglobin levels and that the anemia that
eventually developed after exposure of the dog or rabbit to LD5o dosages
could not be accounted for by cessation of hematopoiesis or hemorrhage
and postulated, as have many others (Heineke, 1905; Schwartz et al.,
1947; Dunlap et al., 1944; Selling and Osgood, 1938), that an abnormal
hemolysis of erythrocytes was probably involved.

Recently Furth and associates (Storey, Wish, and Furth, 1950; Furth
et al., 1951; Kahn and Furth, 1952; Ross, Furth, and Bigelow, 1952),
using a number of techniques including P32 and Fe59 labeling of the red
cells and I131 labeling of the plasma combined with regular hematologic
and histopathologic investigation, have clarified many of the problems
concerning postirradiation anemia. This work, based on studies of mice,
rabbits, and dogs X-irradiated in the LD5o range and above, may be
summarized as follows:

1. Erythropoiesis ceases within 24 hours after irradiation and is not
resumed for 7-14 days.

2. During this same period there is a reduction in the red-cell mass as
determined by isotopic studies (Fe59 and P32), and there is a simultaneous
decrease in the plasma volume. This change masks the magnitude of the
drop in erythrocyte mass, especially during the first postirradiation week,

1040

RADIATION BIOLOGY

and accounts for the fact that during this period erythrocyte, hemoglobin,
and hematocrit determinations are frequently within the normal range.

3. Reduction in red-cell mass results from (a) death of normally aging
red cells and (6) the loss of "normal" erythrocytes from the circulation
by heightened capillary permeability and thus widespread minute
extravasation into lymphatic tissues and tissue spaces. Tagged red cells
(Fe59 and P32), as well as tagged plasma, were found to disappear more
rapidly from the circulation of irradiated than from that of nonirradiated
animals. The leakage of erythrocytes is a major factor in the production

CD E

So
o^

UJ

X

o>

i 1 i 1 1 r*f* — i 1 1 1 r

V**

J I L

J L

J I 1 L

tn

UJto

M

0 ^
o: -

1 OJ

I- o

>-u>

0= o
UJ -

"i i i I 1 1 1 r

i 1 r

\ ~-*D~-^

V

>&=»

~^=*= • =

j_

_L

_L

_L

_L

_L

_L

0 10 20 30 40 50 60 70 80 90 100 110 120

TIME AFTER EXPOSURE, days

• 'CONTROL o -o800r »•-— « 500 r

Fig. 16-6. Effect of single doses of 500 and 800 r of total-body roentgen irradiation on
the hemoglobin and erythrocyte values of the peripheral blood of rabbits.

of anemia. The fate of the red cells thus extravasated as a result of
primary or secondary endothelial injury is phagocytosis and eventual
hemolysis.

No significant anemia, as judged by mean averages, has been observed
in groups of rabbits exposed to dosages below 500 r. In individual
animals, anemia may appear with exposures as low as 300 r. A progres-
sively larger fraction develops anemia following exposure to 500 r and
above (Jacobson et al., 1947; Jacobson, Marks, and Lorenz, 1949).
Figure 16-6 illustrates the anemia observed in rabbits after exposure to
800 r. Rats, mice, dogs, cats, swine, and guinea pigs develop anemia in
response to lower doses. Figure 16-7 illustrates the anemia in guinea
pigs after exposure to 220 r and above. In the absence of gross hemor-
rhage the anemia is normochromic and reaches a maximum at about 14

HEMATOLOGIC EFFECTS OF RADIATION

1041

days; recovery (if the exposure was in the LD50 range) is usually com-
pleted in three to four weeks. With dosages above the LD50 the anemia
develops more precipitously and is more severe, and the recovery process
is delayed.

The immediate precursors of the erythrocyte are not significantly
reduced in number in the peripheral blood of rabbits (Jacobson and
Marks, 1947) and rats (Lawrence and Lawrence, 1930) with doses below
100 r. With doses above 100 r, reduction in reticulocytes becomes

0 10 20 30 40

TIME, days
O 220 r «260r A 310 r D 360 r • 420 r

Fig. 16-7. The effect of single doses of total-body roentgen irradiation on the red-cell
count, hemoglobin, and reticulocyte values of hybrid guinea pigs.

progressively more significant in all the common laboratory animals.
Doses in the LD50 range reduce the reticulocytes from a normal of 1-4
per cent to less than 0.1 per cent. Recovery after such exposure begins
at about the same time in the various species. As illustrated in Fig. 16-4,
a compensatory elevation above the normal range occurs as recovery of
erythropoietic tissue proceeds.

SINGLE EXPOSURE TO FAST NEUTRONS

Lawrence and Lawrence (1936) and Lawrence, Aebersold, and Lawrence
(1936) reported the first study on exposure of mammals to fast neutrons.

1042

RADIATION BIOLOGY

This work, which was performed on rats, indicated that the pattern of
hematologic effects was qualitatively similar to that produced by X
radiation. The observations of Lawrence have been extended to other
laboratory animals, including mice and rabbits (Lawrence, Aebersold, and
Lawrence, 1936; Henshaw et al., 1946; Jacobson and Marks, 1947).
Uranium-pile fast neutrons have been explored and compared with
cyclotron-produced fast neutrons (Zirkle, 1947). Henshaw exposed mice
to single doses of 26, 50, 78, 90, or 105 n. These studies paralleled

1 0,000

CO 5

10

15 20 25 3C

1

35

TIME

o o 26n

o — — o 55 n

FROM EXPOSURE, days

e e 68n o o 89n

o — — » 7fin « «> 97n

—- « I06n

• 128n

e> —

© 9n

9

Fig. 16-8. Effect of single doses of fast neutrons on the heterophil values of the periph-
eral blood of rabbits.

studies on rabbits exposed to cyclotron fast neutrons, in which dosages of
9, 26, 55, 68, 76, 89, 97, 106, or 128 n were used. No difference in the
hematologic effect of the LD5o dose of cyclotron or pile fast neutrons
appears to exist.

Figures 16-8, 16-9 show the effect of cyclotron fast neutrons on the
leukocyte values of rabbits, and Table 16-1 gives the comparative effect of
X rays and fast neutrons on the lymphocyte response in rabbits.

An initial increase in heterophils follows exposure to fast-neutron
dosages of 9-128 n, and within the first 24 hours there is also an initial
rise. This is followed, after dosages ranging from 26-128 n, by a reduc-
tion, reaching a minimum in 3-5 days. No significant reduction in
heterophils occurs after an exposure to 9 n. Eventual recovery to

HEMATOLOGIC EFFECTS OF RADIATION

1043

normal limits is observed in all groups exposed to total-body dosages
below 128 n.

The number of lymphocytes in the peripheral blood is reduced after
the administration of doses ranging from 9-128 n. This reduction is
apparent as early as 3 hours after exposure, with a maximum reduction at
2-3 days. Although the lymphocyte values of animals exposed to 9 n are
again within the normal control range by 5 days, the lymphocytes of those
animals exposed to 26 n and above remain below control values for more
than five weeks.

10,000

5000

2000

CO

>
o
o

i

CL

1000

500

200 -

100

C 0

10 15 20

TIME FROM EXPOSURE.

25
days

30

-• CONTROLS

-a 9n
-* 26n

®— — o 55n

e e 68n

a — — » 76n

o o 89n

35

-o 97n
-« I06n
-• I28n

Fig. 16-9. Effect of single doses of fast neutrons on the lymphocyte values of the
peripheral blood of rabbits.

A severe normochromic anemia is observed in animals exposed to a
fast-neutron LD5o- The maximum anemia, as is also observed after
exposure to a 30-day LD50 of X radiation, appears between the fourteenth
and sixteenth days after exposure. Recovery to normal values occurs by
23 days. Reticulocytes are reduced to 0.1 per cent or less by the third
day in animals exposed to doses of 128 fast-neutron units or above.

After doses of 80-89 and 100 108 n, a maximum reduction of approxi-
mately 72 per cent in the platelet values of the peripheral blood occurs
between the fifth and tenth day, with return to normal values by 15 and
23 days, respectively, in the two exposure levels.

An abortive rise in heterophils and lymphocytes occurs (between the

1044 RADIATION BIOLOGY

fourth and tenth day) after fast-neutron exposure and is comparable to
that observed following the exposure of rabbits to X rays.

The morphologic changes in the cellular constituents of the peripheral
blood after fast-neutron exposure are identical with those occurring in
the peripheral blood of rabbits after "comparable" doses of X rays.
These observations in rabbits are comparable with those after single
exposure to X rays (Figs. 16-1, 16-3). Henshaw's observations (1946) of
mice likewise are qualitatively similar to X-ray studies of these species
when the differential factor of effectiveness between X rays and fast
neutrons is considered.

COMPARISON OF EFFECTS PRODUCED
BY CYCLOTRON FAST NEUTRONS AND X RAYS

Sacher and Pearlman (1947) have made a statistical analysis of exten-
sive hematologic data obtained by Jacobson and Marks (1947) and by
Jacobson et al. (1947) from rabbits exposed to X rays and fast neutrons
and have derived a ratio of the relative effectiveness of these two radia-
tions. Using the data on the effect of fast-neutron and X-ray exposure on
the number of circulating heterophils and lymphocytes, they estimated
the X:n ratio to be of the order of 6.3. These estimates are in essential
agreement with the X:n ratio derived from survival data on rabbits
(using the same type of instruments) by Hagen and Zirkle (1950).
Evans (1948) derived a ratio of 8.1 in Swiss mice. It should be pointed
out again that the n unit is arbitrary and that the X : n ratio given does
not mean that neutrons are 6.3 times more effective than X rays in pro-
ducing the same biological effect.

FACTORS CONCERNING COAGULATION
FOLLOWING SINGLE EXPOSURES

A prolonged bleeding time, impaired clot retraction, fragility, thrombo-
cytopenia, and a prolonged whole-blood clotting time were observed in
laboratory animals following single exposures to dosages of penetrating
radiation in the LD60 range and above by Shouse, Warren, and Whipple
(1931), Allen and Jacobson (1947a), Allen et al. (1948), Cronkite (1950),
Cronkite et al. (1949, 1950), Prosser et al. (1947), and others. The only
cellular element in the circulating peripheral blood known to be concerned
with this problem is the platelet.

Platelets are, in general, found to be reduced in the peripheral blood of
all the various species of laboratory animals after exposure to dosages in
the range that reduces the polymorphonuclear cells. Qualitatively, the
megakaryocytes and granulocyte precursors are of approximately equal
sensitivity to irradiation injury. According to Lawrence, Dowdy, and
Valentine (1948) and Cohn (1952) the platelet values of the peripheral
blood of the rat are significantly reduced after whole-body exposure to

HEMATOLOGIC EFFECTS OF RADIATION

1045

300 r and above. Lorenz (1951) has demonstrated that a severe thrombo-
cytopenia occurs in guinea pigs after exposure to 200 r. Mice, according,
to Jacobson, Robson, and Marks (1950) and Jacobson, Simmons, Bethard,
et al. (1950), are approximately equal in sensitivity in this respect to rats.
With 600 r the maximum reduction occurs at about 9 days after irradia-
tion (Fig. 16-10). Swine (Cronkite et al., 1949) and dogs (Allen and
Jacobson, 1947a, b; Prosser et al., 1947) are roughly comparable with
guinea pigs in sensitivity. In rabbits (Jacobson et al., 1947; Jacobson,
Marks, and Lorenz, 1949), platelets are markedly reduced after exposure
to 500 or 800 r. Recovery of the platelets to normal values in all these
species occurs by about the fourteenth to twenty-first day after LD50

n -

10
9

Q>
O

\-
bJ

<

_J
CL

-

-^ />±

;r^x^v

^\>

0^

\

£

-

\

/
/

\

/

\

\
\

/
/

*

\

/

/

-

-

l i f i 1

i

0

3
TIME

6 9 12
AFTER X RAY,

19

23

15
days

• • CONTROLS o--o OPERATED CONTROLS

t-i600r WITH SPLEEN EXTERIORIZED
♦— • 1025 r WITH SPLEEN EXTERIORIZED

Fig. 16-10. Comparative effects of 600 and 1025 r of total-body roentgen irradiation
on the platelet values of CFi female mice.

exposure. With doses above the LD50 the recovery of platelet values is,
in general, a function of the dose.

The factors concerned with the hemorrhage appearing in experimental
animals exposed to penetrating radiation may be multiple. With doses in
the LD50 range or above in any of the species studied, a severe thrombo-
cytopenia occurs, but at this time it is not known with certainty whether
or not this single cellular reduction alone accounts for the widespread gross
and microscopic hemorrhage which occurs in many of these species but
which has been studied most extensively in the dog (Allen and Jacobson,
1947a, b; Allen, Moulder, and Enerson, 1951) and swine (Cronkite et al,
1949; Cronkite, 1950). Platelets are considered to have a number of
functions that are important in irradiation hemorrhage. Allen (1952)
lists these functions as follows: (1) platelet agglutination and adherence
to the site of vascular injury, (2) a source of thromboplastin, (3) an

1046 RADIATION BIOLOGY

accelerator of prothrombin conversion, (4) an effective antiheparin, and
(5) a contributor to the integrity of the capillary wall. In the absence of
platelets in animals exposed to massive doses of irradiation, hemorrhage-
gross or microscopic — may occur on the basis of failure of these functions.
The severity of hemorrhage will vary from animal to animal, however,
and local lesions such as ulcerations, abrasions, or systemic infection may
also affect the severity of the hemorrhagic manifestation. In general,
prothrombin studies of dogs (Allen et al., 1948) and rabbits (Jacobson,
Marks, Gaston, Allen, et al., 1948) subjected to total-body irradiation
in the LD5o range or above remained within normal limits except termi-
nally. It is generally agreed that the defects of increased bleeding time,
delayed clot retraction, and delayed prothrombin conversion result from
the thrombocytopenia. The delay in prothrombin conversion or con-
sumption can be corrected by addition of platelet-rich hemophilic dog
plasma but not by platelet-poor hemophilic dog plasma (Cronkite et al.,
1950). The increased clotting time observed in dogs (Allen and Jacob-
son, 1947a, b; Allen et al., 1948), goats and swine (Cronkite, 1950),
rabbits (Jacobson, Marks, Gaston, Allen, et al., 1948), and guinea pigs
(Haley and Harris, 1949) may be related, in part or entirely, to platelet
reduction and delayed prothrombin conversion. Other factors known to
be concerned with clotting, such as fibrinogen and calcium blood levels,
are normal in irradiated animals. Allen and Jacobson (1947a) and Allen
et al. (1948) originally reported finding evidence for a heparin-like circulat-
ing anticoagulant in dogs exposed to 450 r which could usually be restored
to normal by administration of protamine sulfate or toluidine blue.
More recently Allen has found that, although after irradiation an increase
in the whole-blood clotting time (Lee- White) is usually demonstrable in
the dog, no evidence for a circulating heparin-like anticoagulant can be
demonstrated unless, in addition, transfusions have been given and
transfusion reactions have occurred. He has repeatedly called attention
to the discrepancy that is not infrequently observed between the platelet
number in the circulating blood and the whole-blood clotting time. He
has reported the appearance of a prolonged whole-blood clotting time
before platelets were appreciably reduced (Fig. 16-11) and a return of
whole-blood clotting time to normal a week or more before evidence of
platelet recovery could be detected in the peripheral blood. Although he
attributes this abnormality in whole-blood clotting principally to physio-
logic disturbances associated with thrombocytopenia, he believes that
other disturbances in the hemoplastic mechanism cannot be excluded at
this time. The role of increased capillary fragility in the hemorrhagic
syndrome induced by irradiation is probably of major importance. The
relation of thrombocytopenia in the causation of increased capillary
fragility is not understood nor has it been adequately explored. The fre-
quent association of petechial hemorrhage with thrombocytopenic states

HEMATOLOGIC EFFECTS OF RADIATION

1047

tends to implicate the platelet in preservation of the integrity of the
capillary wall. On the other hand, increased capillary fragility and
petechial hemorrhage are observed in a number of toxic states, including
severe systemic infection, even though platelet values of the peripheral
blood are normal. The importance of the widespread microscopic hemor-
rhage that occurs presumably secondary to direct or indirect injury to the
capillary wall has recently been emphasized by Furth et al. (1951) . Even
in the absence of gross hemorrhage a more severe anemia occurs after
whole-body exposure to dosages above the LD5o than can be accounted for

en o
IF

00

-

90

450 r

80

-

70

-

60

50

-

40

-

30
20

\
\

10

1

! i i : i i "° i

2 4 6 8 10 12 14

TIME AFTER X RAY. days
'CLOTTING TIME o o PLATELETS

16

Fig. 16-11. Effect of a 450-r single dose of total-body roentgen irradiation on the
platelet values and clotting time (Lee- White) of 25 dogs. {Allen et al., 1948.)

by cessation of erythropoiesis alone. Furth explains this discrepancy on
the basis of increased microscopic hemorrhage resulting from "leaking"
through the capillary walls. No histologic abnormalities have been
demonstrated that account for this loss of vascular integrity.

HEMATOLOGIC EFFECTS OF CHRONIC TOTAL-BODY EXPOSURE

TO EXTERNAL RADIATIONS

The penetrating radiations to which laboratory animals have been
exposed chronically (repeated or continuous exposures) include X rays,
7 rays (radium and pile gammas), fast neutrons (cyclotron and pile-
produced), and slow neutrons. Survival, carcinogenesis, and other
biological effects have been studied in several species of mammals after
exposure to these radiations. Adequate hematologic studies have been

1048 RADIATION BIOLOGY

made in animals exposed chronically to 7 rays, fast neutrons, and X rays.
The work of Lorenz, Eschenbrenner, et al. (1946), Lorenz, Heston, et al.
(1946), Lorenz, Uphoff, and Sutton (1949), and Lorenz (1951) on 7-ray
exposure has been extensive and exceptionally well controlled. Evans
(1948) has studied the effects of fast neutrons and, to a lesser extent, the
effect of X rays on the hematopoietic system.

CHRONIC EXPOSURE TO X RAYS AND FAST NEUTRONS

For purposes of comparison of the biological effectiveness of X rays and
fast neutrons, Evans (1948) exposed groups of Swiss mice to daily doses
of 80 r (X rays) and 10 "N" (neutron units arbitrary). These doses
were chosen as the basis for comparing the median lethal doses of the two
radiations that gave a ratio (in effectiveness) of 8.1 r = IN. The radia-
tions were given daily for 25 days, and the accumulated dosages were
2000 r and 250 N. The hematologic responses were similar in both
groups. The fall in the leukocyte values of both groups was comparable
through the 30 days of observation at which time the values were below
1000/cu mm.

The erythrocyte and hemoglobin values of the fast-neutron and X-ray
exposure groups remained within control limits for about one week and
thereafter fell steadily, reaching levels of approximately 2 million and
3-4 g, respectively, on the twenty-eighth day. On these findings and
other data including survival and histopathologic examination of post-
mortem material, Evans concluded that the effects of these two radiations
at this dosage level were qualitatively and quantitatively similar.

CHRONIC EXPOSURE TO REPEATED SMALL DOSES OF FAST NEUTRONS

In another experiment, Evans (1948) exposed groups of Swiss and
CFi male and female mice to dosages of 0.014, 0.07, 0.14, and 1.4 N/day.
These dosages, according to his calculations, are equivalent biologically
to a value of 1 N = 35 r. Exposure of the Swiss mice was continued for
ninety weeks and of the CFi mice for eighty-three weeks.

Although the mean leukocyte values of all exposure groups were con-
sistently lower than those of the control groups, only in the 1.4 N/day
exposure group were significant effects observed in the peripheral blood.
In this latter group, reduction in the leukocyte values was apparent
within four weeks after beginning of exposure, and a reduction in the
mean count of about 50 per cent was apparent at twenty-four weeks when
the exposure was discontinued. This leukocyte reduction was largely
due to the lymphocyte reduction; no effect on erythrocyte, hemoglobin,
reticulocyte, or platelet values was noted during the period of exposure
of any of the four groups.

Suter (1947) and Ingram and Mason (1950b) studied the hematologic
effect of chronic X irradiation in dogs, rabbits, rats, and monkeys. With

HEMATOLOGIC EFFECTS OF RADIATION 1049

10 r of X ray per day the erythrocyte values of the dog and rabbit were
reduced by sixteen weeks, whereas it required thirty-two weeks for
erythrocyte values to decrease in the rat. The same number of roentgens
per day was also required to bring about a depression of the leukocyte,
absolute heterophil, and platelet values in the rabbit by four, five, and
twelve weeks, respectively, whereas a daily dose of only 6 r/day produced
depressions of these elements in the dog by sixteen and eight weeks.
The leukocyte and absolute lymphocyte values of the rat decreased in
four weeks with 10 r/day, but the same elements were decreased in
monkeys by one to two weeks by the same dose.

Suter (1947) also studied the effects of chronic exposure to 10.2 n/week
on the peripheral blood of the dog, rabbit, and rat. A decrease in the
erythrocyte values was produced by thirty-eight, thirty-two, and six
weeks, respectively. The leukocytes and absolute neutrophils were
reduced in the dog by 2%, weeks and the absolute lymphocytes by 4>^
weeks. These same elements were decreased in the rabbit by 5, 73^, and
2}4 weeks, respectively, whereas in the rat only l}4, %, and 1% weeks,
respectively, were required to bring about a reduction with 10.2 n/week.

CHRONIC EXPOSURE TO SMALL DAILY DOSES OF FAST NEUTRONS

Fast-neutron doses of 4.3 n/day, 6 days per week, produced a dramatic
and uniform decrease in leukocyte levels in CFi female mice, involving
both the lymphocytes and the heterophils (Henshaw, Riley, and Staple-
ton, 1947). Death occurred at eight to twelve weeks. In another group,
daily exposures of 1.15 n were compared with 8.6 r of pile-produced y rays
(Henshaw, Riley, and Stapleton, 1947). The roentgen-to-neutron ratio
of 1:7.5 for the acute killing of mice was the criterion used for selecting
these dosages. Only slight hematologic changes, if any, were produced
with these doses after forty to sixty weeks of exposure. Henshaw
reported, however, that a daily exposure to 0.25 n of fast neutrons or to
8.6 r of pile 7 rays had the same effect on survival of mice, and thus an
r:n ratio of 1:35 for daily doses was calculated for these radiations.
This investigator is of the opinion that threshold responses of the periph-
eral blood are at least ten times less sensitive than survival. Mice (C-58)
treated with single weekly doses of 51.6 r (X rays) showed slightly lower
leukocyte values, with a terminal drop at thirty weeks, than those
exposed to the equivalent of 8.6 r of 7 rays per day, 6 days per week. A
significant, gradual, and progressive decline in erythrocyte values was
apparent with 51.6 r/week (X rays), whereas 8.6 r/day (7 rays) produced
only a slight and minimal reduction of the erythrocyte values.

CHRONIC EXPOSURE TO GAMMA RAYS (RADIUM)

The general plan of exposure employed by Lorenz, Heston, et al. (1946)
and Lorenz, Heston, and Eschenbrenner (1947) is referred to here only to

1050 RADIATION BIOLOGY

orient the reader; details may be found elsewhere. The experimental
animals were placed in cages arranged about a centrally located radium
source. The cages were placed at varying distances from the 2-g radium
source so that given daily doses were delivered during an 8- or a 24-hour
period of exposure. In Lorenz's original experiments (1946), continuous
exposure (24 hours per day) was used. The data from these early experi-
ments are available. Since more complete studies have been done using
8-hour daily exposures, only these later experiments will be considered.

Three species were studied: (1) mice, LAFX genetically homogeneous
hybrids, strain A, C-57 black, and C3H, (2) rabbits, crosses of Dutch and
American Blue, and (3) guinea pigs, genetically heterogeneous hybrids
and inbred strains.

Groups of the three species of animals were exposed to doses of 0.11,
1.1, 2.2, 4.4, or 8.8 r for 8 hours per day for periods extending in some
experiments to more than three years.

Observations on Mice. Male and female LAFi mice received total
dosages as high as 5880 r in the 8.8-r exposure group and correspondingly
lower at the other exposure levels (Table 16-3).

The blood data indicate considerable radioresistance of the hemato-
poietic system. There was a decrease in all cellular hematologic con-
stituents in the peripheral blood of animals exposed to 8.8 r daily. The
mean reduction in erythrocyte, hemoglobin, and platelet values was
relatively minimal and was not more than 30 per cent at any time. This
reduction occurred slowly over a period of seventy-nine weeks. Indi-
vidual animals, however, did develop severe anemia. Only a question-
able reduction in the erythrocyte or hemoglobin and platelet values
occurred in the groups exposed to 4.4 r. No anemia or thrombocytopenia
was observed in the groups exposed to 2.2 r and below.

The mean leukocyte count was reduced in the 8.8-, 4.4-, and 2.2-r
groups, but this was a reflection of the lymphocyte reduction. The
heterophils remained within control limits. In general, female mice
were more radiosensitive than males as far as hematologic findings were
concerned.

Data available on C3H and C3Hb females and strain A males show that
these strains are all comparable to LAFi in respect to their hematologic
response to this chronically administered radiation.

Observations on Female Rabbits. Evidence of irradiation damage to
the hematopoietic system was slight even though the highest accumulated
dose was approximately 12,000 r. During the first 100 weeks of exposure
the total white count of all experimental groups was lowered. The
lymphocyte reduction is evident and is made more pronounced by a
concomitant relative increase in heterophil values in all exposure groups
except those at the 0.1 1-r level. As was observed in mice and guinea
pigs, these changes became apparent within the first few weeks after the

HEMATOLOGIC EFFECTS OF RADIATION

1051

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1052

RADIATION BIOLOGY

exposure was begun and remained fairly constant at this level for the
first 100 weeks. No significant effect on erythrocyte and hemoglobin
values was observed in any of the exposure groups.

Observations on Guinea Pigs. As is indicated in Table 16-4, the expo-
sure levels for this species were the same as for mice and rabbits. Within
a few weeks after the beginning of exposure a decrease in the leukocyte
values of the peripheral blood occurred in both males and females of the
five experimental groups, and the depression persisted throughout the
experiment (Fig. 16-12). The leukocyte depression was significant
(P 5i 0.05) even in the group exposed to 0.11 r. The degree of

Table 16-4. Survival of Hybrid Guinea Pigs Exposed to Gamma Radiation-
Daily (8 Hours)0

Daily
dose, r

Number

of
animals

Mean age

at start,

days

Mean survival

time since

beginning of

exposure, days

Mean
accumu-
lated
dose, r

Duration
of experi-
ment,
days

Largest
accumu-
lated
dose, r

0

24

151

1372 ± 95

2246

0.11

17

137

1457 ± 129

180

2280

300

1.1

17

196

1224 + 119

1380

1993

2200

2.2

18

197

978 ± 63

2170

1410

3100

4.4

18

185

653 ± 83

2900

1467

6500

8.8

18

185

187 ± 27

1670

502

4400

8.86

6

199

1192 ± 142

1527

270

8.8C

6

230

968 ± 124

....

1259

540

° Acute exposure groups included.
6 Exposed for 31 days.
c Exposed for 61 days.

leukocyte depression was correspondingly greater with the higher daily
exposure.

The reduction in the leukocyte values of the various experimental
groups was caused by a decrease in lymphocytes in the 0.11- and 1.1-r
groups. In the 2.2-, 4.4-, and 8.8-r groups it was caused in part by a
decrease in lymphocytes but was largely a result of the more significant
reduction in heterophils. In fact, the characteristic terminal blood
picture is one of profound neutropenia, the proportion of heterophils
often being as low as 1-5 per cent. No significant alteration in the
eosinophil or monocyte values was observed in these experiments. In
general, there were likewise no significant morphologic changes in the
leukocytes in any of these exposure groups.

No anemia of significance was produced in male or female guinea pigs
exposed chronically to 0.11 and 1.1 r. The male guinea pigs exposed to

HEMATOLOGIC EFFECTS OF RADIATION

1053

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1054 RADIATION BIOLOGY

2.2 r, however, developed a normochromic anemia of only moderate
significance beginning approximately seventy-nine weeks after exposure.
Within a period of forty-nine weeks both the male and female guinea pigs
of the 4.4-r group developed a moderate normochromic anemia that
gradually increased in severity. A reduction of about 25 per cent in
hemoglobin and erythrocyte values occurred. Females exposed to 2.2
or 4.4 r developed a lesser anemia than the males at these exposure levels,
but a severe terminal anemia occurred in most of the animals, both males
and females, in these two exposure levels. Within nineteen weeks,
female and male guinea pigs exposed to 8.8 r developed an anemia that
increased in severity through sixty and sixty-nine weeks, respectively,
and all animals died with a terminal anemia.

The platelets per cubic millimeter in both sexes exposed to 0.11 r daily
(8 hours) remained comparable to those of the control animals throughout
the experimental period. Female guinea pigs at the 1.1-r level had a
normal mean platelet value when first sampled at forty-nine weeks after
exposure began: the value was reduced slowly, reaching a mean slightly
below control levels that was maintained throughout the remainder of the
experiment. The platelet values of the male guinea pigs at this same
level, however, fell slowly between the forty-ninth and seventy-ninth
weeks, recovered by the hundredth week, and remained within normal
limits thereafter.

In male and female guinea pigs exposed to 2.2, 4.4, and 8.8 r daily
(8 hours), the reduction in platelet values was progressive, the degree
of reduction roughly corresponding to the daily dose. The terminal
platelet counts for individual animals were as low as 20,000/cu mm and,
in many cases, lower. All animals exposed at the rate of 8.8 r daily
(8 hours) died with anemia or thrombocytopenia, or both. The majority
of the animals exposed to 4.4 and 2.2 r daily (8 hours) likewise died of
the same disorders.

Chronic exposure to the three dose rates of 8.8, 4.4, and 2.2 r daily (8
hours) is sufficient to produce a terminal anemia or thrombocytopenia, or
both, in the guinea pig. Although Lorenz, Heston, et at. (1946) state
that it is difficult to decide from the data whether or not dose rate is more
important than total accumulated dose in the induction of the severe
terminal pancytopenia, they are inclined to believe that the dose rate is
the more important factor. This opinion is based on the fact that no
terminal anemia has been observed in the 1.1-r group even though the
maximum accumulated dose was approximately 2200 r. This dose was
sufficient to induce terminal anemia in the 8.8-, 4.4-, and 2.2-r groups.
It seems that in the 1.1-r group the injury is compensated for by repair
processes, but it is possible that in species with a similar sensitivity of
the hematopoietic system, but with a considerably longer life span,
injury would eventually overcome the repair process.

HEMATOLOGIC EFFECTS OF RADIATION

1055

PANCYTOPENIA INDUCED BY CHRONIC EXPOSURE
TO GAMMA RADIATION

In a series of experiments Lorenz (1951), Lorenz, Jacobson, and Sutton
(1950), and Lorenz et al. (1950) studied the hematologic recovery pattern
of guinea pigs exposed to 8.8 r (8 hours per day) until an anemia of ca. 2.8
million erythrocytes/cu mm had appeared. Leukopenia and thrombo-
cytopenia were also invariably present when anemia reached this stage.
The dose required to produce an anemia of this magnitude varied from
950-1047 r. After removal of the animals from the radiation field, three
distinct hematologic recovery patterns were apparent. These three
patterns will be designated as groups a, b, and c (Fig. 16-13).

t/i.

SI

o 2

^■■°-v-°-^-a''o.^' ■ta.-o'-'S

J L

J I L

J I L

-20 -10 0 10 20 30 40 50 60 70 80 90
TIME, weeks

110

130

• • CONTROL x x GROUP A, WHICH RECEIVED A TOTAL

ACCUMULATED DOSE OF I047±275r
o -o GROUP B, 9701 130 r o -a GROUP C, 950±59r

Fig. 16-13. Effect of limited chronic y irradiation at 8.8 r daily (8 hours) on the red-
cell count of inbred guinea pigs (family 2). (Lorenz, 1951.)

Group a. The erythrocyte, hemoglobin, and hematocrit values of the
animals with recovery pattern a continued to decrease, and no recovery
was apparent, or slight unsustained recovery was observed between six
and nine weeks after the animals were removed from the exposure. The
platelet values fell no lower after removal of the animals from the radia-
tion field and, in fact, rose slowly. The leukocyte values increased
rapidly after termination of the exposure. The rise in leukocytes repre-
sented a rise in both lymphocytes and heterophils. Contrary to the
complete disappearance of reticulocytes following an acute X-ray expo-
sure or a 7-ray exposure of this magnitude, no reduction of reticulocytes
occurred in these three groups ; in fact, a rapid increase above the control
value appeared after termination of the exposure. All animals in group
a died within approximately nine weeks after removal from the radiation
field.

Group b. The hematologic findings in the animals with recovery
pattern b, during the first nine weeks after termination of the exposure,
were approximately the same as the a pattern. After that time the

1056 RADIATION BIOLOGY

recovery processes continued for a period of several weeks. In a few
animals the increase in leukocytes approached normal, but on an average
it amounted to approximately one-half normal. Beginning at approxi-
mately the twelfth week and continuing to the twenty-fourth week, one
animal after another developed a recurrent pancytopenia and died.
There was a persistent increase in the percentage of circulating reticulo-
cytes, and the color index also increased. Hemorrhage appeared to play
no important role in the terminal picture since gross or microscopic evi-
dence of hemorrhage was minimal or absent.

Group c. The third pattern, c, was one of complete recovery. The
initial postirradiation hematologic findings were identical with those of
the other two patterns. Recovery from the anemia began approximately
six weeks after the termination of the exposure. A rapid increase of all
cellular elements during the subsequent six weeks was followed by a
gradual approach to normal. This was achieved most rapidly by the
heterophils and was delayed longest in the platelets. However, full
recovery took place over a period of many months. It is of interest to
note that, although the increased red-cell diameter (macrocytosis) per-
sisted for about sixty weeks, reticulocytes returned to normal after
approximately ten weeks.

The histologic examination of animals that died in the three groups
may be summarized as follows: The bone marrow of the animals of
group a showed a slight-to-considerable decrease in cellularity with an
approximately normal proportion of the different cellular components.
In the animals that died in this group and in those of group b that died
earliest, the degree of cellularity of the bone marrow was normal or nearly
normal or even hyperplastic, but the proportion of the various cellular
components was not normal. In the animals of group c the bone marrow
was comparable to that of nonirradiated controls. Hemorrhage played
no significant role in the development of the terminal anemia in these
three groups of animals since gross or microscopic evidence of hemorrhage
was minimal or absent.

HEMATOLOGIC EFFECTS OF EXPOSURE
TO INTERNALLY ORIGINATING RADIATION (RADIOELEMENTS)

So far as the peripheral blood is concerned, the sensitivity and rapidity
of the effect on blood-forming tissue of the radioelements and subsequent
reflection in the hematologic picture depend on the dosage and the extent
of localization within hematopoietic tissue. With all the radioisotopes
studied by Jacobson, Marks, and Lorenz (1949), whether they localized
in bone or more generally throughout the body, a reduction in the
lymphocyte values of the peripheral blood was found to be the most
sensitive indicator of acute or subacute effect. This was also true for

HEMATOLOGIC EFFECTS OF RADIATION

1057

radioelements such as radioiodine, which localizes promptly in the thy-
roid gland.

PHOSPHORUS (P32)

Jacobson, Skirmont, et al. (1948) studied the effect of a single intra-
peritoneal injection of 1 juc/g of body weight of P32 on normal and poly-
cythemic rats. A significant depression of leukocyte values per cubic
millimeter was produced in both groups. No apparent effect was pro-
duced on the hemoglobin, erythrocyte, or hematocrit values in either
group. On the basis of hematologic data, no apparent difference in the
sensitivity of the hematopoietic system to a single dose of P32 (1 juc/g) or
to 300 r of total-body X irradiation was evident in rats (Fig. 16-14).

90

80

70

£ 30

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COBALT
ADMINISTRATION,

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P32 ONLY

X RAY ONLY

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16

24 32 40 48 56 64
TIME AFTER TREATMENT, days

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72

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DAYS BEFORE

TREATMENT

Fig. 16-14. The effect of 300 r of total-body X irradiation or a single dose of P32 (1
fic/g of body weight) on the hematocrit values of normal rats and those having cobalt-
induced polycythemia.

However, after a total dose of 4.5 /xc/g of body weight given intraperi-
toneal^ in 1.5-Mc/g doses every three weeks, Bethard, Skirmont, and
Jacobson (1950) reported a decrease in hematocrit and hemoglobin values
by the twenty-second week, which reached a low level by twenty-eight

weeks.

Warren and Dixon (1949) studied the hematologic effect of radiophos-
phorus on developing chicks (6-28 days). In 6-day-old chicks, given
subcutaneously 235 nc of phosphorus, a rapid decrease in lymphocytes,
reaching less than 1000/cu mm in 4 days, occurred followed by a gradual
increase to normal values by 55 days. The erythrocytes fell gradually
the first 8 days, dropped sharply to 0.63 million/cu mm by 15 days, and

1058 RADIATION BIOLOGY

returned to normal value in 41 days. The leukocyte values fell rapidly
during the first 4 days and then gradually disappeared completely from
the blood in 15 days, recovering slowly and only approaching normal
values at the end of 48 days. The same general pattern of peripheral-
blood changes was found by Warren following administration of 300 /ic to
19-day-old chicks and 760 /zc to 28-day-old chicks. The larger doses
caused a fatal anemia.

Dixon (1948) injected chickens subcutaneously with 300 or 400 mc of
P32 every week for three weeks. A virtual agranulocytosis and thrombo-
cytopenia developed by 7 days and remained thus during the period of
observation (17 days). Erythrocyte values decreased steadily reaching
0.5 million/cu mm by 17 days.

Raper and Barnes (1951), Raper, Zirkle, and Barnes (1951), and Raper,
Henshaw, and Snider (1951) studied total-surface radiation with (5 rays
in rabbits. With doses ranging from 1500-59,000 rep, no direct effect on
the cellular constituents of the peripheral blood was noted. Sporadic
increase in the heterophils began three weeks after irradiation with
10,000-15,000 rep, but no evidence of individual effect on the leukocyte
values was detected, nor was there evidence to substantiate the claim of
lymphocyte stimulation by superficial irradiation. The lack of effect is
considered to be consistent with physical properties of radiation and
anatomical features of the rabbit.

SODIUM (Na24)

A moderate-to-severe leukopenia and lymphopenia followed injection
of radiosodium (Na24) given intraperitoneally to mice in doses of 23-96
juc/g of body weight. With doses of 48-95 /xc/g of body weight, death of
the animals occurred by 8 days (Jacobson and Simmons, 1946a).

BARIUM-LANTHANUM (Ba-La14°)

A dose of 1.9 /xc/g of body weight administered intraperitoneally to
mice produced a significant and rapid reduction of leukocyte, lymphocyte,
and heterophil values that was sustained for approximately 60 days.
Rapid reduction in these values and death within 15 days followed the
administration of 17 juc/g of body weight. No significant change
occurred in erythrocyte or hemoglobin values by 60 days with these
doses (Jacobson, 1946). The radiation effects of these two isotopes in
the mouse may be entirely due to /3 rays (Finkle, Snyder, and Tompkins,
1946), although both 0 and 7 rays are emitted.

YTTRIUM (Y91)

Yttrium (Y91) administered by gavage to rats in a single dose of
20 Mc/g of body weight produced a marked initial elevation in the erythro-

HEMATOLOGIC EFFECTS OF RADIATION 1059

cyte and hemoglobin values followed by a severe anemia that reached
its maximum at 20 days. A 75 per cent reduction in lymphocyte values
occurred within 3 days with a return to normal values by 35 days. A dose
of 10 fic/g of body weight produced a temporary lymphopenia, whereas
no hematologic effect was observed after single doses of less than 10 /xc/g
of body weight. Daily ingestion of 0.3-2.0 /xc/g of body weight of
yttrium produced a lymphopenia that reached a maximum at 90 days.
There was prompt return to normal values upon discontinuation of the
yttrium (Jacobson and Simmons, 1946b).

STRONTIUM (Sr89)

Hematologic studies of the peripheral blood of rabbits, rats, and mice
after intraperitoneal or stomach-tube administration of strontium (Sr89)
as a chloride in doses from 0.015-14.5 /xc/g of body weight indicated that
reduction in heterophil and lymphocyte values was about equal and that
these cells were sensitive indicators of acute and subacute effects. No
anemia occurred in mice following doses of 0.034 /xc/g of body weight or
lower, but moderate anemia was produced with a dose of 2.0 /xc/g of
body weight in this species. The minimal dose exerting a detectable
reduction in heterophil values in rats and mice was 0.22 and 0.25 /xc/g of
body weight. In rabbits an intraperitoneal dose of 1.0 /xc/g of body
weight produced a significant reduction of heterophil values, and 3.0 /xc/g
of body weight produced a significant reduction in hemoglobin and
erythrocyte values (Simmons and Jacobson, 1946).

Of interest in connection with strontium (Sr89) toxicity is the fact that,
in mice injected with a dose of 2.0 /xc/g of body weight, only a leukopenia
became apparent even though the bone marrow was markedly depleted
or aplastic (Jacobson, Simmons, and Block, 1949). Concomitant with
the development of aplasia in the marrow a myeloid metaplasia developed
in the spleen, which was sufficient to compensate for the lack of erythro-
cyte production in the marrow. In splenectomized mice a severe
anemia developed since compensatory blood formation in the spleen was
not possible (Fig. 16-15).

RADIUM

Hematologic studies on the peripheral blood of rats, mice, and rabbits
after intraperitoneal, intravenous, or intracardial injections of radium
chloride indicate that a macrocytic anemia develops following parenteral
administration of radium chloride in doses between 0.1 and 0.2 /xg.
Although doses below this range produced an anemia in rats and mice, no
anemia was produced in the rabbits with this dose. A reduction in
leukocyte values occurred with doses in the 0.1-/xg range in the rabbit,
whereas doses as high as 0.02-0.03 /xg resulted in leukocyte reductions in
rats and mice (Jacobson and Simmons, 1946c).

1060

RADIATION BIOLOGY

PLUTONIUM (Pu239)

There appeared to be no significant species difference in the sensitivity
of the hematopoietic system of the mouse, rat, or rabbit to plutonium
given intravenously or intramuscularly as a citrate in doses of 0.001-
0.119 Mc/g of body weight to rabbits and in doses of 0.003-0.125 /zc/g of
body weight to the rat. Doses of 0.0062 /xc/g of body weight produced
a moderate-to-severe and sustained leukopenia in all three species and
caused an anemia in mice and rats resulting in early death in these two

0 7 28 49 70 91

TIME AFTER INJECTION, days
t>— -a SPLENECTOMY PLUS Sr89 (2.0 fic/q)
a— ^SPLENECTOMY ALONE
o oSr89 ALONE (2.0^c/g)

Fig. 16-15. Erythrocyte values of normal and splenectomized rats injected with a
single dose of Sr89, 2.0 /ic/g.

species (135 days). Doses above 0.0063 jxc/g of body weight produced
a correspondingly more severe and sustained anemia, leukopenia, and
reticulocyte and platelet reduction in mice and rats (Jacobson and
Simmons, 1946c).

The fact that effects on the hematopoietic system are seen in the
peripheral blood of plutonium-injected animals whereas comparable
doses of injected radium produce no such changes and yet give rise to
bone tumors earlier and in greater number may be related to difference in
the site of deposition of the two elements.

GOLD (Au198)

Wheeler, Jackson, and Hahn (1951) studied the hematologic effect of
radiogold in 14 dogs receiving intravenously 1 ^c/kg. All dogs showed

HEMATOLOGIC EFFECTS OF RADIATION 1001

a decrease in leukocyte values, but only one had a marked leukopenia.
In most dogs the sedimentation rate was increased. Liver function
tests were essentially negative. The hemoglobin and hematocrit values
decreased over a period of several weeks.

GALLIUM (Ga72)

Dudley, Louviere, and Shaw (1951) administered radiogallium as a
citrate to five species of animals. The only effect of radiation observed
was a reduction in total leukocytes in the mouse, rabbit, and guinea pig
with dosages of 2.3-9.0 mc of Ga72/kg. The degree and duration of the
leukopenia vary with the dose and species. Repeated injections of
Ga72 in dogs and rabbits indicated that this element is a cumulative
poison for these species.

MORPHOLOGIC CHANGES IN PERIPHERAL-BLOOD CELLS
PRODUCED BY IONIZING RADIATIONS

Morphologic changes in the various cellular constituents of the periph-
eral blood of animals, which are observed after exposure to penetrating
radiation such as X ray, fast neutrons, and radioactive elements, have
been described by a number of authors. These morphologic alterations
are not specific for the various types of radiation. In fact, all the morpho-
logic abnormalities that have thus far been described can be produced by
such radiomimetic substances as the nitrogen mustards. Exposure to
repeated small dosages of X rays or fast neutrons produced little or no
morphologic change except macrocytosis and, occasionally, the appear-
ance of giant platelets. Whether or not an anemia develops in mammals
exposed to ionizing radiation given in repeated dosages depends on the
sensitivity of the species and the size of the dose. In the event that
anemia develops, then all the characteristic morphologic changes are
seen in the red cells, e.g., anisocytosis, poikilocytosis, and macrocytosis.
The observations of Lorenz, Heston, et al. (1946) on mice, guinea pigs,
and rabbits exposed to dosages as high as 8.8 r/day until the accumulated
dose was 5000 r or more showed surprisingly little evidence of morpho-
logic change and form the basis for this statement.

Exposure to a single dose of externally applied radiation, such as X ray
or fast neutrons, or irradiation from a single massive dose of an isotope
that localizes in the blood-forming tissue produced morphologic changes
that were detectable within a few hours after the exposure. These
changes, which have been described by Aubertin and Beaujard (1908),
Heineke (1904), Kroemeke (1926), Henshaw (1943-1944a, b), Evans
(1948), Dunlap (1942), Clarkson, Mayneord, and Parson (1938), Jacob-
son and Marks (1947), Jacobson et al. (1947), and Jacobson, Marks, and
Lorenz (1949), are briefly summarized.

1062 RADIATION BIOLOGY

In general, the number of morphologic abnormalities observed are
directly proportionate to the radiation dosage sustained by the blood-
forming tissue. There is little evidence that cells are injured in the
peripheral blood (Furth et al., 1951). The injury that results in the
observed abnormalities in the peripheral blood is undoubtedly sustained
or produced by the irradiation effects on the various maturation stages of
the precursors in the blood-forming tissues. With doses in the LD50
range or above, an anemia was invariably produced in all the species of
laboratory animals that have been investigated. In the presence of an
anemia, anisocytosis, poikilocytosis, macrocytosis, microcytosis, and
polychromasia were found. The degree of these red-cell abnormalities
depends, in general, on the type and chronicity of the anemia.

Nucleated erythrocytes were observed soon after exposure but reached
a maximum between the tenth and twenty-fifth day after irradiation.
This represents the period of maximum anemia in the peripheral
blood and the stage of maximum regeneration of erythropoiesis in the
marrow.

Degenerative morphologic changes in the platelets were found by
Jacobson, Marks, and Lorenz (1949) in the peripheral blood of animals
following median lethal doses and above of roentgen radiation. Giant
platelets were frequently seen concomitant with reduction in platelet
values. Except for increased size and distinctive granular degeneration,
no further morphologic alterations were found in the platelets of the
peripheral blood. Only on rare occasions were megakaryocytes found in
the peripheral blood after total-body roentgen irradiation in the LD50
range.

The morphologic changes in the lymphocytes of the peripheral blood
increased with increasing amounts of radiation. Clumping of nuclear
chromatin was seen early. Histologic studies indicate that this change,
which occurred with relatively small doses (~50 r), may actually be
reversible in some lymphocytes (Jacobson, Marks, and Lorenz, 1949).
As it became more pronounced, pale bluish opaque areas were encount-
ered. In addition, lymphocytes were seen with rounded chromatin
masses seemingly free within the nuclear membrane. At the time of
maximum lymphocyte reduction in the peripheral blood after irradiation,
mononuclear cells with a somewhat basophilic cytoplasm and with
nucleoli were often prominent. These latter cells are indistinguishable
from "blast" forms. Occasionally mitotic blast cells appeared after
semilethal or higher doses. Other lymphocytes appeared to have split
nuclei and nuclei with considerable fragmentation. With the LD5o range
of acute total-body irradiation, these destructive changes were more pro-
nounced. Remnants of nuclear fragments, without nuclear outline and
which could not be distinguished as belonging to either the myeloid or
lymphoid series, appeared. Many lymphocytes were bilobed, whereas

HEMATOLOGIC EFFECTS OF RADIATION 1063

others were difficult to distinguish from monocytes. During the peak
of lymphocyte destruction, phagocytic monocytes with engulfed nuclear
debris were found in the peripheral-blood smears. Similar changes were
described by Henshaw et al. (1946) in mice following large single lethal
doses of 7 rays.

During the phase of rapid reduction of granulocyte values in rabbits,
abnormal forms of this series were seen in the peripheral blood. These
cells showed nuclear deformity and disintegration to the extent that it
was impossible to determine at which stage of maturity the cells were
damaged (Jacobson and Marks, 1947; Jacobson et al., 1947; and Jacob-
son, Marks, and Lorenz, 1949). The nuclear membrane was destroyed,
and in most, instances, bluish nonspecific granules of uneven contour
filled the entire cell, covering the nucleus and cytoplasm to such an
extent that it was impossible to identify the cell other than to say it was
a damaged or degenerating leukocyte. Other bizarre forms were also
seen that were difficult to describe and impossible to categorize. The
cytoplasm of the heterophil may undergo changes following irradiation.
Occasionally it contained vacuoles and had a mottled appearance. In
some instances the granules of the heterophils appeared to be damaged
since cells were found in which some granules clung to the nuclear mem-
brane while the remainder of the cytoplasm remained clear. Another
type of cell seen in rabbits was one that resembled a basket cell but dif-
fered from it in having one or two distinct chromatin masses that were
deep blue against a network of destroyed perichromatin and cytoplasmic
disintegration.

Atypical eosinophils characterized by irregularity in the granules,
both with regard to size and shape, were occasionally seen in the blood
smears of dogs following LD5o or higher doses of total-body roentgen
irradiation. Giant polymorphonuclear leukocytes were frequently seen
which appeared swollen and contained multilobed nuclei spread over the
cytoplasm-like globules, having an extremely fine filament branching
into eight or more lobes.

DIRECT VERSUS INDIRECT EFFECT OF IRRADIATION
ON BLOOD FORMATION

Lawrence, Valentine, and Dowdy (1948) reviewed the problem of
indirect effects and concluded that the hematologic or histologic evidence
for indirect effects of irradiation on nonirradiated blood-forming tissue
was unconvincing. Jacobson et al. (1949, 1950) found no suppression
of lymphopoiesis in the lead-shielded appendixes of rabbits given 800 r
of total-body X irradiation. Exposure of rabbits to this dose with shield-
ing of only a single Peyer's patch of the intestine did not appear to
inhibit or destroy the lymphocytes in the patch (Jacobson et al., 1952).
It must be borne in mind that dosages above the LD5o may produce

1064 RADIATION BIOLOGY

changes in the hematopoietic tissue of the shielded body part if the
general toxicity produced by the irradiation is severe.

"STIMULATION" OF BLOOD-FORMING TISSUE BY RADIATION

The evidence that small or large doses of penetrating radiation stimu-
late blood formation directly is unconvincing (Bloom, 1948). On the
other hand, secondary or "compensatory" increases in certain of the
cellular constituents of the peripheral blood and in the blood-forming
tissue have been observed repeatedly. Such compensatory "stimula-
tion " is invariably preceded by a reduction of certain cellular constituents
of the peripheral blood and destruction or inhibition of cells within the
blood-forming tissue (Bloom, 1948; Jacobson, Marks, and Lorenz, 1949;
Murphy, 1926). These findings are based on histologic or hematologic
observations and are to be differentiated from chemical or metabolic
changes within cells that may occur after irradiation and that might be
labeled "stimulation." This is also true when the problem of indirect
effects of irradiation is considered.

MEASURES MODIFYING DESTRUCTIVE EFFECTS OF IRRADIATION

OR AFFECTING RECOVERY

Many substances that have been used to treat hematologic disorders in
the human being have also been screened for their possible prophylactic
or therapeutic value. Simmons et at. (1946) and Jacobson, Steamer, and
Simmons (1947) found that, in experimental animals, substances includ-
ing folic acid, liver extract, ascorbic acid, and pentnucleotide were of no
demonstrable value in reducing radiation injury or in hastening recovery
from this injury.

Only those prophylactic measures that have shown conclusive evidence
of effect on the hematologic recovery pattern from radiation injury will be
discussed in the following sections.

PROPHYLACTIC MEASURES

Induced Hyperplasia of Blood-forming Tissue. Jacobson, Marks,
Gaston, Simmons, et al. (1948) reported that the induction of erythro-
blastic hyperplasia by a hemolytic agent (phenylhydrazine hydrochloride)
or by repeated phlebotomy prior to irradiation prevented or reduced the
atrophy of the bone marrow observed in normal adult rabbits after expo-
sure to 800 r of total-body X irradiation. As is shown in Figs. 16-4,
16-16, and 16-17, phenylhydrazine produced an anemia, a reticulocytosis,
and a hyperplastic bone marrow. Although an anemia was produced in
normal rabbits which reached a maximum in 14 days, no further anemia
occurred in animals with a phenylhydrazine-induced anemia at the time
of irradiation, and, as can be noted by examining the reticulocyte values

HEMATOLOGIC EFFECTS OF RADIATION

1065

of histologic preparations, the production of erythrocytes was maintained
within essentially normal limits. The failure of a mid-lethal dose (800 r)
to produce further anemia under these conditions or to interfere materi-
ally with recovery is probably dependent on the multiplication of the
surviving primitive erythroblast precursors or on the functional survival
of even more primitive cells such as reticular cells. That this phenom-
enon was not due to a specific effect of the phenylhydrazine itself on the
cells of the marrow is indicated by the fact that phlebotomy before
irradiation afforded similar protection. Schack and MacDuffee (1949)
have corroborated these findings using the technique of exposing mice to a

10

O

C/l

>
o

o
cc

i
i-
>
cc

10
9
8
7
6
5
4
3
2
l

.-o-^

800 r

J_

J_

J L

J_

_L

I I I I I I L

-8 -6 -4 -2 0

PHENYLHYDRAZINE
ADMINISTRATION

2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34
TIME AFTER X RAY, days

o o PHENYLHYDRAZINE AND 800 r OF TOTAL-BODY X IRRADIATION

o o 800 r OF TOTAL" BODY X IRRADIATION ONLY

• • CONTROL © @ PHENYLHYDRAZINE

Fig. 16-16. Effect of a single dose of 800 r of total-body roentgen irradiation on the
erythrocyte values in the peripheral blood of normal rabbits and rabbits with a
phenylhydrazine-induced anemia. {Originally published in Science, 107: 249, 1948.)

low oxygen tension (simulated high altitude of 15,000 ft) and thus induc-
ing erythroblastic hyperplasia prior to irradiation. Bethard, Skirmont,
and Jacobson (1950) have demonstrated that, during the period of
maximum erythroblastic hyperplasia induced by cobalt, radiophosphorus
produces lesser inhibition of erythropoiesis than in normal rats (Fig.
16-18). More recently, Valentine and Pearce (1952) have compared the
regenerative capacity of erythroid tissue in irradiated and nonirradiated
cats. Immediately prior to irradiation (200 r, whole body) the cats were
bled sufficiently to reduce the peripheral erythrocyte count by 40 per cent.
It was found that recovery from the anemia induced by hemorrhage
immediately prior to irradiation was only slightly less rapid than in the
control nonirradiated cats. Exposure to low oxygen tension after irradia-
tion had no effect on recovery of hematopoiesis after 400 or 500 r according
to Smith, Dooley, and Thompson (1948).

1066

RADIATION BIOLOGY

The mechanism whereby phenylhydrazine-induced anemia or phlebot-
omy reduces the inhibition of erythropoiesis expected after a given dose of
X radiation is probably unrelated to the observation of Dowdy, Bennett,
and Chastain (1950) in which survival of rats exposed to lethal doses of
X radiation was enhanced by reducing oxygen tension available to the
animals to about 6 per cent during irradiation. This latter procedure,

Ht

4B

w

4C

Fig. 16-17. The effect of 800 r of total-body roentgen irradiation on normal and
hyperplastic bone marrow of the rabbit. (IA,B,C) Range of normal control marrow.
(2A,B,C) Bone marrow after phenylhydrazine-induced hyperplasia, 3, 4, and 6 days,
respectively, after phenylhydrazine withdrawal. (3A,J3,C) Bone marrow of phenyl-
hydrazine-induced hyperplasia at 1, 3, and 5 days, respectively, after 800 r and 3, 4,
and 6 days, respectively, after phenylhydrazine withdrawal. (AA,B,C) Bone marrow
of normal rabbits exposed to 800 r at 1, 3, and 5 days after 800 r. Magnification 16 X.
(Originally published in Science, 107: 248, 1948.)

for all practical purposes, stops metabolic activity, whereas phenyl-
hydrazine or phlebotomy actually enhances the metabolic activity of the
cells concerned in erythropoiesis. The mechanism whereby phenyl-
hydrazine- or phlebotomy-induced hyperplasia reduces the expected
radiation effect is not known, but it must be assumed that some metabolic
change within the cells renders them less susceptible to radiation.

Use of Estrogens. Treadwell, Gardner, and Lawrence (1943) demon-
strated that estradiol benzoate, given 10 days prior to X radiation in the

HEMATOLOGIC EFFECTS OF RADIATION

10G7

LD50 range and above, significantly decreases mortality. The mechanism
underlying this fact is not resolved, but studies by Patt, Straube, et at.
(1949) have conclusively shown that the recovery of the hematopoietic
tissue and subsequent return of certain constituents of the peripheral
blood occurred earlier in mice given estrogens before radiation than in
control irradiated nonestrogen-treated mice. The observed hematologic
effect of estrogen pretreatment indicates that a greater effect is exerted
on regeneration of granulocytes and erythrocytes than on lymphocytes.

Other Prophylactic Measures. Sulfhydryl enzyme systems inactivated
by irradiation are reactivated in vitro by the addition of glutathione
and other similar compounds (Barron et at., 1949). Patt, Tyree, et at.

80
70

! P32 INJECTION —

I IHII!

■'j^Zz.

COBALT

CONTROLS

vyfc-P32 PLUS
"'"' COBALT

p32-
COBALT ADMINISTRATION

INITIAL

P32 INJECTION

PLUS P32 PLUS

COBALT

19 22 25
Tl M E, weeks
Fig. 16-18. Effect of repeated injections (1.5 juc/g) of P32 on the hematocrit values of
the peripheral blood of rats with and without cobalt-induced polycythemia.

(1949) and Patt, Smith, and Jackson (1950) demonstrated that the
administration of cysteine just prior to total-body irradiation of rats and
mice increases survival of rats and, to some extent, hastens the recovery
of blood-forming tissue in rats. Glutathione has since been shown by
Cronkite, Brecher, and Chapman (1951) to produce an effect similar to
cysteine on survival of mice and on hematopoietic recovery. Enhance-
ment of hematopoietic recovery has been reported after severe oxygen
deprivation (Bennett, Hanson, and Dowdy, 1951), but no hematologic
data are available on animals subjected to cyanide intoxication during
irradiation (Bacq et al., 1950). The mechanism by which estrogens,
cysteine, or glutathione modify the hematologic picture is not known.
That the mechanism is different from the one observed after phenyl-
hydrazine or phlebotomy is clear since the destruction of hematopoietic

1068

RADIATION BIOLOGY

tissue occurs even though cysteine or estrogens have been given prophy-
lactically. The decreased effect observed in the peripheral blood is due
to a more rapid regeneration of the hematopoietic tissue from the more
radioresistant precursors such as reticular cells, whereas the effect
observed when phenylhydrazine is given prophylactically is that hemato-
poietic tissue, including reticular cells, blast forms, and perhaps even more
mature forms, survives and is immediately capable of accelerated produc-
tion. Regeneration of granulocytic tissue is more rapid than regenera-
tion of lymphatic tissue when cysteine has been given prophylactically if
the peripheral blood can be used as a measure of the production of
granulocytes and lymphocytes. On the other hand, phenylhydrazine,
although producing a panhyperplasia of the bone marrow, produces a
more extreme erythroblastic hyperplasia which persists even after irradia-
tion. Erythropoiesis is the major initial regenerative activity of the
bone marrow under these conditions.

"THERAPEUTIC" MEASURES

Death of animals exposed to single total-body dosages of ionizing
radiations in the lethal range is assumed to be due to the destruction of
certain tissues in the body and failure of functional reconstitution of one

Table 16-5. Survival of Mice Exposed to Various Dosages of X Radiation

WITH AND WITHOUT SHIELDING OF THE SURGICALLY EXTERIORIZED SPLEEN

Lead

shielding
of spleen

Dosage,
r

Number
of mice

Number
of 28-day
survivors

Survival,
per cent

Yes

1025

176

134

76.0

No

1025

431

10

2.3

Yes

1100

82

30

36.5

No

1100

15

0

0.0

Yes

1200

42

23

54.7

No

1200

6

0

0.0

Yes

1300

156

5

3.2

Yes

1300''

48

15

31.2

No

1300

28

0

0.0

Yes

1600

46

0

0.0

No

1600

a Technique differed in that the artery at the distal tip of the spleen was not severed,
and thus no infarct occurred in the spleen.

or more of the tissues (e.g., the hematopoietic system) that individually
or collectively are vital to the survival of the animals. Although many
reasons for the importance of the blood-forming tissue to survival can be
enumerated, it cannot be stated with certainty that death or survival of
the irradiated animal invariably depends on the direct or indirect results

HEMATOLOGIC EFFECTS OF RADIATION

10G9

of hematopoietic dysfunction. In the studies that are discussed in the
following sections, a close parallelism between survival and hemato-
poietic regeneration is obvious, but it must be borne in mind that this cor-
relation may be more apparent than real and that failure of tissues other
than the blood-forming tissue may be critical. More precise methods of
study of other organ systems and of the dependence of organ systems on
one another may be necessary before the factors contributing to death or
survival of irradiated animals can be clearly separated.

Lead Shielding of the Exteriorized Spleen and Other Parts of Body. Lead
shielding of the surgically exteriorized spleen (average weight 0.1 g) of

<0

O

en

Ld

H
>
o
o
a.
i
I-
>-
a.
u

12

I I

10
9
8
7
6
5
4
3

2

I
0

fc^^»-

Jaa-s^>--0-

/

^:d

\

X

\

V

/

J_

_L

J_

-3

23

29

© 0

6 9 12 15 19

TIME AFTER X RAY, days
CONTROLS O O OPERATED CONTROLS

1300 r WITH SPLEEN SHIELDING
AND DISTAL VESSEL NOT CUT
600 r WITH SPLEEN EXTERIORIZED
AND DISTAL VESSEL CUT

Fig. 16-19. Comparative effect of 600 r of total-body roentgen irradiation (spleen
exteriorized and distal vessel cut) and of 1300 r with lead shielding of the exteriorized
spleen (distal vessel not cut) on the erythrocyte values of CFi female mice.

adult mice during exposure to 1025 r of total-body X irradiation markedly
enhances survival (Table 16-5). After exposures up to 1300 r. no anemia
and only a transient leukopenia and thrombocytopenia appear in spleen-
shielded mice, whereas a severe pancytopenia follows exposure to 600 r
without spleen shielding (Figs. 16-19, 16-20). Recovery of hemato-
poietic tissue in mice exposed to 1025 r with spleen shielding occurs by 8
days, but no hematopoietic recovery is noted during this interval in
unshielded mice, Fig. 16-21 (Jacobson, Simmons, Marks, et at., 1950).
Recovery of the lymphatic tissue in the wall of the gastrointestinal tract
in spleen-shielded mice parallels recovery of the hematopoietic tissue
elsewhere. These observations led to the theory that the mechanism of
recovery from radiation injury under these conditions was on a humoral
basis and that the factor (or factors) responsible was produced by the

1070 RADIATION BIOLOGY

cells of the shielded tissue (Jacobson, Simmons, Bethard, et at., 1950;
Jacobson, Simmons, Marks, et at., 1950).

The survival of mice exposed to 1025 r of total-body X irradiation is
approximately 30 per cent if part of the exteriorized liver (0.8 g), the
exteriorized intestine (2.5 g), the entire head (3.0 g), or one entire hind
leg up to the thigh (1.5 g) is lead shielded during exposure. Without
shielding, only 0.8 per cent survive this dose; with spleen shielding at

I-

o

o

20

10
8
6

4
3

I
0.8

0.6

0.4
0.3

0.2

-3 0 3 6 9 12 15

TIME AFTER X RAY, days

19

-•CONTROLS 0---0 OPERATED CONTROLS

e ®600r WITH SPLEEN EXTERIORIZED

AND DISTAL VESSEL CUT

cr-— A 1300 r WITH SPLEEN SHIELDING
AND DISTAL VESSEL NOT CUT

Fig. 16-20. Comparative effect of 600 r of total-body roentgen irradiation (spleen
exteriorized and distal vessel cut) and of 1300 r with lead shielding of the exteriorized
spleen (distal vessel not cut) on the leukocyte values of CF! female mice.

least 76 per cent survive. Shielding one exteriorized kidney (average
weight 0. 19 g) does not enhance survival. Recovery of the hematopoietic
tissue, as judged by histopathologic and hematologic examination, is
under way by 8 days in liver- or intestine-shielded animals, whereas after
lead shielding of the head, recovery of these tissues is delayed even longer,
and recovery of hematopoietic tissue is nil with kidney shielding (Jacob-
son, Simmons, Marks, and Eldredge, 1951; Jacobson, Simmons, Marks,
Gaston, et al., 1951).

The effect of spleen or appendix shielding on the survival of irradiated
rabbits has not been carefully studied, but it is clear that no enhancement
of survival occurs such as is observed in spleen-shielded mice or rats.

HEMATOLOGIC EFFECTS OF RADIATION

1071

^* «WT\ '

Fig. 16-21. Femoral marrow of mice 8 days after exposure to 1025 r of total-body X
irradiation. (.4) Control untreated. (B) Control operated only. (C) Total-body
X irradiation without spleen protection. (D) Total-body X irradiation with spleen
lead-shielded during irradiation.

Spleen or appendix shielding in the rabbit during exposure to 800 or
1000 r appears not to affect survival appreciably even though regenera-
tion of blood-forming tissue precedes the recovery of this tissue in the
animals without spleen or appendix shielding (Jacobson, Simmons,
Marks, Gaston, et al., 1951).

1072 RADIATION BIOLOGY

Allen et al. (1948) have reported that 450 r of total-body X irradiation
is invariably lethal to dogs. With head shielding (Allen, 1951), however,
mortality following this dose is reduced to 75 per cent, and other aspects
of the usual postirradiation syndrome, such as hemorrhage and evidences
of infection, are greatly reduced or absent. Further work on the dog
comparing the relative effectiveness of shielding parts such as the head,
spleen, intestine, limbs, and liver after exposure to various dosages of
total-body X irradiation will be of interest if for no other reason than to
obtain baselines on the potential effectiveness of these tissues for com-
parison with mice, rats, and rabbits and to accumulate some facts on the
potential production of the factor on a tissue-weight basis, type of tissue
shielded, etc. The species differences that are apparent cannot be ade-
quately evaluated since, for example, considerable differences may exist
between rabbit and mouse spleen in terms of the potential production of
the factor (or factors) involved in survival or early regeneration of
hematopoietic tissue. In this connection the work of Bond et al. (1949),
in which the abdomens of rats were shielded with lead during total-body
X irradiation, is of interest. The LD50 for these abdominal-shielded rats
was 1950 r compared with ca. 700 r for nonshielded control rats. For
very practical reasons it would be of interest to determine more precisely
the relative importance of the various abdominal tissues in enhancing
survival on a weight basis and to assess more adequately the volume-dose
factor. This has been done to some extent with rats by Gershon-Cohen,
Hermel, and Griffith (1951), Jacobson et al. (1949), Jacobson, Simmons,
Bethard, et al. (1950), Jacobson, Simmons, Marks, et al. (1950), and
Jacobson, Simmons, Marks, Gaston, et al. (1951) with mice, but further
data for all species are needed. As was pointed out by Bond et al. (1949),
the radiosensitivity of the part of the body irradiated may be more impor-
tant than the gram-roentgens sustained by the balance of the body. To
this must be added the fact that the actual or potential production of the
factor (under consideration in this section) by the shielded or nonirradi-
ated tissue may be more important in determining survival of the animal
than the radiosensitivity of the tissue in the radiation field and, within
certain limits, more important than the gram-roentgens sustained by the
balance of the body.

The amount of tissue shielded in the intestine- and liver-shielding
experiments in mice was greater by factors of 25 and 8, respectively, than
that shielded in the mouse spleen-shielding experiments, yet survival was
considerably less. These findings suggest that the potential production
of a factor (or factors) by the intestine and the liver is not as great as that
by splenic tissue but yet is sufficient to institute recovery early enough in
a sufficient number of cells of the body to have a definite effect on survival.

It is conceivable that practically all tissues of the body are capable of
producing the factor (or factors) concerned in recovery from radiation

HEMATOLOGIC EFFECTS OF RADIATION 1073

injury (Jacobson, Simmons, Marks, and Eldredge, 1951; Jacobson, Sim-
mons, Marks, Gaston, et al., 1951), but certain tissues and, in particular,
the hematopoietic system have a greater potential production per unit
volume.

There are several differences between head or limb shielding and spleen
implantation (to be described later) and spleen shielding. According to
generally accepted concepts, the reduction in volume dose when the head,
limb, or intestine is shielded must be considered as playing a role in the
reduction of mortality since these structures represent a fairly large pro-
portion of the body weight (15, 7.5, and 12.5 per cent, respectively). On
the other hand, the shielded spleen weighs only 0.1 g (0.005 per cent of the
body weight) and spleen implants weigh 0.010 g (0.0005 per cent of the
body weight) which eliminates from consideration the volume-dose factor
of shielded spleen or implanted spleen. The fact that the head, hind
limbs, and intestine contain reticuloendothelial tissue and other tissue of
mesenchymal origin is probably more important than the volume-dose
consideration.

Postirradiation Spleen Trans-plantation. Transplantation of spleens
(total weight, 10 100 mg) from baby or adult mice into the peritoneal
cavity of mice within 2 hours after exposure of the recipient adult mice to
1025 r of total-body X irradiation significantly increases the survival
(ca. 50 per cent) of the irradiated mice and hastens regeneration of
hematopoietic tissue (Jacobson, Simmons, Marks, and Eldredge, 1951;
Jacobson, Simmons, Marks, Gaston, et at., 1951). Transplantation of
spleens into the peritoneal cavity of mice 1 or 2 days after exposure to
1025 r of total-body X irradiation likewise enhances survival (ca. 25 per
cent) but not as effectively as earlier transplantation. Implantation of
muscle into the peritoneal cavity after exposure of mice to 1025 r of total-
body X irradiation has no beneficial effect on survival.

If splenectomy is performed in mice prior to irradiation, followed by
implantation of fresh spleens into the peritoneal cavity after irradiation,
survival is enhanced, indicating that an intact spleen is not required to
make the transplant effective. Surgical removal of the transplanted
spleens from the peritoneal cavity of mice 1 and 2 days after the irradia-
tion-transplant procedure has invariably been followed by death of the
animals. It is assumed that the ineffectiveness of this procedure is due
to the fact that the implanted spleens are not vascularized prior to
removal and are therefore incapable of elaboration and distribution of
the factor. Gross and microscopic observations on mice surviving the
irradiation-transplant procedure reveal that the implanted spleen or
spleens have vascularized and eventually appear as normal splenic tissue.
Revascularization and reconstitution of the implanted spleen are usually
well under way by the sixth day after implantation. Transplantation of
splenic tissue 2 days after irradiation is admittedly less effective in

1074 RADIATION BIOLOGY

enhancing survival of mice exposed to 1025 r than earlier transplantation,
but it has not been determined when a state of irreversibility has been
reached. Actually, if establishment of a vascular supply to the trans-
plant is essential to the manufacture and transport of the factor (or
factors) in question, a conservative guess would be that supplying an
optimum amount of the factor to mice as late as 6 days after exposure to
1025 r would still significantly increase the survival.

Administration of Suspension of Mashed Embryos. Intraperitoneal
administration of a suspension of 12-day-old mouse embryos is effective
in enhancing survival of mice exposed to 1025 r of total-body X irradia-
tion (Jacobson, 1952; Jacobson, Marks, and Gaston, 1951). This sus-
pension, prepared in the cold with or without the addition of normal
physiological saline or buffered saline, when given intraperitoneally in a
dosage of from 0.5-1.0 ml, 2-6 hours after irradiation of the recipient,
results in 30 per cent survival. Suspensions of baby or adult spleens
prepared and administered in a similar manner have thus far been ineffec-
tive in enhancing survival of mice exposed to 1025 r but have been effec-
tive in enhancing the survival of mice exposed to 800 r (Jacobson, 1952;
Jacobson, Marks, and Gaston, 1951). Chick embryo suspensions (age
of embryos, 11-14 days) prepared and administered in a similar manner
to the mouse embryo suspensions have been reported by Marks (unpub-
lished data, 1952), Stroud and Brues (unpublished data, 1952), and
Jacobson (1952) to be ineffective in enhancing survival of mice exposed to
X radiation in the LD5o range or above.

The factor (or factors) in the embryo or spleen suspensions responsible
for recovery from radiation is probably the same as that responsible for
the effectiveness of the spleen shielding and spleen implants. Two
possible explanations for the effectiveness of cell suspensions are obvious:
(1) that the cells in the suspension quickly implant and begin elaboration
of the factor or factors effective in initiating tissue regeneration through-
out the body or (2) that the peritoneal cavity serves as an incubator that
allows the cell suspension to remain alive and to elaborate the factor
responsible for increased survival and tissue regeneration in irradiated
mice.

Homologous Bone-marrow Injection. Lorenz et al. (1951) have shown
that, although 900 r is the LD99 for genetically homogeneous hybrid
LAFi mice, approximately 75 per cent survive this dose if bone marrow
aspirated from the long bones of normal nonirradiated mice of the same
strain is injected intravenously within an hour after the exposure. If the
bone marrow is administered intraperitoneally, survival is slightly less
(ca. 50 per cent). The author estimates that the total weight of the
injected marrow is approximately 1.5 mg. The recovery of the hemato-
poietic tissue of the bone-marrow-treated mice, as in the spleen-shielded,
spleen-implanted, or embryo-suspension-injected mice, is hastened.

HEMATOLOGIC EFFECTS OF RADIATION 1075

Jacobson (1952) obtained similar results in CFi female mice with intra-
venous injections of marrow from normal mice. Rekers (1948, 1950),
Talbot and Pinson (1951), and Talbot and Gertsner (1951) have reported
negative and equivocal beneficial effects on the radiation syndrome in
dogs and rats, respectively, after bone-marrow administration.

The suspension of bone marrow that Lorenz and Jacobson injected
contained mature and immature free cells such as granulocytes and mega-
karyocytes as well as free and fixed macrophages, reticular cells, and
endothelial tissue. It seems likely that the cells injected establish as
scattered foci of hematopoietic tissue and produce a factor (or factors)
responsible for survival of the animal and that this factor is identical
with that postulated in the spleen-shielding, spleen-implantation, and
embryo-suspension experiments. Bone-marrow injection is not as effec-
tive as spleen shielding or spleen transplants in enhancing survival or
hastening recovery of hematopoietic tissue in mice exposed to 1000 r or
more. Since it is not known which cells in the shielded tissue or trans-
planted tissue (including bone marrow) are the most important in bring-
ing about the effect, no adequate data are available to compare adequately
the relative effectiveness of splenic tissue and bone marrow in enhancing
survival from radiation injury.

Heterologous Transplants and Cell Suspensions. Lorenz, Congdon, and
Uphoff (1952) have reported evidence of the effectiveness of heterologous
tissue transplants on recovery from radiation injury. Within an hour
after exposure of mice to 900 r (LD99) of total-body X irradiation, approxi-
mately 25 mg of freshly aspirated guinea-pig bone marrow (in buffered
saline) was injected intravenously into the irradiated mice. None of the
control irradiated mice survived, but 40 per cent of the irradiated mice
which received intravenous guinea-pig bone-marrow suspension survived
the 28-day period of observation. As mentioned previously the survival
of this strain of mice injected with homologous bone marrow (1.5 mg)
after exposure to 900 r is ca. 75 per cent. The number of animals used
by Lorenz, Congdon, and Uphoff (1952) in the heterologous transplant
experiment is too small to warrant a positive conclusion. If further
data corroborate these findings, proof that the factor (or factors) is
humoral would be available.

It would not seem likely that heterologous bone-marrow injection could
seed the hematopoietic tissue with cells which, by multiplication, repopu-
late the tissue. If heterologous tissue is effective as suggested by Lorenz,
it seems more likely that this heterologous tissue lives, at least tempo-
rarily, in its new environment and produces a noncellular substance (or
substances) that aids in recovery from the radiation injury.

Po stir radiation Parabiosis. Barnes and Furth (1943) first demon-
strated that parabiosis diminished the deleterious effects of irradiation.
When one member of a parabiotic pair was irradiated, the pathological

1076 RADIATION BIOLOGY

changes in the blood and blood-forming tissues of the irradiated mouse
were less conspicuous than those in a single mouse similarly irradiated,
while in the nonirradiated parabiont, the changes were only slight. More
recently Brecher and Cronkite (1951) reported that approximately
50 per cent of rats exposed to 700 r of total-body X irradiation survive
if they are joined to normal nonirradiated litter mates within a few hours
after irradiation. None of the controls given the same does of irradiation
survived the 28-day period of observation. Hematopoietic regeneration
was more rapid in the irradiated rats with a parabion than in the irra-
diated controls. These findings, like those of Lorenz et al. (1951),
corroborate Jacobson's previously reported observation that effective
postirradiation "therapy" is a reality. Like the embryo-suspension
experiments described, however, the experiments of Brecher do not point
out the cellular source nor the identity of the effective factor (or factors)
concerned.

Relation between Quantity of Shielded or Implanted Splenic Tissue and
Effects on Survival. Two separate observations indicate that a definite
relation exists between the quantity of implanted or shielded tissue and
the effect as measured by hematopoietic regeneration or survival from
irradiation (Jacobson, Simmons, Marks, Gaston, et al., 1951).

1. The transplantation of two spleens (weight, ca. 5 mg) from 7- to
12-day-old baby mice into the peritoneal cavity of mice immediately
after irradiation (1025 r) does not enhance survival, although the trans-
plantation of four spleens (weight, ca. 10 mg) is effective in reversing the
process in time to allow recovery of the animal (Jacobson, Simmons,
Marks, Gaston, et al., 1951).

2. In the regular spleen-shielding technique the main splenic pedicle is
left intact, but a small blood vessel at the distal tip of the spleen is
severed to facilitate exteriorization and lead shielding of the spleen.
Invariably from one-fourth to one-half the spleen proximal to the severed
vessel becomes infarcted and undergoes liquefaction necrosis. If this
vessel is not cut and the whole spleen is shielded and remains intact, 100
per cent of the animals survive exposure to 1025 r rather than the expected
77 per cent. In fact, with a total-body exposure to 1300 r, only 3.4 per
cent of the animals survive if the distal vessel of the spleen is cut during
the shielding procedure, whereas 26.9 per cent survive this exposure with
spleen shielding if the distal vessel is not cut (Jacobson, Simmons, Marks,
Gaston, et al., 1951). Hematopoietic regeneration is complete by 8 days
in the animals exposed to 1300 r with spleen shielding and in which the
distal vessel to the spleen is not cut. In the animals, exposed to this
dose with spleen shielding but with the distal vessel cut, regeneration is
not as rapid (Figs. 16-19, 16-20). This observation like observation 1
indicates that the quantity of the factor being produced is directly related

HEMATOLOGIC EFFECTS OF RADIATION 1077

to the number of cells available in the shielded or implanted tissue and
the amount of the factor available determines the survival of the animal.
It would appear as though the repair process must be initiated in a mini-
mum number of cells in the body of the irradiated animal to ensure sur-
vival of the animal exposed to dosages of 1000 r or more.

The Humoral Theory of Cell Regeneration. In mice and rabbits which
had spleen shielding or spleen transplants, regeneration may occur from
the scattered "free cells" in the lymphatic tissues and bone marrow that
survive the radiation, but heteroplastic regeneration from reticular cells
is prominent. Thus colonization from the shielded tissue and repopula-
tion by multiplication of these colonized cells, if a factor at all, is only one
aspect of the recovery process. The shielded tissue in some way restores
the functional capacity of the reticular cells to repopulate the hemato-
poietic tissues. The shielded tissue may likewise restore the functional
capacity of the residual free cells, which are not destroyed by irradiation,
to multiply and thus repopulate the hematopoietic tissues. Cells coming
from the shielded or implanted tissue cannot at the moment be dis-
tinguished from the residual free cells. If the cells, which do migrate out
from the shielded tissue, do "lodge" in hematopoietic tissue, then it is
also possible that they too contribute by division and multiplication and
also by elaboration of the factor (or factors) under discussion.

Clamping Off Splenic Circulation during Irradiation-shielding Procedure.
The survival of mice in which the circulation to the shielded spleen is
clamped off during exposure of the animal to 1025 r and in which the
clamp is released immediately after irradiation is approximately the
same as survival in animals with spleen shielding but without clamping
(Jacobson, Simmons, Marks, and Eldredge, 1951; Jacobson, Simmons,
Marks, Gaston, et al., 1951). Histologic recovery of the hematopoietic
system is the same as in the spleen-shielded animals without clamping of
the splenic pedicle. This observation was convincing evidence that the
presence of shielded tissue in the circulation was not required during the
period of irradiation in order for survival to be enhanced and hemato-
poietic regeneration to proceed. Reintroduction of the spleen into the
circulation after irradiation could thus be considered an effective post-
irradiation "therapeutic" approach to the problem.

Splenectomy after Spleen Shielding. Surgical extirpation of the ini-
tially shielded spleen at intervals after exposure of mice to 1025 r of total-
body X irradiation shows that a beneficial effect (survival > 70 per cent
and early regeneration of hematopoietic tissue) has already been exerted
if the shielded spleen is left intact in the circulation for 1 hour (Jacobson,
Simmons, Marks, Gaston, et al., 1951). Leaving the spleen in the circula-
tion for longer periods such as 6 or 24 hours or 2 or more days does not
increase the percentage of animals surviving. In a previous communica-

1078

RADIATION BIOLOGY

tion (Jacobson, Simmons, Marks, and Eldredge, 1951) it was reported
that, if splenectomy was performed within 10 minutes after the irradia-
tion-spleen-shielding procedure, none survived. Further work, how-
ever, has shown that leaving the spleen in the circulation for as little as 5
minutes is sufficient to significantly increase the survival of mice exposed
to 1025 r (Jacobson, Simmons, Marks, Gaston, et al., 1951). Full
recovery of the blood-forming tissues is delayed longer in mice splenecto-
mized 5 minutes after irradiation than in mice splenectomized 24 hours

6 9 12 15 19 22

TIME AFTER X RAY, days

29

• • CONTROLS

o -o SPLENECTOMY CONTROLS

o » 1025 r WITH LEAD SHIELDING

o o 1025 r WITH LEAD SHIELDING; SPLENECTOMY

WITHIN 5 MINUTES AFTER X IRRADIATION
s s 1025 r WITH LEAD SHIELDING; SPLENECTOMY

24 HOURS AFTER X IRRADIATION

Fig. 16-22. Effect of splenectomy on the reticulocyte values of mice exposed to 1025 r
of total-body roentgen irradiation with lead shielding of the exteriorized spleen.

after irradiation (Fig. 16-22). If the originally shielded spleen (whether
or not the pedicle is clamped during the irradiation) is not removed, com-
plete regeneration of hematopoietic tissue occurs earlier than in mice with
splenectomy 24 hours after the shielding procedure (Jacobson, Marks,
et al., 1951). These facts indicate that the intact spleen may release
enough of the factor in a few moments to enhance significantly the sur-
vival but that, if left in the circulation longer, an earlier and more com-
plete regeneration of hematopoietic tissue occurs.

Total-body Exposure to 1025 r plus 200-r Increments to Spleen. Mice
have been exposed to 1025 r of total-body X irradiation, and the spleen
has been given various increments of the total-body dose. Doses up to
and including 200 r may be given to the spleen at the same time as 1025 r

HEMATOLOGIC EFFECTS OF RADIATION 1079

of total-body exposure without reducing survival below the 75 per cent
that is expected from the earlier spleen-shielding studies (Jacobson,
Simmons, Marks, Gaston, et al., 1951). In contrast to animals with
spleen shielding and thus no irradiation of the spleen, these animals
become moderately anemic and develop a severe leukopenia that persists
beyond the twelfth day. Recovery of erythropoiesis as judged by the
circulating reticulocytes begins by the sixth day, Fig. 16-22 (Jacobson,
Marks, et al., 1951). Histologic studies show that recovery of the
blood-forming tissue is qualitatively complete by 10-12 days (Jacobson,
Simmons, Marks, Gaston, et al., 1951). Even with dosages of 400, 500,
or 600 r to the spleen and 1025 r to the body, survival is significantly
higher (59, 50, and 34 per cent, respectively) than in mice exposed to
1025 r without spleen shielding (1.1 per cent). Recovery of hemato-
poietic tissue is delayed progressively longer with increasing increments of
the total-body dose delivered to the spleen. These data indicate that the
capacity of the splenic tissue to elaborate the factor is still partially
retained or recovery of the tissue in the spleen that produces the factor
occurs early enough to enhance survival even with doses as high as
600 r to the spleen and 1025 r to the body. These observations tend to
add support to the hypothesis that the factor (or factors) responsible for
recovery from radiation injury under these circumstances is derived from
more primitive but more radioresistant cells, such as reticular cells,
rather than from the free cells of the hematopoietic system, such as
lymphocytes and granulocytes. The spleen is, for all practical purposes,
devoid of these free cells after a dose of 500 r, whereas the basic reticular
network remains "histologically " intact. It is interesting that, following
the delivery of a lethal dose to the body (1025 r) and an LD50 (500-600 r)
to the spleen, ca. 50 per cent of the animals survive.

EFFECT OF COMBINED PROPHYLACTIC
AND THERAPEUTIC MEASURES

The fact that a reduction in the mortality of animals exposed to lethal
dosages of total-body X irradiation could be effected by pretreatment
with estrogens (Treadwell, Gardner, and Lawrence, 1943) as well as by
spleen shielding (Jacobson et al., 1949) suggested to Simmons, Jacobson,
and Marks (1950-1951) that these two measures might produce an addi-
tive effect on survival. Accordingly, Simmons tested this hypothesis and
found that (1) mortality of mice exposed to 1025 r of total-body X irradia-
tion was 100 per cent, (2) 61.5 per cent of the mice survived this dose if
estrogens were given prior to irradiation, (3) 82.3 per cent survived if the
spleen was shielded during exposure to 1025 r, and (4) 100 per cent
survived 1025 r if the techniques of pretreatment with estrogens as well
as spleen shielding were employed. Bethard, Skirmont, and Jacobson
(1950) and Bethard and Jacobson (1951), employing the same general

1080 RADIATION BIOLOGY

approach, found that cysteine and spleen shielding similarly had an
additive effect on survival of mice exposed to X radiation. Furthermore,
it was found that, when the techniques of pretreatment with estrogens
and cysteine and spleen shielding during irradiation were all combined,
an additive effect on survival was observed (Bethard, Simmons, and
Jacobson, 1951). Jacobson (1952) found that pretreatment with
cysteine followed by 1025 r of total-body X irradiation and postirradia-
tion intraperitoneal transplantation of normal spleen were also additive
in enhancing survival. Cysteine fails to produce any enhancing effect on
survival of irradiated mice when given after 1100 r of total-body X
irradiation whether or not the mice had spleen shielding during irradiation
(Jacobson, 1952). These studies, although interesting, shed no light on
the obvious question of whether or not the pretreatment or prophylactic
techniques are related to the therapeutic techniques from the standpoint
of mechanism.

ANTIBODY-FORMATION STUDIES

The fact that a single total-body exposure to X rays inhibits antibody
formation is well documented (Benjamin and Sluka, 1908; Hektoen,
1918). The time of antigen injection relative to irradiation determines
to a large measure the degree of inhibition (Benjamin and Sluka, 1908;
Hektoen, 1918). Exposure of rabbits to dosages of total-body X irradia-
tion ranging from 250-800 r effectively suppresses the development of
appreciable antibody titer of particulate antigens administered in the
24 hours before or the 48 hours after irradiation (Benjamin and Sluka,
1908; Hektoen, 1918). It would appear from these observations that
irradiation either destroys the cells concerned with antibody formation or
reduces their functional capacity to react in the normal way to the
injected antigen. It is not known with certainty in which cells of the
body this function normally resides, but it has been postulated in a
broad sense that the cells of the reticuloendothelial system are responsible.
As suggested by Hektoen (1915) the degree of inhibition of antibody
formation correlates well with the extent of damage to the blood-forming
tissue induced by irradiation.

It has been demonstrated by Jacobson, Robson, and Marks (1950) that,
if the spleen or the appendix of the rabbit is surgically exteriorized and
shielded with lead during total-body exposure to 800 r, the capacity to
form antibodies to a particulate antigen injected 24 hours after irradiation
is retained. In another series of experiments this observation was carried
a step further (Jacobson and Robson, 1952). Spleen-shielded rabbits
were exposed to dosages of 800 or 500 r of total-body X irradiation.
Twenty-four hours later the spleen was removed surgically. After
another 24-hour period (48 hours after irradiation) a particulate antigen
(sheep red cells) was given intravenously. The capacity of these animals

HEMATOLOGIC EFFECTS OF RADIATION 1081

to form antibodies (anti-sheep-cell hemolysin) was compared with various
control groups given the same antigen at the same time relative to the
irradiation as the experimental animals. The capacity to form anti-
bodies to the injected antigen was retained in the rabbits, given 800 or
500 r of total-body X irradiation, which had spleen shielding during
irradiation and the spleen left intact in the circulation for 24 hours and
then removed surgically even though the antigen was given 24 hours
after splenectomy and 48 hours after irradiation. The fact that these
rabbits retained the capacity to form antibodies even though hemato-
poietic tissues in the body were as yet atrophic and that control rabbits
exposed to the same dose did not retain this capacity is considered to be a
result of the functional restoration of cells in the body (such as free and
fixed macrophages and reticular cells) by a humoral (noncellular) sub-
stance entering the general circulation from the originally shielded spleen
during the 24 hours prior to splenectomy.

COMMENT

It seems extremely unlikely that cell migration from the shielded or
transplanted tissue and subsequent proliferation of these cells account for
the reconstitution of hematopoietic tissues and increased survival of
irradiated animals or that neutralization of some toxin produced by
irradiation can account for these findings. Perhaps the latter possibility
cannot be positively excluded on the basis of the available data. The
evidence accumulated strongly suggests that the factor (or factors)
responsible for recovery from radiation under these circumstances is non-
cellular and may be required only for the initiation of the repair process.
The factor (or factors) may be quite labile or, as is more likely, may be
produced in an effective quantity only by living cells. These cells may be
present in shielded or implanted tissue or may migrate out and produce
the factor under discussion. The factor may be a single substance such
as an enzyme or coenzyme necessary for the functional reconstitution of
many different cell types in the several organ systems, or several different
factors may be concerned. Salisbury et al. (1951) found in a small
number of dogs that early direct cross transfusion between irradiated
(LD90, X ray) and nonirradiated dogs significantly reduced mortality,
reduced the severity of the expected hematopoietic depression, and
reduced the severity of the usual clinical signs of irradiation sickness.
Salisbury's experiments (1951) should be expanded in numbers of animals
and should be more adequately controlled to make the significance of the
experiment more clear cut.

The fact that 75 per cent of mice that have lead shielding of the spleen
during exposure to 1025 r and then splenectomy 1 hour after the irradia-
tion-spleen-shielding procedure survive would lead to the expectation
that early administration of whole blood is effective. This seems logical

1082 RADIATION BIOLOGY

since whatever the spleen accomplishes under these circumstances must
be through the blood stream. That the factor (or factors) must be
supplied early is also strongly supported by the fact that, in mice given a
lethal dose of radiation, spleen transplants are more effective on the day
of irradiation than on the second day after irradiation. In other words,
supplying the factor responsible for initiating the functional repair in
some unknown minimum number of cells throughout the body must be
accomplished early enough to reverse the processes that ordinarily end in
death of the animal. Once the factor is adequately supplied, as is
clearly demonstrated by the splenectomy experiments, the process of
repair is initiated ; yet histologic evidence of repair or regeneration is not
apparent for 4 or more days.

Indirect transfusion initiated on the fourth postirradiation day has
been reported to be unsuccessful in significantly increasing survival of
X-irradiated dogs (Allen et at., 1952). This apparent failure of whole-
blood transfusion to enhance survival and reduce morbidity effectively is
understandable if it is assumed that (1) the amount of the factor (or
factors) present in the blood per unit volume is small and therefore rela-
tively large amounts of blood would be necessary to initiate effectively
the recovery process or (2) the factor (or factors) was administered too
late and in too small a quantity after exposure of the recipient to initiate
the repair process in time and widely enough in cells of the body to have
a critical effect on morbidity and survival. The preliminary experiments
of Swisher and Furth (1951) indicate that the morbidity of dogs exposed
to dosages of X irradiation in the mid-lethal range is reduced if as little
as 250 cc of whole blood, collected in ACD solution from a compatible
donor, is administered to irradiated dogs shortly after exposure. The
ineffectiveness of cell-free extracts obtained from extirpated splenic tissue
or embryos may only indicate that too small an amount of the factor is
present or too small an amount of the factor is obtained in the extracts
from these tissues by present methods. A method of preservation or con-
centration of the factor (or factors) or a more sensitive method of assay may
be necessary before a positive result can be obtained by cell-free extracts.
If the factor is present in whole blood in a concentration sufficient to
alter the radiation syndrome even when given in relatively small amounts
soon after irradiation of the recipient, then varying the conditions and
methods of the administration of whole blood may supply important
clues to the identification of the factor (or factors).

HEMATOLOGIC EFFECTS OF RADIATION 1083

REFERENCES

(Information regarding the availability of government reports indicated by an asterisk
may be obtained from the Office of Technical Services, Department of Commerce,

Washington, D.C.)

Adams, W. S., and J. S. Lawrence (1947) The negative effect of folic acid on irradia-
tion leukopenia in the cat. USAEC Report MDDC-1538.*

Allen, J. G. (1947-1948) A method for the in vitro titration of heparin. USAEC
Report ANL-4147,* pp. 89-95.

- (1951) The beneficial effects of head shielding in 20 dogs exposed to 450 r
total-body X radiation. USAEC Report ANL-4625,* pp. 60-61.

(1952) Pathogenesis of irradiation hemorrhage. Conference on blood clot-
ting and allied problems. Trans. 5th Josiah Macy Jr. Foundation, pp. 213-246.

, C. E. Basinger, J. J. Landy, M. H. Sanderson, and D. M. Enerson (1952)

Blood transfusion in irradiation hemorrhage. Science, 115: 523-526.

- and L. O. Jacobson (1947a) Hyperheparinemia: cause of the hemorrhagic
syndrome associated with total body exposure to ionizing radiation. Science,
105: 388-389.

- and (1947b) Irradiation hemorrhage. Proc. Inst. Chicago, 16:

376-377.

, P. V. Moulder, and D. M. Enerson (1951) Pathogenesis and treatment of

the postirradiation syndrome. J. Am. Med. Assoc, 145: 704-711.

M. Sanderson, M. Milham, A. Kirschon, and L. O. Jacobson (1948) Hepa-

rinemia (?), an anticoagulant in the blood of dogs with hemorrhagic syndrome

associated with total body exposure to roentgen rays. J. Exptl. Med., 87: 71-86.
Aubertin, C, and E. Beaujard (1905) Action comparee des rayons X sur le sang

dans les leucemias myelogene et lymphatique. Compt. rend. soc. biol., 58: 177.
and - - (1908) Action des rayons X sur le sang et la moelle osseuse.

Arch. med. exptl. anat. path., 20: 273-288.
Bacq, Z. M., A. Herve, J. Lecomte, and P. Fischer (1950) Cyanide protection

against X-irradiation. Science, 111: 356-357.
Barnes, W. A., and O. B. Furth (1943) Studies on the indirect effect of roentgen rays

in single and parabiotic mice. Am. J. Roentgenol. Radium Therapy, 49: 662-681.
Barron, E. S. G., S. Dickman, J. Muntz, and T. P. Singer (1949) Studies on the

mechanism of action of ionizing radiations. I. Inhibition of enzymes by X-rays.

J. Gen. Physiol., 32: 537-551.
Benjamin, E., and E. Sluka (1908) Antikoerperbildung nach experimenteller Schae-

digung des haematopoetischen Systems durch Roentgenstrahlen. Wien. klin.

Wochnschr., 21: 311-313.
Bennett, L. R., R. A. Hansen, and A. H. Dowdy (1951) The effect of anoxia on the

hematological injury in rats caused by roentgen irradiation. USAEC Report

UCLA- 156.*
Bethard, W. F., and L. O. Jacobson (1951) Preliminary observations on the combin-
ing effect of cysteine and spleen protection on mortality of mice after exposure to

X radiation. USAEC Report ANL-4571,* pp. 30-31.

— , E. L. Simmons, and L. O. Jacobson (1951) Preliminary observations on

enhanced survival in X-irradiated mice following combined treatment with

cysteine, estrogen and spleen protection. USAEC Report ANL-4625,* pp.

48-49.
, E. Skirmont, and L. O. Jacobson (1950) Effect of radiophosphorus on rats

with cobalt-induced polycythemia. Proc. Central Soc. Clin. Research, 23: 12.

1084 RADIATION BIOLOGY

Bloom, M. A. (1948) Bone marrow, in Histopathology of irradiation from external
and internal sources, W. Bloom, ed. McGraw-Hill Book Company, Inc., New
York, National Nuclear Energy Series, Div. IV, Vol. 221, Chap. 6.

- and W. Bloom (1947) The radiosensitivity of erythroblasts. J. Lab. Clin.
Med., 32: 654-569.

Bloom, W. (1947) Histological changes following radiation exposure. Radiology,
49: 344-347.

(ed.) (1948) Histopathology of irradiation from external and internal

sources. McGraw-Hill Book Company, Inc., New York, National Nuclear
Energy Series, Div. IV, Vol. 221.

and L. O. Jacobson (1948) Some hematological effects of irradiation. Blood,

3: 586-592.

Bond, V. P., M. N. Swift, A. C. Allen, and M. C. Fishier (1949) Sensitivity of the
abdomen of the rat to X-irradiation. Nav. Radiol. Def. Lab. Report ADB-92.*

Brecher, G., and E. P. Cronkite (1951) Post-irradiation parabiosis and survival in
rats. Proc. Soc. Exptl. Biol. Med., 77: 292-294.

Brues, A. M., and L. Rietz (1948) Acute hematologic radiation response of the
guinea pig as a function of lethality. USAEC Report ANL-4227,* pp. 183-187.

Clarkson, J. R., W. V. Mayneord, and L. D. Parson (1938) The effect of X-irradia-
tion on the blood and lymphoid tissue of tumor-bearing animals. J. Path. Bact.,
46: 221-235.

Cohn, S. H. (1952) Effects of total body X-irradiation on blood coagulation in the
rat. Blood, 7: 225-234.

Cronkite, E. P. (1950) The hemorrhagic syndrome of acute ionizing radiation illness
produced in goats and swine by exposure to the atomic bomb at Bikini, 1946.
Blood, 5: 32-45.

— , G. Brecher, and W. H. Chapman (1951) Mechanism of protective action
of glutathione against whole body irradiation. Proc. Soc. Exptl. Biol. Med., 76:
396-398.

— , B. Halpern, D. P. Jackson, and G. V. LeRoy (1950) A study of the hemor-
rhagic state in dogs after a lethal dose of two million volt X-rays. J. Lab. Clin.
Med., 36: 814.

-, F. W. Ullrich, D. C. Eltzholtz, C. R. Sipe, and P. K. Schork (1949) The
response of the peripheral blood of swine to whole body X ray radiation in the
lethal range. Nav. Med. Research Inst. Report NM 007039,* Report 21, Apr. 7.

Czepa, A. (1923-1924) Wachstumsfoerdernde und functionssteigernde Roentgen-
Radium-Wirkung. Strahlentherapie, 16: 913.

Davis, R. W., N. Dole, M. J. Izzo, and L. E. Young (1950) Hemolytic effect of radia-
tion. Observations on renal bile fistula dogs subjected to total body radiation
and on human blood irradiated in vitro. J. Lab. Clin. Med., 35: 528-537.

Desjardins, A. U. (1932) The radiosensitiveness of cells and tissues and some medical
implications. Science, 75: 569-575.

Dixon, F. J. (1948) Radiation induced hemorrhagic disease in chickens. Proc.
Soc. Exptl. Biol. Med., 68: 505-507.

Dowdy, A. H., L. R. Bennett, and S. M. Chastine (1950) Study of the effect of
varying oxygen tensions on the response of mammalian tissue to roentgen irradia-
tion. USAEC Report UCLA-55.*

Dudley, H. C, L. J. Louviere, and J. C. Shaw (1951) Effects of injection of radio-
gallium (Ga72). Nav. Med. Research Inst. Report NM 007 081.06.10,* Sept, 15.

Dunlap, C. E. (1942) Effects of radiation on the blood and the hemopoietic tissues,
including the spleen, the thymus and the lymph nodes. Arch. Path., 34: 562-608.

, J. C. Aub, R. D. Evans, and R. S. Harris (1944) Transplantable osteogenic

sarcomas in rats by feeding radium. Am. J. Path., 20: 1-21.

HEMATOLOGIC EFFECTS OF RADIATION 1085

Evans, T. C. (1948) Effects of small daily doses of fast neutrons on mice. Radi-
ology, 50: 811-833.

Finkle, R. D., R. H. Snyder, and P. C. Tompkins (1946) Acute radiotoxicity of
(Ba-La)140 in rats and mice. Part I. Preparation and administration of the
emitters. USAEC Report MDDC-1248.*

Furth, J., G. A. Andrews, R. H. Storey, and L. Wish (1951) The effect of X irradia-
tion on erythrogenesis, plasma and cell volumes. Soc. Med. J., 44: 85-92.

Gershon-Cohen, J., M. B. Hermel, and J. Q. Griffith (1951) The value of small lead
shields against the injurious effect of total-body irradiation. Science, 114: 157-
158.

Hagen, C. W., L. O. Jacobson, R. Murray, and P. Lear (1944) Effects of single
indirect doses of X rays on rabbits. USAEC Report MDDC-999. *

and R. E. Zirkle (1950) Comparative biological actions of cyclotron fast

neutrons and X rays. I. Lethal action in mice and rabbits. USAEC Report
CH-3903.*

Haley, T. J., and D. H. Harris (1949) The response of the guinea pig to 200 r acute

body X-irradiation. USAEC Report AECU-357.*
Hayer, E. (1934) Ergebnisse von experimentellen Studien am peripheren weissen

Blutbild nach Roentgenbestrahlung. Strahlentherapie, 50: 193-236.
Heineke, H. (1903) Ueber die Einwirkung der Roentgenstrahlen auf Tiere. Munch.

med. Wochnschr., 50: 2090-2092.

(1904) Ueber die Einwirkung der Roentgenstrahlen auf innere Organe.

Mlinch. med. Wochnschr., 51: 785.

(1905) Experimented Untersuchungen ueber die Einwirkung der Roent-
genstrahlen auf innere Organe. Mitt. Grenzg. Med. Chir., 14: 21-94.

Hektoen, L. (1915) The influence of the X ray on the production of antibodies.
J. Infectious Diseases, 17: 415-422.

(1918) Further studies on the effects of the roentgen ray antibody production.

J. Infectious Diseases, 22: 28-33.

Hennesey, T. G., and R. L. Huff (1950) Depression of tracer iron uptake curve in
rat erythrocytes following total body X-irradiation. Proc. Soc. Exptl. Biol.
Med., 73: 436-439.

Henshaw, P. S. (1943-1944a) Experimental roentgen injury. I. Effects on the
tissues and blood of C3H mice produced with single small whole body exposures.
J. Natl. Cancer Inst., 4: 477-484.

(1943-1944b) Experimental roentgen injury. II. Changes produced with

intermediate-range doses and a comparison of the relative susceptibility of
different kinds of animals. J. Natl. Cancer Inst., 4: 485-501.

, E. F. Riley, and G. E. Stapleton (1947) The biologic effects of pile radia-
tions. Radiology, 49: 349-360.
-, R. S. Snyder, E. F. Riley, G. E. Stapleton, and R. E. Zirkle (1946) Com-

parative late effects of single doses of fission neutrons and of gamma rays.

USAEC Report MDDC-1254.*
Ingram, M., and W. B. Mason (1950a) The effects of acute exposure to roentgen

radiation on the peripheral blood of experimental animals. USAEC Report

UR-122.*
and (1950b) The effects of chronic exposure to roentgen radiation

on the peripheral blood of experimental animals. USAEC Report UR-121.*
Isaacs, R. (1934) Relation to cell types in leukemia sensitivity to radiation. Folia

Haematol., 52: 414-425.
Jacobson, L. O. (1946) Acute radiotoxicity of (Ba-La)140 in rats and mice. Part V.

The effect of (Ba-La)140 on the hematological constituents of the peripheral

blood of rats and mice. USAEC Report MDDC-1261.*

1086 RADIATION BIOLOGY

Jacobson, L. 0. (1952) Evidence for a humoral factor (or factors) concerned in recov-
ery from radiation injury: a review. Cancer Research, 12: 315-325.

- and E. K. Marks (1947) Comparative action of cyclotron fast neutrons and
X rays. Part II. Hematological effects produced in the rabbit by fast neutrons.
USAEC Report MDDC-1372.*

— , , and E. O. Gaston (1951) Effect of mouse embryo tissue on survival

of irradiated mice. USAEC Report ANL-4625,* pp. 46-47.

-, , - - J. G. Allen, and M. H. Block (1948) The effect of nitrogen

mustard and X-irradiation on blood coagulation. J. Lab. Clin. Med., 33: 1566-
1578.

, , , and M. J. Robson (1952) Histologic studies of rabbits

exposed to 800 r total body radiation with lead shielding of a single Peyer's patch.
USAEC Report ANL-4794.*

-, , , , and R. E. Zirkle (1949) The role of the spleen in

radiation injury. Proc. Soc. Exptl. Biol. Med., 70: 740-742.

, , , E. L. Simmons, and M. H. Block (1948) Studies on radio-
sensitivity of cells. Science, 107: 248-250.

, , and E. Lorenz (1949) The hematological effects of ionizing radia-
tions. Radiology, 52: 371-395.

, , E. L. Simmons, and E. O. Gaston (1951) Comparative hematologic

studies of X-irradiated mice subjected to spleen shielding and related experi-
mental procedures. USAEC Report ANL-4713,* pp. 23-37.
- - - - -, C. W. Hagen, and R. E. Zirkle (1947) Effects of X rays on
rabbits. Part II. The hematological effects of total body X irradiation on the
rabbit. USAEC Report MDDC-1174.*

and M. J. Robson (1952) Factors effecting X-ray inhibition of antibody

formation. J. Lab. Clin. Med., 39: 169-175.
, , and E. K. Marks (1950) The effect of X-radiation on antibody

formation. Proc. Soc. Exptl. Biol. Med., 75: 145-152.
and E. L. Simmons (1946a) Acute radiotoxicity of injected Na24 for mice

and rats. II. The effect of the Na24 on the leucocytes of the peripheral blood of

mice. USAEC Report AECD-2036. *
- and - - (1946b) Effects of insoluble ingested Y91. VIII. Hematological

effects. USAEC Report AECD-2037. *
and - (1946c) Studies of the metabolism and toxic action of injected

radium. Part II. The hematological effects of parenterally administered

radium. A comparison of plutonium and radium effects. USAEC Report

AECD-2372.*

— , — — , W. F. Bethard, E. K. Marks, and M. J. Robson (1950) The influence

of the spleen on hematopoietic recovery after irradiation injury. Proc. Soc.

Exptl. Biol. Med., 73: 455-459.

— , , and M. H. Block (1949) The effect of splenectomy on the toxicity

of Sr89 to the hematopoietic system of mice. J. Lab. Clin. Med., 34: 1640-1655.

— - — , E. K. Marks, and J. H. Eldredge (1951) Recovery from radiation

injury. Science, 113: 510-511.

E. O. Gaston, M. J. Robson, and J. H. Eldredge (1951)

Further studies on recovery from radiation injury. J. Lab. Clin. Med., 37:

683-697.

— - — - — M. J. Robson, W. F. Bethard, and E. O. Gaston (1950).

The role of the spleen in radiation injury and recovery. J. Lab. Clin. Med., 35:

746-770.

— , E. Skirmont, E. K. Marks, E. Gaston, and M. Block (1948) The effect of

HEMATOLOGIC EFFECTS OF RADIATION 1087

total-body X irradiation and P32 on the peripheral blood of normal rats and
rats with cobalt-induced polycythemia. USAEC Report ANL-4147,* pp. 22-
32.

S. P. Steamer, and E. L. Simmons (1947) The effect of folic acid on radiation

induced anemia and leukopenia. J. Lab. Clin. Med., 32: 1425.
Kahn, J. B., and J. Furth (1952) The pathogenesis of postirradiation anemia.

USAEC Report ORNL-1186.*
Kroemeke, F. (1926) Ueber die Einwirkung der Roentgenstrahlen auf die roten

Blutkoerperchen. Strahlentherapie, 22: 608-652.
Lacassagne, A., and J. Lavedan (1924) Les modificationes histologiques due sang

consecutives aux irradiationes experimentales. Paris med., 51: 97-103.
Lawrence, J. H., P. C. Aebersold, and E. O. Lawrence (1936) Comparative effects

of X rays and neutrons on normal and tumor tissue. Proc. Natl. Acad. Sci.

U.S., 22: 543-557.
and E. O. Lawrence (1936) The biological action of neutron rays. Proc.

Natl. Acad. Sci. U.S., 22: 124-133.
- and associates (1949) Quoted in E. P. Cronkite, The hematology of ionizing

radiations, in Atomic medicine, C. F. Behrens, 1st ed. Thomas Nelson & Sons,

New York, p. 109.

— , A. H. Dowdy, and W. N. Valentine (1948) The effects of radiation on

hemopoiesis. Radiology, 51: 400-413.

W. N. Valentine, and A. H. Dowdy (1948) The effect of radiation on

hemopoiesis. Is there an indirect effect? Blood, 3: 593-611.

Linser, P., and E. Helber (1905) Experimentelle Untersuchungen ueber die
Einwirkung der Roentgenstrahlen auf das Blut und Bemerkungen ueber die
Einwirkung von Radium und ultraviolettem Lichte. Deut. Arch. klin. Med., 83:
479-498.

Lorenz, E. (1951) Recovery pattern of the blood picture in guinea pigs following a
limited total body exposure to chronic gamma radiation. J. chim. phys., 48:
264-274.

— , C. C. Congdon, and D. Uphoff (1952) Modifications of acute irradiation
injury in mice and guinea pigs by bone marrow injections. Radiology, 58: 863
877.

— , A. Eschenbrenner, M. Deringer, and W. E. Heston (1946) Biologic action
of gamma and X rays. I. Exposure of mice to daily doses of gamma radiation at
two rates: 5.5 r/hr and 0.11 r/8 hr. J. Natl. Cancer Inst., 6: 349-353; Cancer
Research, 6: 485.

— , W. E. Heston, and A. B. Eschenbrenner (1947) Biological studies in the
tolerance range. Radiology, 49: 274-285.

— , - -, L. O. Jacobson, A. B. Eschenbrenner, M. B. Shimkin, M. K. Deringer,

J. Doniger, and R. Schweisthal (1946) Effects of long-continued whole body
irradiation with gamma rays on mice, guinea pigs, and rabbits. Parts I-IV.
USAEC Report MDDC-653, * pp. 654-656.

— , L. O. Jacobson, and H. Sutton (1950) Part I. Survival and blood picture of
hybrid guinea pigs exposed to long-continued total-body gamma radiation.
USAEC Report ANL-4401,* pp. 38-52.

— , - , , and R. Schweisthal (1950) Part II. Recovery from anemia

induced in guinea pigs by a limited exposure to chronic total-body gamma radia-
tion. USAEC Report ANL-4401,* pp. 53-71.

— , D. Uphoff, T. R. Reid, and B. Shelton (1951) Modifications of irradiation
injury in mice and guinea pigs by bone marrow injection. J. Natl. Cancer Inst.,
12: 197-201.

1088 RADIATION BIOLOGY

— , , and H. Sutton (1949) The 30-day LD50 and accompanying blood

picture of hybrid guinea pigs. USAEC Report ANL-4333,* pp. 57-65.
Minot, G. R., and R. Spurling (1924) Effect on blood of irradiation, especially short

wave length roentgen ray therapy. Am. J. Med. Sci., 168: 215-241.
Mottram, J. C, and S. Russ (1921) Lymphopenia following exposure of rats to

"soft X rays and beta rays of radium." J. Exptl. Med., 34: 271.
Murphy, J. B. (1926) The lymphocyte in resistance to tissue grafting. Monograph

21, Rockefeller Institute for Medical Research.
Murray, R., M. Pierce, and L. O. Jacobson (1948) The histological effects of X-rays

on chickens with special reference to the peripheral blood and hemopoietic

organs. USAEC Report AECD-2303.*
Patt, H. M., D. E. Smith, and E. Jackson (1950) The effect of cysteine on the

peripheral blood of the irradiated rat. Blood, 5: 758-763.
-, R. L. Straube, E. B. Tyree, M. N. Swift, and D. E. Smith (1949) Influence

of estrogens on the acute X-irradiation syndrome. Am. J. Physiol., 159: 269.
, E. B. Tyree, R. L. Straube, and D. E. Smith (1949) Cysteine protection

against X irradiation. Science, 110: 213-214.
Prosser, C. L., E. E. Painter, H. Lisco, A. M. Brues, L. O. Jacobson, and M. N. Swift

(1947) The clinical sequence of physiological effects of ionizing radiation in

animals. Radiology, 49: 269-365.
-, , and M. N. Swift (1946) Physiology of dogs exposed to single total-
body doses of X rays. USAEC Report CH-3738. *
Raper, J. R., and K. K. Barnes (1951) The effects of external irradiation with beta

rays on the peripheral blood of rabbits, in Biological effects of external beta

radiation, R. E. Zirkle, ed. McGraw-Hill Book Company, Inc., New York,

National Nuclear Energy Series, Div. IV, Vol. 22E, Chap. 10.
, P. S. Henshaw, and R. S. Snider (1951) Effects of periodic total-surface

beta irradiation, in Biological effects of external beta radiation, R. E. Zirkle, ed.

McGraw-Hill Book Company, Inc., New York, National Nuclear Energy Series,

Div. IV, Vol. 22E, Chap. 14.
-, R. E. Zirkle, and K. K. Barnes (1951) Comparative lethal effects of external

beta irradiation, in Biological effects of external beta radiation, R. E. Zirkle, ed.

McGraw-Hill Book Company, Inc., New York, National Nuclear Energy Series',

Div. IV, Vol. 22E, Chap. 3.
Rekers, P. E. (1949) Transplantation of bone marrow into dogs that have received

total body single dose radiation. USAEC Report AECD-1966.*
, M. P. Coulter, and S. L. Warren (1950) Effect of transplantation of bone

marrow into irradiated animals. Arch. Surg., 60: 635-667.
Ross, M. H., J. Furth, and R. R. Bigelow (1952) Capillary fragility caused by

ionizing radiations. Changes in cellular composition of the lymph. Blood,

7: 417-428.
Russ, S. (1921) The immediate effect of X rays on the blood lymphocytes. Arch.

Radiol. Electrotherapy, 26: 146.

— , H. Chambers, and G. M. Scott (1921) Further observations of the effect

of X rays upon the lymphocytes. Arch. Radiol. Electrotherapy, 25: 377.

-, and J. C. Mottram (1919) Experimental studies with small

doses of X-rays. Lancet, 196: 692.
Sacher, G. A., and N. Pearlman (1947) Comparative action of cyclotron neutrons

and X-rays. Part III. Statistical analysis of blood data. USAEC Report

MDDC-1387.*
Salisbury, P. E., P. E. Rekers, J. H. Miller, and N. F. Marti (1951) Effect of early

cross transfusion on X-irradiated dog. Proc. Soc. Exptl. Biol. Med., 78: 226-229.

HEMATOLOGIC EFFECTS OF RADIATION 1089

Schack, J. A., and R. C. MacDuffee (1949) Increased radioresistance of red bone

marrow after anoxia. Science, 110: 259-260.
Schwartz, S. E., E. Katz, L. M. Porter, L. O. Jacobson, and C. J. Watson (1947)

Studies of the hemolytic effect of radiation. USAEC Report CH-3760.*
Selling, L., and E. E. Osgood (1938) Action of benzol, roentgen rays and radioactive

substances on the blood and blood-forming tissues, in Downey's handbook of

hematology. Paul B. Hoeber, Inc., Medical Book Department of Harper &

Brothers, New York, Vol. 4, pp. 2693-2801.
Shouse, S., S. L. Warren, and G. H. Whipple (1931) Aplasia of marrow and fatal

intoxication in dogs produced by roentgen radiation of all bones. J. Exptl. Med.,

52: 421-435.
Siegel, P. W. (1920) Die Veranderungen des Blutbildes nach gynakologisehen

Rontgen-, Radium- and Mesothoriumtiefenbestrahlung und ihre klinische

Bedeutung. Strahlentherapie, 11: 64-139.
Simmons, E. L., and L. O. Jacobson (1946) Radiotoxicity of injected Sr89 for rats,

mice, and rabbits. Part IV. The hematological effects of enterally and paren-

terally administered Sr89 in mammals. USAEC Report MDDC-1387.*
, , and E. K. Marks (1950-1951) Effect of pre-injection of estradiol

benzoate plus spleen shielding on the survival of mice following X irradiation.

USAEC Report ANL-4571.*

-, N. Pearlman, and C. L. Prosser (1946) The effectiveness of drugs

in preventing or alleviating X-ray damage. USAEC Report MDDC-1277.*
Smith, W. W., R. Dooley, and E. O. Thompson (1948) Simulated high altitude

following whole-body radiation of mice. J. Aviation Med., 19: 227-237.
Steamer, S. P., E. L. Simmons, and L. O. Jacobson (1947a) The effects of single

dose total body X irradiation on the peripheral blood of the rat. Anat. Record,

99: 576-577.
( , and (1947b) The effects of total body X-irradiation on the

peripheral blood and blood-forming tissues of the rat. USAEC Report MDDC-

1319.*
Storey, R. H., L. Wish, and J. Furth (1950) Changes in cell and plasma volumes

produced by total body X irradiation. Proc. Soc. Exptl. Biol. Med., 74: 242-244.
Suter, G. M. (1947) Response of hematopoietic system to X rays. USAEC Report

MDDC-824.*
Swift, M. N., C. L. Prosser, and E. S. Mika (1946) Effects of Sr89 and X-radiation

on goats. USAEC Report AECU-108.*
Swisher, S. N., and F. W. Furth (1951) The effect of early small exchange trans-
fusion on the X-irradiated dog. Proc. Soc. Exptl. Biol. Med., 78: 226-229.
Talbot, J. M., and H. B. Gerstner (1951) Bone marrow implants in the treatment

of radiation sickness. USAF School of Aviation Med., Project 21-47-001.*
- and E. A. Pinson (1951) The experimental use of bone marrow in acute

radiation injury. Military Surgeon, 108: 412-417.
Taylor, H. D., W. D. Witherbee, and J. B. Murphy (1919) Studies on X ray effects.

I. Destructive action on blood cells. J. Exptl. Med., 29: 53-73.
Thomas, M. M., W. D. Witherbee, and J. B. Murphy (1919) II. Stimulative action

on the lymphocytes. J. Exptl. Med., 29: 75.
Treadwell, A., W. U. Gardner, and J. H. Lawrence (1943) Effect of combining

estrogen with lethal doses of roentgen-ray in Swiss mice. Endocrinology, 32:

161-164.
Valentine, W. N., W. D. Adams, and J. S. Lawrence (1947) Blood platelets and rate

of utilization in the cat. Blood, 2: 40-49.
and M. L. Pearce (1952) Studies on the radiosensitivity of bone marrow.

1090 RADIATION BIOLOGY

I. The relative sensitivity of erythroid and myeloid elements. Blood, 7: 1-13.
Warren, S. L., (1943) Effects of radiation on normal tissue. Arch. Path., 35: 121.
and D. J. Dixon (1949) Effects of continuous radiation on chick embryos

and developing chicks. II. Bone marrow, lymphoid and peripheral blood.

Radiology, 52: 869-882.

and G. H. Whipple (1922) Roentgen ray intoxication. II. A study of the

sequence of clinical, anatomical, and histological changes following a unit dose of

X ray. J. Exptl. Med., 35: 203-211.
Wheeler, B., M. A. Jackson, and P. Hahn (1951) Hematology of the dog following

intravenous administration of radioactive colloidal gold. Am. J. Physiol., 166:

323-327.
Wuensche, H. W. (1938) Fortlaufende Untersuchungen ueber des Einflusses der

Roentgenstrahlen auf das Knochenmark. Arch. Exptl. Path. Pharmakol., 189:

581.
Zirkle, R. E. (1947) Components of the acute lethal action of slow neutrons. Radi-
ology, 49: 271-273.
(1950) Radiobiological additivity of various ionizing radiations. Am. J.

Roentgenol. Radium Therapy, 63: 170-175.

Manuscript received by the editor Mar. 15, 1952

CHAPTER 17

Histological Changes after Irradiation

William Bloom

Department of Anatomy and Institute of Radiobiology and Biophysics,

The University of Chicago

and Margaret A. Bloom

Chicago

Introduction. Visible changes in cells: Cell membrane — Organelles — Interphase
nucleus — Dividing cells. Changes in tissues. Bone marrow. Lymphatic organs.
Thymus. Spleen. Bone. Cartilage. Male generative system. Female generative
system. Skin: Epithelium — Derma. Epidermal derivatives: Hair — Nails — Skin glands
— Mammary gland. Gastrointestinal system: Mouth, nasopharynx, and esophagus—
Stomach and intestines — Salivary glands — Liver — Gall bladder and biliary passages —
Pancreas. Urinary system: Kidney — Urinary bladder. Respiratory system. Nervous
system. Eye. Endocrine glands: Adrenal — Hypophysis — Thyroid — Parathyroid. Dis-
cussion. References.

INTRODUCTION

Within the first ten to fifteen years after their discovery, it was shown
that ionizing radiations, in amounts we recognize today as under 1500 r,
would produce marked degenerative changes in the skin and its append-
ages, parts of the gastrointestinal tract, the blood-cell-forming organs
(and thus in the blood), the gonads, conjunctiva and lens, bone, and
cartilage. It was also found that the skin might become ulcerated and
even cancerous. Moreover, it was learned that irradiation could destroy
some cancers and could produce embryological monsters. In the years
which have elapsed since then much knowledge has been added, but
much more is required before the visible effects of radiation on cells can
be presented in a complete and systematic fashion.

Advances have been retarded by a number of factors. One of these is
our inability to evaluate dosages in the older reports before the introduc-
tion of the roentgen. Another is the difficulty of comparing the effects on
a particular organ of total-body irradiation with local irradiation of that
organ, since systematic studies on this theme are lacking. Further
variables which must be taken into account are the time-intensity factor
for a given dosage and, closely related to this variable, whether the radia-

1091

1092 RADIATION BIOLOGY

tion is given at one time or in divided doses. Finally, medical interest in
radiation has focused much of the investigation on the higher animal
forms and on a relatively limited dose range determined by therapy in
man and toxicity and survival studies in animals.

Depending in part on the amount of radiation applied and in part on
the degree of susceptibility of the cells irradiated, the resulting cellular
changes may be classified roughly into two groups. The first comprises
subtle effects recognizable only by alterations in the actual and potential
functions of the cells (including those which become manifest only in
their progeny). The second group consists of more obvious changes,
detectable by the optical microscope: (1) degenerative phenomena which
may be reversible but which usually lead to cell death, (2) inhibition of
mitosis, and (3) abnormal mitosis. Other morphological criteria must be
sought especially through study of intracellular enzymes and of changes
in submicroscopic structure.

Since all cells near an embedded radium needle will die if exposed long
enough, there are no completely resistant cells. But with amounts of
radiation which permit survival of laboratory animals for only a short
time, many cells appear to be unaffected. Among the more sensitive
cells, however, gradations in susceptibility are revealed.

In a number of organs the visible effect of a given dose of radiation is
the death of a given type of cell, while other cell types may show no
obvious damage although some of them may undergo great changes of
another kind. It is thus impossible to compare their reaction to radia-
tion using cell death as the sole criterion. For instance, spermatogonia
in most mammals are destroyed by exposure to 800 r (total-body irradia-
tion with 200-kv X rays) while spermia may show no obvious change.
Yet the spermia may have been injured, although the effects will appear
later only if one of them fertilizes a normal egg. For the other cells of
the testis (spermatocytes, spermatids, Sertoli cells, and interstitial cells)
we can find no comparison of relative radiosensitivities, as revealed by
cellular degeneration, beyond the observation that they are much more
resistant than the spermatogonia. Similar statements can be made
about the other organs, both the sensitive and the more resistant ones.
We are thus in a position to evaluate only roughly the relative radio-
sensitivity of a few types of cells while the other cells of the body may be
grouped together as comparatively radioresistant. The data so far
determined do not permit an evaluation of all cell types of the body in
exact terms such as might be expressed by LD.,o or LDioo values for those
cells. This lack was stressed by Stafford L. Warren (Duggar, 1936,
p. 475).

Although relative radiosensitivity is a convenient term for distinguishing
varying degrees of susceptibility shown by the different parts of an
organism to the same amount of radiation, it is meaningless unless the

HISTOLOGICAL CHANGES AFTER IRRADIATION 1093

cells or tissues are being compared for the same end effect of radiation and
unless the physical conditions of irradiation are the same. The idea of
radiosensitivity of a given cell (or organism) must be further qualified to
include consideration of its functional state at the time of irradiation and,
in many cases, its present or future reproductive activity, and the species
and age of the animal to which it belongs. Other terms should be used
for those conditions which do not satisfy these criteria.

In contrast to a few types of injuries which cause immediate damage to
cells (thermal burn, for example) the effects of irradiation, like those of
many toxic agents, become apparent only after some time has elapsed.
This interval may vary from a few minutes to months or years, depending
mainly on the cell type, and the criterion of the injury. For instance,
vast numbers of lymphocytes are destroyed in a few hours by 600 to 800 r
of 200-kv X rays, while cells of the epidermis may not die for several
days after three times this amount of radiation, or months or years may
elapse before cataracts develop in the lens.

With the possible exception of certain effects due to specific ionization,
the results of the several kinds of ionizing radiations from external sources
given in equivalent amounts are identical (W. Bloom, 1948a). The same
biological changes as those produced by X rays and neutrons result when
the radiations are from internal sources, but these vary with the distribu-
tion of the isotope (the chemical nature of the element determining its
localization, being diffuse in the case of Na24 or highly localized with Sr89),
the type of "particle" given off, and the half-life of the isotope (and its
daughters). The continuous nature of internal irradiation is in contrast
to that from most external sources which only exceptionally have been
applied continuously.

The cold-blooded vertebrates are more resistant than the warm-blooded
ones and, going down the scale of animal (and plant) complexity, the
forms become increasingly resistant. Nevertheless, the kinds of cellular
changes seen in vertebrate cells also occur in lower forms, although the
amount of radiation necessary to elicit these changes will usually be much
greater. The younger the animal the more susceptible it will be; this is
also true for its cells, at least for extremes in age. It is well known that
embryos are much more sensitive than postnatal animals and that resist-
ance increases progressively with age. For instance, the nerve sheath
cells in a three-week chick suffer great damage with 800 r total-body
irradiation, whereas these same cells are not damaged with this dose when
the animal is 11 weeks old (Snider, 1948b). Changes in temperature and
metabolic rate affect the sensitivity of a particular cell strain, as shown by
greater resistance at lower temperatures and when the circulation of
blood is interrupted during irradiation.

It must be kept in mind that all the effects of ionizing radiations on
cells, ranging from nonvisible effects on chromosomes to rapid cell death,

1094 RADIATION BIOLOGY

can be brought about by certain chemical agents. This is true, for
instance, for the hematopoietic organs, intestine, and testis after injection
of appropriate amounts of the nitrogen or sulfur mustard gases. These
and many other substances are also potent mutagenic agents. Moreover,
benzol has for many years been known to deplete the blood-forming
tissues in much the same fashion as X rays.

Although there is nothing known that is specific for radiation effects,
it is often possible within certain dose ranges to diagnose radiation
damage. Thus, "The combination of two or more of the nonspecific
characteristics of radiation is strongly presumptive evidence that the
injury is in fact the result of radiation. Thus the combination of giant
and irregular nuclei, hyaline connective tissue, and thick-walled hyalin-
ized blood vessels would be difficult to explain on any other basis than
that of a late response to irradiation " (Shields Warren, 1944). However,
the acute changes due to nitrogen mustard and irradiation are identical.

For detailed consideration of the several thousand papers dealing with
changes from irradiation from external and certain internal sources, the
reader is referred to a number of extensive reviews (W. Bloom, 1948a;
Colwell, 1935; Desjardins, 1924, 1926, 1928, 1931; Dunlap, 1948; Ellinger,
1941; Giese, 1947; Lacassagne and Gricouroff, 1941; Packard, 1931;
Regaud and Lacassagne, 1927 ; Shields Warren, 1942, 1943, 1944; Stafford
L. Warren, 1936). Here we can only attempt to summarize the more
important of these changes. Whenever possible, the data emphasized
will be from experiments on laboratory animals, where control of dosage,
intervals, and a large number of subjects under identical conditions is
possible. In most of the organ systems the effects of moderate single
doses of external irradiation will be described first, followed by the changes
resulting from smaller or larger doses, then those after repeated small
doses, and finally the effects of internal irradiation.

VISIBLE CHANGES IN CELLS

The first changes of irradiation are usually manifest as nuclear changes,
often those involving the mitotic process. Most of the evidence since
1903 (Perthes, 1903) indicates that the nucleus is more sensitive to
irradiation than the cytoplasm. This view has received direct experi-
mental confirmation in the comparison of the effects of irradiation of
cytoplasm plus nucleus with cytoplasm alone in fern spores (Zirkle, 1932).
Similar conclusions were reached on Arbacia eggs (Henshaw, 1938).
However, the opposing view that nuclear changes are secondary to
changes in the cytoplasm has been strongly maintained (Duryee, 1949;
see also Chap. 10 by Giles and Chap. 11 by Carlson).

Cell Membrane. There have been several reports that the cell mem-
brane is damaged by irradiation. The most convincing instance, in the

HISTOLOGICAL CHANGES AFTER IRRADIATION 1095

authors' opinion, is that of cells observed in tissue culture after enormous
dosages of irradiation. Here it has been seen that vesicles may form at
the edge of the cell and that changes in form of the membrane, as in
ameboid motion, may cease. In all such cases the changes in the cell
membrane have followed severe, irreversible damage to the nucleus. It
has been reported that both nucleated and nonnucleated red cells swell
after irradiation, and this has been interpreted as being due to injury to
the cell membrane. However, the swelling may be simply a result of
passage of water through the membrane because of an increased number
of ions in the cell. Loss of stainability of the membrane has also been
reported. The few instances in which this occurs probably are late
stages after high dosage. Certainly it may be accepted that in the great
mass of irradiated cells exposed to 1000 to 3000 r, the cell membrane does
not lose its stainability. Early swelling of cells as a whole occurs more
strikingly in some cell types than in others. It may also appear late, as
in the fibroblasts of inflammatory tissue.

There is evidence in certain plants and in some cells in tissue culture of
an immediate decrease in viscosity of the cytoplasm after intense
irradiation. Some cells in culture have been observed to slow down,
others to move faster, at least for a time. Cytoplasmic movements
may proceed for many hours after the nucleus is obviously coagulated as
a result of intensive irradiation.

Organelles. It has been postulated that destruction of the centrosome
with its diplosome will prevent the cell from dividing. The proof for this
is absent, although it is presumably correct. Ciliary motion stops after
irradiation. It has been claimed that changes in the mitochondria are the
first morphological evidences of radiation effects. This conclusion has
been denied by others on the basis of sectioned material (W. Bloom,
1948b; Fogg and Shields Warren, 1937) and on observations of living
cells irradiated in tissue culture (W. Bloom, unpublished observations).
We have found no evidence that mitochondria degenerate early; on the
contrary, they may persist unchanged long after the cells are obviously
undergoing irreversible degenerative changes.

Minor changes in the Golgi net after irradiation of tumor cells have
been described (Fogg and Shields Warren, 1937). This subject should
be investigated further.

Neurofibrils and myofibrils occur in cells that are among the most
radioresistant in the adult animal. We have found no reports on changes
in these fibrils as a result of irradiation with therapeutic dosages or
with small multiples thereof. Also, no characteristic changes have been
described in cellular inclusions (fat, carbohydrates, chromophil substance,
pigment granules). However, it has been noted that, in many types of
cells after irradiation, there may be an increase in fat content, and there
are also reports on purported changes in the glycogen in liver cells. Such

1096 RADIATION BIOLOGY

changes are probably secondary to effects produced in other organs. On
the other hand, iron pigment from ingested erythrocytes accumulates in
macrophages in the reparative phase after irradiation, and brown pigment
develops in the skin.

Inter-phase Nucleus. One of the first changes to be seen in sections as
well as in living tissue cultures is a clumping of chromatin. This may
occur in both sensitive and resistant cell strains. It is probably reversi-
ble, since it may be seen in all types of cells of the body a short time after
irradiation and, depending on the dosage and the cell type, may or may
not disappear after a few hours. Other changes, probably not reversible,
include vacuolization of chromatin particles and nucleoli and thickening
of the nuclear membrane. In the course of the next few hours such cells
may show progressive pyknosis, the nuclear material condensing into
one, two, or three large chromatin masses (with or without vacuoles).
Sometimes the nuclei may disintegrate without passing through a
pyknotic stage. There is nothing specific about these nuclear changes
after irradiation, for similar effects may be produced by a variety of toxic
agents.

In addition to these changes, nuclei may show marked budding in
certain cell types (hepatic epithelium, megakaryocytes), particularly
after long-continued action of radioactive isotopes accumulating in the
organ. Multilobular nuclei may be found in myelocytes after irradia-
tion. The swelling of nuclei is believed to be one of the most constant
results of irradiation (Failla and Sugiura, 1939). It is clearly seen in the
intestinal or gastric epithelium during the regenerating phase (Pierce,
1948). A pronounced swelling of the nucleus has not been a striking
feature of irradiated cells in tissue culture (W. Bloom, unpublished
observations).

Dividing Cells. One of the most striking nuclear effects of irradiation
is the rapid cessation, after 200 to 800 r, of practically all mitosis in all
the organs which normally are in a state of continuous regeneration.
This interference with cellular division may be apparent about \-i to 1
hour after irradiation and may last 12 to 15 hours or much longer. The
cause of this inhibition is not known. In many tissues its duration
varies roughly with the dose. It is equally remarkable that "not all
mitoses are inhibited, even in a heavily irradiated tissue culture" (Shields
Warren, 1942, 1943). Most cells which had undergone the first morpho-
logically evident mitotic changes seem to go through the rest of the
process, although they may show lagging or otherwise damaged chromo-
somes. Fragmented chromosomes are thought to heal either with or
without visible defects, but, in all such cases, genie material is undoubt-
edly lost. In total mounts of the cornea of amphibian larvae a marked
clumping and eventual pyknosis of the chromatin at any stage of mitosis

HISTOLOGICAL CHANGES AFTER IRRADIATION 1097

have been described as a primary effect of radiation (Alberti and Politzer,
1923) . Some of these dumpings may be pseudo-amitotic. Then, follow-
ing the period of inhibition of mitosis, the secondary effect appears,
manifested by karyorrhexis in cells which were resting at the time of
irradiation. In animals given a second irradiation 10 or 15 days after
the first, the primary effects were complicated by secondary effects from
the first irradiation.

Many of the cells which are in a resting state at the time of irradiation,
as well as some that are in mitosis, may show .their first evidence of
damage when they divide some time later and develop visible chromo-
somal defects and abnormalities. A high percentage of such cells may
die during this mitosis (Halberstaedter and Back, 1942; Kemp and Juul,
1930 ; Strangeways and Hopwood, 1926) . In some instances, the chromo-
somes may become "sticky" and pairs of chromosomes may adhere to
each other (Giese, 1947). If this occurs throughout the chromosomal
content of a cell, polyploidy may result.

CHANGES IN TISSUES

Nerve and muscle, two of the four major tissues, are relatively un-
affected by doses of radiation which elicit wide ranges of susceptibility in
the other two primary tissues, the various connective tissues and epi-
thelia. Some of the differences in sensitivity can be explained by the
fact that, in some tissues, resting cells and cells about to divide may
not show effects of damage until they divide. For example, in the
epidermis there are comparatively few cells dividing at any one moment,
and the full effects of irradiation do not become manifest until many of
the resting cells divide. Hepatic epithelium is normally untouched by
relatively large amounts of radiation. But if a large part of the liver is
removed, the rest will regenerate with great numbers of mitoses and
irradiation will affect the organ.

The connective tissues differ from the other tissues in two important
respects: (1) the preponderance of intercellular substance, and (2) an
intimate relation to the blood vascular system (although cartilage and
parts of bone are exceptions). In all organs, to a greater or less extent,
connective tissue separates the blood vessels from the parenchyma.
Vascular connective tissue throughout the body is the tissue which
reacts, by inflammation, to local injury. Much has been written about
the effects of radiation on intercellular substances (see section on the
derma). In the hematopoietic tissues the free cells are many times more
susceptible to damage by irradiation than the cellular stroma. The
adipose, pigment, tendinous, and mucous connective tissues have not
been studied sufficiently for radiation effects.

1098

RADIATION BIOLOGY

Fig. 17-1. Bone marrow of rabbit after total-body irradiation with 800 r of 200-kv
X rays. The marrow at 24 hours (A) shows mainly various stages of granulocyte
formation. The arrows point to debris, mostly of precursors of red blood cells. At 9
days (B) the marrow, completely depleted of hematopoietic cells, contains some fat

HISTOLOGICAL CHANGES AFTER IRRADIATION 1099

BONE MARROW

The framework of active mammalian marrow (myeloid tissue) con-
sists of blood sinusoids lined with phagocytic littoral cells and a loose
stroma of reticular fibers with embedded fat cells and primitive and
phagocytic reticular cells. In the meshes of this framework are closely
packed groups of free cells: hemocytoblasts (free stem cells), erythro-
blasts (precursors of red blood corpuscles), myelocytes (precursors of
granular leukocytes), and megakaryocytes. In the normal adult hemo-
cytoblasts are rare.

Ionizing radiations cause destruction of the free hematopoietic cells,
and their replacement by dilated sinuses and a gelatinous or fatty type of
marrow (M. A. Bloom, 1948; Dunlap, 1948; Heineke, 1903; Lacassagne
and Gricouroff, 1941; Martland, 1931; Pappenheim and Plesch, 1912;
Tullis, 1949). The bone marrow is much more susceptible to acute
radiation damage than many of the other organs, and the changes pro-
duced are easily detectable at the LD50 30-day level and, in some species,
even well below this (M. A. Bloom, 1948). Thus, after 800 r total-body
X irradiation in the rabbit, initial damage is revealed by a cessation of
mitosis half an hour after treatment and a progressive degeneration of
hematopoietic cells, already striking at 3 hours. Some cells die very
soon, as evidenced by pyknosis and fragmentation of their nuclei, and
are then phagocytosed by macrophages; others assume abnormal forms
with swollen or distorted, often multilobed, nuclei, and, after an interval,
these too may degenerate. The first of the blood-forming cells to be
destroyed are the erythroblasts and hemocytoblasts, next the myelocytes,
and finally the megakaryocytes. In each series the younger cells are
more sensitive than the older forms of the same cell type. Thus, in the
red-cell-forming series the basophil erythroblasts are more sensitive than
the polychromatophil, and these in turn than the orthochromatic ones.
There is also some erythrophagocytosis during this period. By 4 or 5
days the marrow is almost completely devoid of immature red and white
blood corpuscles. At no stage is there any visible damage to reticular
cells.

The acute destructive effects on the myelocytes are quickly reflected
in the drop in granular leukocytes in the circulating blood. However,
the acute destruction of erythroblasts is not reflected in a similar rapid
drop in the erythrocytes of the blood, since the latter are much longer
lived. The effects of irradiation on the peripheral blood cells and blood

cells and a gelatinous intercellular substance, in addition to an occasional macrophage
(as in the center of the field) and some collapsed vessels. (C) After 10 days, the mar-
row shows a focus of regenerating hematopoiesis. Note the exceedingly large, young
cells with darkly stained cytoplasm (basophilia) and the great prominence of the
nucleoli. These cells are somewhat larger than the normal. 1030 X.

1100 RADIATION BIOLOGY

platelets are discussed in Chap. 16 (by Jacobson). In contrast to the
radiosensitivity of the free hematopoietic cells is the great resistance of
macrophages, reticular cells, and fat cells, none of which show any evi-
dence of damage even with 2000 r or more. This is important since the
reticular cells are more primitive than the free stem cells and are the
most important source of regenerating hematopoietic cells (W. Bloom,
1948a).

First attempts at regeneration of hematopoietic cells, sometimes during
the first week after irradiation, are often abortive at the LD50 level of
total-body X irradiation, but full recovery eventually takes place some
weeks later. Regenerating erythroblasts often contain highly con-
stricted nuclei, and regenerating myelocytes may be greatly enlarged.
Regeneration follows a definite pattern: the gelatinous intercellular sub-
stance is largely replaced by fat cells and the dilated venous sinuses
resume their normal size. A proliferation of stem cells — hemocytoblasts
—arising from mitosis of their own cell kind or by transformation of
reticular cells (homoplastic or heteroplastic origin) is followed by active
erythropoiesis, later by myelocytopoiesis, and finally by the return of
young megakaryocytes. After lower doses of X rays, 400 to 100 r,
cellular destruction is less extensive and recovery occurs earlier. The
effect of 50 r is scarcely perceptible. The morphological and temporal
details of regeneration after very high doses of localized radiation have
not been worked out.

X rays affect the bone marrow much less when administered at the
rate of 80 r per day for several weeks than when the same quantity of
radiation is given in a single dose (M. A. Bloom, 1948). Daily exposure
of mice to external y rays reveals no changes in the marrow with 1.1 r
for 16 months (Eschenbrenner, 1946; Spargo et at., 1951). An increased
gelatinous replacement of the cellular elements of the marrow, particu-
larly in the metaphysis, begins at four months after 8.8 r per day and at
six months after 4.4 r per day. The number of immature cells of the
myelocyte series (particularly hemocytoblasts) increases progressively
from 8 to 16 months in the 8.8-r series. These animals also show a
progressive increase in connective tissue mast cells in the marrow (Spargo
et at., 1951). In guinea pigs 8.8 r per day produces a low-grade anemia
with a sudden terminal ending after some months (Lorenz, Heston,
Jacobson, et al., 1953).

A similar pattern of damage and repair results from equivalent doses
of X rays, fast neutrons, and slow neutrons (M. A. Bloom, 1948; Lawrence
and Tennant, 1937). With all types of radiation, damage to the bone
marrow is roughly proportional to the size of the dose.

All bone-seeking isotopes tend to accumulate at the growing ends of
bones and produce earlier and more severe damage in the marrow there
than in the center of the shaft. A somewhat similar but less striking

HISTOLOGICAL CHANGES AFTER IRRADIATION 1101

gradient of injury occurs with X rays and, to a less extent, with slow
neutrons, but not with fast neutrons. Secondary rays, emitted by the
bone spicules of the metaphysis after X irradiation and to a less extent
after slow-neutron bombardment, may perhaps account for these differ-
ences.

Both depletion and regeneration take place more slowly in response to
the radioactive isotopes than to any of the external ionizing radiations.
Slow degeneration explains the absence of cellular debris, and delayed
regeneration is undoubtedly the effect of the continued bombardment by
long-lived isotopes.

In addition to the degenerative changes characteristic of bone marrow
subjected to external irradiation, an atypical fibrous bone develops from
spindle cells in the marrow cavity after administration of many of the
radioactive isotopes. This will be described in the section on bone.

Study of aspirated vertebral marrow from atomic bomb casualties at
Hiroshima disclosed the disappearance, within the first week, of myeloid
tissue and the presence of atypical cells, many of them resembling plasma
cells. These findings are confirmed and extended by histological sections
of the long bones and sternum obtained at autopsy, in which multiplica-
tion of reticular cells is seen at 12 days (Liebow et al, 1949). The failure
to find in the Hiroshima material the early degeneration of erythroblasts,
previously described on the basis of closely spaced specimens from
laboratory animals (M. A. Bloom and W. Bloom, 1947), probably rests
on the scanty material obtained during the first weeks after the explosion
of the bomb. In the authors' opinion, the residual foci of erythroblasts
described in the Hiroshima material may well be foci of regenerating
erythroblasts.

It may not be amiss to point out here that a variety of toxins can
duplicate in detail the bone marrow changes which follow irradiation.

LYMPHATIC ORGANS

Lymphatic tissue is composed of a framework of reticular cells and
fibers with vast numbers of large, medium-sized, and small lymphocytes
filling the meshes. The relative proportions of lymphocytes to stroma
determine whether the lymphatic tissue in a given location is loose, dense,
or nodular (Maximow and Bloom, 1948).

Lymph nodes and the accumulations of lymphatic tissue in other
organs (tonsils, solitary nodules and Peyer's patches of the intestinal
tract, and appendix) are exceedingly susceptible to ionizing radiations,
and, as in the other hematopoietic organs, the degree of injury is propor-
tional to the dose, within certain ranges. Damage is evidenced by
massive destruction of lymphocytes of all sizes with consequent striking
changes in the lymphatic nodules and reduction in size of the organ.

1102

RADIATION BIOLOGY

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Fig. 17-2. Three stages in the degeneration and beginning regeneration of lymphatic
tissue of mesenteric lymph node of rabbits after total-body irradiation with 800 r of
200-kv X rays. (A), untreated rabbit, shows an active germinal center with the shell
of small lymphocytes extending into the dense diffuse lymphatic tissue. (B) 17 hours
after irradiation, all the lymphocytes in the nodule have been destroyed and many of
those at its periphery are also degenerating. Most of the debris is within macrophages,
especially in the center of the nodule. (C) 24 hours after irradiation; except for a few
scattered lymphocytes, the nodule consists mainly of reticular cells and a few lympho-
cytes, both large and small. (D) 31 days after irradiation; there is a focus of intense
formation of lymphocytes (a new nodule) and beginning repopulation of the diffuse
tissue with lymphocytes. (After De Bruyn.) 220 X.

HISTOLOGICAL CHANGES AFTER IRRADIATION 1103

This destruction of lymphocytes is soon reflected in a drop in the number
of lymphocytes in the peripheral blood (Chap. 16 by Jacobson).

After the administration of 800 r of total-body X radiation to rabbits,
nuclear debris of lymphocytes in the lymph nodes rises from an appreci-
able amount at Yi hour to a maximum at 8 hours (De Bruyn, 1948). It
is usually concentrated in the more active portions of the nodules rather
than at their periphery. Outside the nodules it is diffusely distributed
throughout cortex and medulla, sometimes with a little in the lumen of
small lymphatic vessels. The reticular cells show no evidence of damage.
Prompt phagocytic activity of macrophages results in almost complete
disappearance of debris by 24 hours, leaving large areas of lymphocyte-
poor tissue frequently characterized by dense masses of spindle-shaped
reticular cells. The concentration of these cells is probably due to a
collapse of the stroma rather than to a hyperplasia of the reticular cells,
since none of them are in mitosis. Medium-sized and large lymphocytes
frequently exhibit signs of damage, such as clumping of chromatin, loba-
tion of nuclei, or formation of giant cells, and many, but not all of them,
degenerate. As in the bone marrow, reticular cells and macrophages are
not visibly affected at any stage and are the main potential source of
regeneration of lymphocytes. Heterophil leukocytes appear in great
numbers during the first day and persist for several days, after which they
degenerate and are phagocytosed by macrophages. Edema of the con-
nective tissue about some of the nodes is prominent as late as 10 days
after irradiation. Following the disintegration and removal of the
lymphocytes, most of the lymphatic nodules disappear, leaving a promi-
nent reticular stroma for several weeks.

Early regeneration is diffuse, nodular regeneration occurring much
later. Beginning at 5 days, diffuse areas of medium-sized lymphocytes,
many in mitosis, appear in the cortex and to some extent in the medulla.
Then, three weeks after irradiation, these cells begin to concentrate in
localized areas of the cortex, forming small "bare" germinal centers,
frequently associated with small areas of ectopic myelopoiesis. These
bare nodules gradually grow larger, develop a shell of small lymphocytes,
and by four months resume the appearance of typical nodules. Plasma
cells, noted from 9 days on, especially in the medullary cords, gradually
diminish after 36 days.

In the lymph node, as in the bone marrow of animals exposed to X rays
(100 to 800 r), the damage is correlated with the amount of radiation.
This correlation holds, not only for the quantity of cellular debris, but
also for the alterations in the nodules. In contrast to complete destruc-
tion and delayed regeneration of a majority of the nodules in rabbit
lymph nodes after 800 or 600 r of X rays, after 400 r there is only partial
destruction and a greatly accelerated recovery, and after still lower doses
the changes are correspondingly less marked. After 100 and 50 r there is

1104 RADIATION BIOLOGY

a latent period of 3 hours before damage is evident. At 50 r the only
injury is an increase in debris at 3 and especially at 8 hours which is not
confined to the nodules. Below 50 r no damage is observed.

In different species the lymphatic tissue undergoes the same degree of
damage at a particular dose level, regardless of the lethality of that dose
for the species.

Equivalent doses of fast and slow neutrons and y rays produce the
same histological changes in the lymph node as do X rays.

Repeated doses of 80 r of total-body X irradiation to mice for five weeks
apparently have no cumulative or sensitizing effect on the lymph node and
produce no changes in the amount of lymphatic tissue or in the number
and size of active nodules.

Repeated doses of external y rays to mice, 8.8 r per day, produce
changes in the lymph node only when the irradiation is continued for
10 months or more (Spargo et al., 1951). Lymphocytopoietic activity
decreases, the nodules becoming smaller and fewer until they disappear
completely, and, concomitantly, transport of small lymphocytes through
the cortical sinuses diminishes until there is none at 16 months. Lower
doses, 1.1 or 4.4 r per day, even when continued for 16 months, cause no
significant changes.

The changes in the lymph node after the internal administration of
radioactive isotopes depend on whether the isotopes lodge in or near the
organ. When present in sufficient amounts they produce the same
qualitative changes as those noted after external irradiation. A quanti-
tative comparison of effects of external and internal irradiation is more
difficult, because of the uneven distribution of the isotopes in the various
organs and tissues, the varying rates of excretion from the body, and the
differences in their half-lives.

The intestinal lymphatic tissue undergoes the same histological altera-
tions as the lymph node after external and internal irradiation.

THYMUS

The thymus consists of a framework of reticular cells — mainly of
endodermal but also of mesenchymal origin — whose meshes are filled
with dense masses of small lymphocytes in the outer (cortical) portion of
the organ and with smaller numbers of small lymphocytes and a few
medium-sized ones in the inner (medullary) portion of the organ. The
thymus gradually "involutes" with advancing age, losing lymphocytes
and some stromal cells.

Studies of the thymus after X irradiation reveal a loss in size of the
organ, owing mainly to destruction of lymphocytes. In the rabbit, the
effects of a single dose of 800 r total-body X ray (Murray, 1948b) include:
(1) a destructive phase (1 to 8 hours), during which most of the lympho-

HISTOLOGICAL CHANGES AFTER IRRADIATION 1105

cytes die; (2) a phase of cleanup (8 hours to 2 days), when debris is
removed by macrophages and possibly in part through lymphatic
channels, exposing a condensed epithelial stroma and blood vessels in
the cortex and leaving a few surviving lymphocytes in the medulla;
(3) a phase of inactivity (2 to 9 days), during which shrinkage continues
and the connective tissue becomes prominent ; and (4) a phase of regenera-
tion (10 days to four weeks), characterized by a repopulation with
lymphocytes, proceeding outward from the medulla. The stromal or
reticular cells are not damaged by this or even much larger amounts of
irradiation. The H assail' s bodies, formed of masses of epithelial cells,
are unaffected. There are suggestions of new formation of Hassall's
bodies from stromal cells at 14 days.

At lower doses fewer cells die, the inactive period is shorter, and regen-
eration is more rapid. This sequence of changes is somewhat similar to
that already described for the lymphatic tissue. As in that tissue, the
reaction in different species is the same to equal single doses of X rays,
irrespective of the variations in LD50 levels for these species.

External (3 rays sufficient to kill about 20 per cent of the mice (5000 rep)
do not affect the thymus in these animals. Qualitatively, the reactions
to fast and slow neutrons are the same as to X rays. Quantitatively, the
X-ray /fast-neutron ratio has been estimated to be roughly 5 and the
X-ray/slow-neutron ratio to be roughly 1 (Murray, 1948b).

Long-continued, daily low doses of total-body y rays gradually cause
depletion of lymphocytes — with 8.8 r first at 6 months and with 4.4 r at
8 months (Spargo et al., 1951). With 1.1 r greater depletion than in
controls is first observed at 10 months. Both in this group and in the
4.4-r group attempts at regeneration are present at 14 and 16 months.
In all these animals there is a striking increase in mast cells in the depleted
organs as well as in the surrounding connective tissue. Plasma cells are
not prominent.

The depletion of the thymus following internal irradiation is similar to
that produced by X rays, but it usually occurs more gradually and is
more prolonged. Also, because of localization of the isotopes, the deple-
tion is not always uniform. Irregular depletion results from the uneven
distribution of a emitters within the thymus. For example, after admin-
istration of plutonium, areas containing the isotope, as seen in autoradio-
graphs of adjacent sections, are depleted, whereas nearby uncontaminated
areas are not. When the thymus is irradiated as a result of activity
deposited elsewhere, as Srs9 or Y91 in the sternum, or Y91 or cerium-
praseodymium (275-day) in the lung, there is a gradient of damage.
The parts nearest the localized isotopes suffer the most damage, which
decreases perceptibly away from the focus. However, as might be
expected, the highly diffusible Na24 in doses of 50 to 80 juc/g in mice
(considerably above the LD50 30-day level) causes a rapid, diffuse, and

1106 RADIATION BIOLOGY

severe depletion of the organ in 4 days. In contrast to large single doses
of external irradiation in which there is marked regeneration, there is none
in those areas of the thymus in the range of long-continued bombardment
from isotopes in nearby tissues.

SPLEEN

The spleen is made up of lymphatic tissue which is typical in its
white pulp and atypical in its red pulp. The white pulp ensheaths the
arteries; the red pulp surrounds the radicles of the veins and consists
mainly of reticular cells and fibers and a few lymphocytes ; it is permeated
by circulating blood. Since lymphopoiesis occurs in the while pulp and,
in some mammals, ectopic myelopoiesis in the red pulp, the effects of
ionizing radiations on these functions may be compared here within a
single organ. The white pulp, essentially dense lymphatic tissue with
nodules, reacts in a fashion similar to that of lymph nodes.

After a single exposure of rabbits to 800 r of total-body X irradiation,
the spleen is rapidly reduced in size during the first day, and for a week or
longer it remains small and dense, with relatively small and inconspicuous
areas of white pulp, lacking nodules or germinal centers (Murray, 1948a).
Neither the germinal centers nor the white pulp as a whole recovers
appreciably until about two weeks after treatment. Mitosis stops in a
few minutes, and within the next few hours most of the lymphocytes
throughout the white pulp die and the germinal centers, normally rich in
medium-sized lymphocytes, are obliterated as such, leaving dense masses
of reticular cells. In the red pulp, lymphocytes suffer the same change
as in the white pulp, and the relatively few erythroblasts and myelocytes
present show damage at early intervals and are absent at 5 and 9 days.
The period from 3 to 17 hours is one of rapid phagocytosis by reticular
cells of the debris from this widespread degeneration in both white and
red pulp and from invading heterophil leukocytes. With the removal of
the dead cells, the spleen, particularly the white pulp, shrinks, leaving
the reticular cells prominent. A phase of relative inactivity ensues (1 to
9 days) with further condensation of the reticular cells, the few abortive
attempts at regeneration being by transformation of reticular cells.
Finally (10 days to four weeks) a phase of reconstitution takes place
through mitotic activity of the lymphocytes. The regeneration of
erythroblasts and myelocytes and new formation of megakaryocytes
occur after the resumption of lymphopoiesis.

On the other hand, in mice and rats receiving 350 and 600 r (total-
body), respectively, erythropoiesis is renewed much earlier. It expands
from a few foci in the red pulp to such an extent that it sometimes
encroaches upon the white pulp and apparently interferes with regenera-
tion there. Megakaryocytes, which are numerous in the normal spleen

HISTOLOGICAL CHANGES AFTER IRRADIATION 1107

of these animals, are greatly decreased by 3 days but begin to reappear
in mice at 5 days after 350 r and in rats after three weeks following 600 r.

The sequence and type of effects are similar with lower doses, but fewer
cells die, the inactive period is shorter, and regeneration is more rapid.
The normal structural pattern shows little disruption below 175 r, and
damage to lymphocytes is not seen below 25 r. In different species,
damage to the spleen is of the same magnitude for the same size dose,
irrespective of differences in the LD50 for these species (Murray, 1948a).

Fast and slow neutrons resemble X rays in their qualitative effects on
the spleen. Equivalent fractions of the LD5o 30-day level produce similar
effects. Estimations of their biological effectiveness on the spleen are,
roughly, X/fast neutrons = 4; X/slow neutrons = 0.85.

Daily doses of 80 r of total-body X rays severely deplete the mouse
spleen by 20 treatments, but erythropoiesis is definitely elevated after 24
treatments. Depletion is less than after 350 r (total-body) given in a
single dose.

Seven hundred roentgens of y rays (from an external source of radium)
administered over a three-month period at the rate of 8.8 r per 8-hour day
causes less damage to the spleen of guinea pigs than a single total-body
dose of 175 r of X rays (Lorenz, Heston, Jacobson, et at., 1953). A similar
experiment with mice shows a gradual shrinkage of the organ with pro-
gressive depletion of white pulp (Spargo et al., 1951). In the red pulp
an increase in erythropoiesis begins at 6 months after 8.8 r per day and
becomes progressively greater up to 16 months, with hemocytoblasts
predominating at the latter interval. After 4.4 r per day erythropoiesis
increases only at 14 and 16 months. In all these chronically irradiated
mice there is a progressive increase in mast cells, reaching a maximum in
the 8.8-r group of 4.5 times the number in controls.

Autoradiographs show all isotopes more concentrated in the red pulp
than in the white, the deposition often being especially heavy at the
transition between the two zones. With Zr93-Cb93, Y91, and radium there
is some deposition in the white pulp, mainly around the arterioles.

When radioactive isotopes lodge, even temporarily, in the spleen, the
effects, with one important exception, are similar to those produced by
external irradiation and consist in depletion of white pulp and shrinkage
of the organ. Indeed, the changes due to administration of appropriate
amounts of Na24 cannot be told from those due to X rays. However, the
injection of bone-seeking isotopes such as Y91, Sr89, Ba140-La140, plutonium,
or radium into small mammals causes an extreme depletion of hemato-
poietic cells in the bone marrow, and the red pulp of the spleen develops
an accelerated hematopoiesis. Zr93-Cb93, however, in addition to produc-
ing a marked depletion of splenic white pulp, differs from the other bone-
seeking (3 emitters in not eliciting a marked compensatory hematopoiesis
in red pulp. In fact, erythrophagocytosis is striking in the red pulp

1108 RADIATION BIOLOGY

after injection of this isotope. It is noteworthy that erythropoiesis does
occur (except after Zr93-Cb93) in the red pulp, which is the site of lodgment
of many isotopes. With practically all radioactive isotopes, the mega-
karyocytes, after a preliminary decrease, increase tremendously in num-
ber after weeks and months and often show great nuclear abnormalities
(Murray, 1948a).

BONE

Bone is a specialized connective tissue consisting of bone cells (osteo-
cytes) embedded in a matrix of collagenous fibers and amorphous cement-
ing substance which is heavily infiltrated with small crystals of hydroxy-
apatite, calcium phosphate, carbonate, and small amounts of citrate.
On the surface of a developing spicule of bone are cells called osteo-
blasts, which are believed to make the bone, and some of them become
bone cells. In areas where bone is being destroyed, there are large multi-
nucleated cells called osteoclasts. Their presumed role is the destruc-
tion of bone.

In normal animals growth of the bone in length depends on (1) con-
tinued multiplication of the cartilage cells in the epiphyseal plate and,
near the bone marrow, calcification of the intercellular substance between
them; and (2) continued invasion of the calcified cartilage matrix by
vascular connective tissue from the bone marrow and deposition of bone
in the space thus made available, often on remnants of cartilage which
are not destroyed. With cessation of bone growth, multiplication of the
cartilage cells and invasion by blood vessels stop, and bone is deposited
on the diaphyseal side of the cartilage plate. This plate disappears in
man on reaching adulthood, and continuity develops between the marrow
of metaphysis and epiphysis. The epiphyseal cartilage participates in
the growth of bones for a long time in rats, but in mice the period of
growth lasts for only two or three months.

Bones grow in width by deposition of bone directly on the outer surface
of the bony cylinder (periosteal growth). In addition, with the recon-
struction of a bone which takes place as it grows, new bone may be laid
down inside the bone collar (endosteal growth). This is especially true
toward the growing end of a bone.

Although many believe mature bone is relatively resistant, there is
little doubt that it can be severely damaged by irradiation. Thus a
peculiar eburnation and devitalization of bone (Friedman, 1942) and
massive necrosis after dosages which did not damage the skin (Regaud,
1922) have been described.

There is general agreement that the tissues responsible for growth of
bone can be severely damaged by irradiation. Stunting of the growth of
the long bones has been observed by many investigators. The rat,
because of its rapid and prolonged period of growth, is a sensitive indi-
cator of these radiation effects. After 600 r of X rays (LD50 30-day

HISTOLOGICAL CHANGES AFTER IRRADIATION

1109

Fig. 17-3. (A) Metaphysis of tibia of untreated rat showing the interdigitation of
primary spongiosa with cartilage; 35 X. (B) Metaphysis of rat 9 days after 600 r
total-body X rays (200 kv). Note the complete severance of the bone of the meta-
physis from the cartilage; 40 X. ((') Metaphysis of mouse femur 90 days after 3.6
Mc/g of Sr89. There has been severance of the spongiosa from the epiphyseal plate and
the cartilage has become markedly thickened, with swelling of the cells; 50 X. (D)
Metaphysis of mouse femur 40 days after 1 A<c/g of radium. Note areas of newly
formed, atypical bone in the marrow; 105 X. (After Heller, 1948.)

1110 RADIATION BIOLOGY

level), the continuity of the epiphyseal cartilage of femur and tibia with
the spongiosa is frequently interrupted, resulting in a temporary cessation
of bone growth in length (Heller, 1948a). Preceded by excessive hyper-
trophy of cartilage cells in the zone of ossification at 3 days, the normal
interdigitation of cartilage and metaphyseal spongy bone is gradually
lost. By 9 days the two are completely separated. In the lamellae of
the spongiosa there are some dead bone cells, but none in the dense
cortical bone. Osteoblasts disappear after irradiation but gradually
reappear after some weeks. Only a slight increase in the number of
osteoclasts accompanies the resorption of bone. In some instances this
severance of cartilage and spongy bone may persist for a month, but in
others a new primary spongiosa is laid down against the cartilage plate
as early as 21 days or possibly sooner and growth in length is resumed.
The old, disconnected spongiosa is gradually resorbed. Recovery of the
growth process is complete at 70 days, although the bones are perma-
nently somewhat stunted.

A single dose of 400 r of X rays merely reduces the number of osteo-
blasts at 3 and 7 days while osteoclasts are more numerous in parts of the
metaphysis.

In young chicks (three weeks of age) similar changes occur. Doses of
800 or 1000 r total-body X irradiation cause severance of cartilage from
spongy bone, effectively stopping growth of the bone. After 1000 r there
is a steady progression of damage from 1 to 16 hours, with the disappear-
ance of osteoblasts, increase of osteoclasts around the spongy bone, and a
decrease in the erosion of cartilage. From 7 hours to 4 days occasional
dead osteocytes and a number of empty lacunae are present in the spongy
bone. By 4 days separation of cartilage from spongy bone is extensive,
some of the latter having been resorbed, and the bone stops growing.
After 800 r, the destructive process is the same as after 1000 r, only not
quite so rapid. However, by 5 days severance is extensive, osteoblasts
are absent, and bone growth stops. After 400 r there is a progressive
reduction of osteoblasts around the metaphyseal spicules of bone from 2
to 18 hours, but at this dosage no severance or interruption of bone growth
occurs. Recovery is rapid, with osteoblastic and osteoclastic activity
resumed by 30 hours. No effect on chick bones is noted after 100 r.

In comparison with the effects of external irradiation on bone at LD50
levels, even small fractions of the LD50 of the bone-seeking isotopes cause
disproportionately greater damage, owing to their continued activity at
the points of deposition (Heller, 1948a). Among these isotopes there is a
remarkable similarity of reaction, especially between radium and plu-
tonium, and the /3 emitters Sr89 and Y91. However, the weak (3 particles
of C14, which deposit heavily in bone after injection of NaHC1403, do
not interfere with the growth of bone, even when 2 juc/g is injected (W.
Bloom, 1949).

HISTOLOGICAL CHANGES AFTER IRRADIATION

1111

Autoradiography indicate that deposition of most radioactive materials
is greatest in the areas of maximum growth in length of the bones. The
isotopes are also deposited along the periosteum and parts of the endos-
teum. Where there is intense radioactivity, osteocytes degenerate.
The bone of the metaphysis is devitalized to a greater extent than that
of the cortex. But in rats five and six months after radium dosage of
0.06 juc/g, even the dense cortical
bone contains large numbers of
lacunae devoid of their bone cells.
As after X irradiation, osteoblasts
disappear and interdigitation of bone
and cartilage at the epiphyseal line is
decreased, often leading to complete
separation of epiphyseal cartilage
from spongiosa and arrest of growth
in length. After low doses of short-
lived radioactive isotopes, as after X-
ray dosage, repair occurs and growth
in length is resumed. Longer-lived
agents and higher doses cause an
overgrowth of atypical metaphyseal
bone which later becomes devital-
ized. This bone varies in appearance
by the number of osteocytes, coarse-
ness of connective tissue fibers, and
density of the matrix ; it never shows
evidence of intracartilaginous ossifica-
tion. It may persist in the center of
the shaft months after injection of the
radioactive material. The old meta-
physis with its devitalized bone is
very weak and fractures often occur
here. This is also the site in which
osteogenic sarcomas develop. Several
examples of this occurred in radium
dial painters (Martland, 1931). In
rats receiving doses that permit sur-
vival for a long enough time, this
atypical overgrowth is outstripped by

the normal growth mechanism and growth in length of the bone is
resumed. At still lower doses there are sometimes suggestions of partial
resorption of the old spongiosa and atypical bone even with the long-
lived a emitters.

However, where bone growth has ceased, as in adult mice, the picture

Fig. 17-4. Section of proximal epiphy-
sis of rat tibia, five months after the
injection of 0.125 juc/g of radium
showing the old metaphysis at (a),
new bone at (6), and the new meta-
physis at (c). (After Heller, 1948.)
9X.

1112 RADIATION BIOLOGY

is quite different. For example, five and seven months after the injec-
tion of plutonium or radium (0.03 juc/g), atypical bone often replaces the
marrow in the metaphysis and even much of the shaft (M. A. Bloom and
W. Bloom, 1949).

In a growing bone, isotope-containing bone will be in part resorbed with
the remodeling and in part incorporated in new bone. The isotopes
enter adult bone by incompletely understood processes of exchange with
atoms in the bone-salt crystal complexes. Moreover, in the case of
yttrium and plutonium, at least some of these elements become firmly
adherent to the organic part of the intercellular substance of bone (Copp,
Axelrod, and Hamilton, 1946; Copp, Greenberg, and Hamilton, 1946).

CARTILAGE

This tissue is distributed in the respiratory system, joints, and grow-
ing bones. In the last, a definite portion of the epiphyseal cartilage is in
continuous growth, and a thin zone within it is just as continuously
undergoing provisional calcification and shortly thereafter dissolution and
replacement by newly formed bone. As might be expected, cartilage in
these several positions and different states of activity shows great varia-
tions in susceptibility to damage by irradiation. In growing animals it is
Difficult to assess accurately just how much of the change resulting from
irradiation is due to failure of replacement of cartilage by ingrowth of
bone. In nongrowing animals, on the other hand, it is probable that all
the changes in cartilage are direct effects of irradiation. The response of
cartilage may vary from rather marked resistance in adult animals to a
fairly high degree of sensitivity during the period of rapid growth with
resulting retardation of skeletal growth. On the basis of investigations
on chicks (Perthes, 1903), on young rats (Segale, 1920), and on young
rabbits (Bisgard and Hunt, 1936; Brooks and Hillstrom, 1933), it appears
that localized exposure of an epiphysis of tibia or femur to 1500 r or less
of X rays leads to an arrest of elongation of the bone owing to damage to
the epiphyseal cartilage (Hinkel, 1942). Degenerative changes in certain
cartilage cells, particularly in the zone of proliferation, occur as early as
2 days after irradiation, and at 6 days many of these cells are hypertrophic
and vacuolated, with pyknotic nuclei. The columns of cells and the
intercellular bridges lose their regular arrangement, the epiphysis becomes
thinner, and, after two weeks, the disturbance in the growth mechanism is
manifest in both cartilage and new bone of the metaphysis. By six
months the epiphyseal cartilage has disappeared ; this is much earlier than
it disappeared on the nonirradiated side. There are indications that
growing children are much more sensitive to radiation than these animals
(Gates, 1943). With total-body as well as with localized irradiation,
abnormally swollen cartilage cells have been noted as the first sign of

HISTOLOGICAL CHANGES AFTER IRRADIATION 1113

degeneration in the zone of provisional calcification (Heller, 1948a).
This is observed in rats 3 days after a single dose of 600 r of X rays (total-
body) and is followed by partial or complete separation of epiphyseal
cartilage from the primary spongiosa and eventual recovery with resump-
tion of growth (see section on Bone).

The greater and more sustained damage to cartilage produced by
certain internally administered radioactive isotopes parallels their greater
injurious effects on bone, and varies with the dosage and with the age of
the animal and the consequent stage of growth of its bones. Swelling
and death of cells of the epiphyseal cartilage are noted early after deposi-
tion of most bone-seeking isotopes. The effects are especially severe in
young rats, in which separation of the cartilage from the bone begins at 3
days and is nearly complete 7 days after a Sr89 dose of 2.9 /xc/g (Heller,
1948a). By 11 days the entire epiphyseal cartilage is several times the
normal width and many cartilage cells are dead, especially in the zone
bordering the metaphysis. The articular cartilage cells, particularly
those facing the epiphyseal cavity, are either dead or greatly enlarged.
In some instances the articular cartilage becomes separated into several
portions by connective tissue. By three or five months (depending on
the dose and the particular isotope) that portion of the cartilage not
destroyed by radiation from the deposited isotope has continued its
growth and moved away from the zone of damage; growth in length of
the bones continues in histologically normal although slower fashion, and
the animals are permanently stunted.

In rats injected with radium, 0.5 /xc/g, the width of the cartilage plate,
at least twice the normal at two months, increases still more at three and
five months. At this last interval the cartilage has been invaded by
vascular connective tissue, which extends transversely from the region
of the periosteum and separates large portions of the hypertrophic
cartilage, often 30 cell layers deep, from the rest.

In mice, in which growth in length of the bone ceases early, Sr89 also
causes an abnormal hypertrophy and vacuolization of cartilage cells at
most intervals, but conspicuous from 90 to 197 days after 3.6 juc/g and as
late as 70 days after 0.86 /xc/g. The cartilage plate becomes thicker with
marked irregularities owing to the increased number of hypertrophic
cells. After higher dosages, from 5 to 15 juc/g, the same changes occur,
but earlier. The effect of radium is similar but even more severe, despite
its doubtful deposition in the epiphyseal cartilage at early intervals and
its absence at 40 days. From 8 to 40 days after a radium dose of 1 /xc/g,
swollen and degenerating cartilage cells of the epiphyseal cartilage
encroach even into the deeper layers of the plate. Eighty days after a
dose of 0.5 iic/g, multiplication of the cartilage cells has increased the
width of the plate two or three times and dead cartilage cells are numer-
ous. The irregularity of this invasion of connective tissue results in

1114 RADIATION BIOLOGY

tremendous variations in width of the epiphyseal plate and great
abnormality in the appearance of the whole metaphysis.

MALE GENERATIVE SYSTEM

Total-body irradiation, whether it is carried out with single or repeated
doses of X rays or 7 rays or single exposure to fast neutrons, produces the
same pattern of damage and repair in the testis. The degenerative
changes are mainly induced through injury to the free stem cells. With
all types and amounts of external and internal radiations that produce
cellular depletion, the testis is reduced in size and there is a gradual
cessation of spermatogenesis (Albers-Schonberg, 1903; Barratt and
Arnold, 1911-1912; Bergonie and Tribondeau, 1905; Heller, 1948b;
Regaud and Blanc, 1906; Schinz and Slotopolsky, 1925).

A single total-body exposure of rabbits to 800 r of X rays destroys the
spermatogonia. This causes an interruption in the production of new
spermatocytes and consequently of spermia several weeks later. Cell
death, evidenced by dead spermatogonia (and only rarely by a dead
spermatocyte) 1 day after treatment, is less obvious at 3 and 5 days
(Heller, 1948b). By 9 days spermatogonia are inconspicuous in most
tubules, which contain only Sertoli cells, a decreased number of spermato-
cytes (some damaged), spermatids and spermia, and great numbers of
multinucleated and bizarre cells. Beginning regeneration (at 30 to 35
days) is indicated by the presence on the basement membrane of baso-
philic outstretched cells or very early spermatogonia (Heller, 1948b).
Some of the tubules even contain a few spermatocytes at this time.
Spermatogenesis returns to normal by four months in some specimens,
while in others the tubules contain Sertoli cells alone. There is some-
times edema of the intertubular connective tissue.

After 600 r the cessation of spermatogenesis followed by regeneration is
similar to that after the higher dose except that not quite so many tubules
are depleted and regeneration begins earlier. Attempts at recovery are
indicated at 21 days by the presence in a few tubules of spermatogonia in
all stages of development, more tubules showing this appearance by 31 days
and at least half the tubules showing some degree of recovery by 45 days.

In the rat the effects of 600 r of X rays on the testis are the same as in
the rabbit, even in time sequence. However, in different rats 132 days
after exposure, there are great differences in the degree of recovery.

The damage to the mouse testis after 350 r of X rays is much like that
in the rabbit after 600 r except that regeneration begins earlier.

The cumulative effect of repeated small doses of radiation is more
striking on the testis than on any other organ of the same animal, although
not quite so marked as on the ovary. The testes of mice irradiated daily
with 80 r of X rays show progressive damage. Twenty-four hours after

HISTOLOGICAL CHANGES AFTER IRRADIATION • 1115

Fig. 17-5. (A)-(D) Sections of rat testes after whole-body irradiation with 600 r of
200-kv X rays. {A) 21 days; the tubule contains Sertoli cells and groups of spermia;
all spermatogenic cells are absent; 350 X. (B) 31 days; the tubule is further collapsed
and contains only Sertoli cells; 350 X . (C) 70 days, beginning regeneration of sperma-
togenesis. Note the almost continuous layer of dark-staining spermatogonia at the
periphery of the tubule, the larger spermatocytes, and the clumps of smaller pale-
staining spermatids; 350 X. (D) 8 hours after irradiation; showing the intact
spermatocytes and three degenerating spermatogonia indicated by the arrows; G75X.
(E) Section of rat testis 24 days after rat was given 5Mc, g of Zr93-Cb93. The tubule
is filled with necrotic cells which stain darkly; 260 X. (After Heller, 1948.)

1116 » RADIATION BIOLOGY

four doses, dead spermatogonia are seen; 24 hours after nine doses,
spermatogonia are rare and there is a marked decrease in spermatocytes ;
after 14, 20, 24, and 35 daily doses of 80 r, there is a gradual elimination of
spermatogenesis until only the Sertoli cells remain.

In the testis of guinea pigs exposed daily to 8.8 r of y rays from a
radium source, there is early destruction of some spermatogonia while
others develop into spermatocytes and eventually into spermia. With
continued irradiation, no regeneration of spermatogonia occurs and the
damage is progressive. After 37 daily treatments (326 r) spermatogenesis
is absent (Heller, 1948b). After 46 exposures (405 r) and 81 exposures
(713 r), only Sertoli cells remain and nearly all cellular debris has been
removed from the lumens of the tubules. From 37 days on, large
vesicular cells resembling fibroblasts are prominent around the tubules
and among the interstitial cells. However, after 81 exposures the
epididymis still contains spermia.

Histologically, the findings are the same in mice similarly treated, but
complete cessation of spermatogenesis is less rapid. Although there is
considerable generalized degeneration after two months of irradiation
(8.8 r per 8-hour day), practically complete depletion of the tubules
(except for Sertoli cells) is not observed until the eight-month interval
(Eschenbrenner, 1946; Spargo et at., 1951). In mice irradiated with 4.4 r
per day, the testis shows only slight degenerative changes up to 10
months. Twelve to 16 months of treatment cause slight but progres-
sively increasing degeneration. Irradiation at the rate of 1.1 r per day
produces no visible changes in the testis up to 16 months.

Exposure of rabbits and mice to fast neutrons in comparable fractions
of the LD50 30-day level results in similar but even more severe damage
to the testis than X irradiation (Heller, 1948b; Lawrence and Tennant,
1937). In addition to these degenerative and regenerative changes in
the tubules, the interstitial connective tissue in the mouse contains focal-
ized masses of plasma cells and some areas of hematopoiesis. There is
also some inflammation in the epididymis. The testes of young rabbits
show the peak of damage at 21 days, when every tubule contains a dense
amorphous material and an occasional huge basophil cell. In the sur-
rounding connective tissue there is frequently considerable extravascular
and intravascular erythropoiesis.

Beta rays from an external P32 source produce extensive damage at
the periphery of the testis near the source of the radiation. The damage
decreases toward the interior of the organ. In the least damaged por-
tions regeneration is rapid, but the outside margin is incompletely
recovered 28 days after exposure to 2500 rep. Besides the dead cells in
the severely injured marginal tubules, giant and bizarre cells are often
prominent (Heller, 1948b).

Although the effects of internally administered radioactive isotopes are

HISTOLOGICAL CHANGES AFTER IRRADIATION 1117

the same in kind as of the externally applied radiations, the extremes of
damage are much greater at the LD50 30-day level for the internally
administered substances; corresponding with their localized deposition,
the resulting damage tends to be localized. Radium, plutonium, and
Zr93-Cb93 appear to be more destructive to the germinal epithelium than
Sr89, P32, Ba140-La140, and Y91. Following very low doses, injury and
repair may occur concomitantly, making it difficult or impossible to
evaluate the damage.

After all types of irradiation, external and internal, the stem cells are
the most sensitive and the interstitial cells are extremely radioresistant.
Except in a few animals after neutron and radium treatment, the Sertoli
cells show no significant changes.

There are but scanty reports on the male accessory organs and passages.
They all seem to be relatively radioresistant.

FEMALE GENERATIVE SYSTEM

Irradiation of the mammalian ovary causes marked atrophy of the
organ and may produce temporary or even permanent sterility if the
dosage is high enough (Halberstaedter, 1905; Lacassagne, 1913; Lacas-
sagne and Gricouroff, 1941 ; Shields Warren, 1942, 1943). If sterilization
is not permanent, there may be a reawakening of sexual activity after the
passage of months and even normal pregnancies may ensue. Dependent
on and following the changes in the ovary there is atrophy of tubes,
uterus, vagina, and mammary gland.

The sensitivity of the ova and of the attached follicular cells varies
with their functional states at the time of irradiation (Lacassagne, 1913).
There are unusually marked species differences in sensitivity. The
corpora lutea and interstitial cells are relatively very radioresistant.

Irradiation of the ovaries of a rabbit with 1200 r produces dramatic
effects (Lacassagne and Gricouroff, 1941). In primitive ova (surrounded
by a single layer of epithelium) the signs of degeneration are pyknosis or
chromatolysis and fragmentation of the cell. Debris of ovocytes is
phagocytosed by the follicular cells, but these also may degenerate and
the whole follicle disappear in 3 or 4 days. However, some primitive ova
escape destruction and after five months or more they may begin to
develop. When irradiated in the beginning growth phase, first the ovum
and then follicular epithelium degenerate within 2 weeks. In irradiated
large, growing follicles the first changes are seen in the follicular epi-
thelium, but the residue of large ova may persist for months. In mature
follicles the follicular epithelium disappears early after irradiation, while
the ovum, with more or less visible damage, may persist for some time.
There may be hemorrhages in the follicles. Apparently, definitive
destruction of all ova may take place with 2000 to 2500 r.

1118 RADIATION BIOLOGY

On the other hand, interstitial gland cells show little if any effects of
such an exposure, although these glands gradually decrease over some
months and imperceptibly disappear, owing to failure of new cells to
develop from corpora lutea. Interstitial cells give rise to cords and
"glands." Some also may develop as ingrowths from surface epi-
thelium. It should be noted that the interstitial glands, although
prominent in the ovaries of rodents, are inconspicuous and may even be
absent in the human ovary.

The corpora lutea are not affected by doses which destroy ova and
growing follicles. Indeed, irradiation a few hours after copulation does
not hinder development of corpora lutea, although such corpora lutea
degenerate early. It has been reported that 500 to 1300 r administered
before ovulation has no effect on development of corpora lutea, while
after larger doses (1300 to 1800 r) small corpora lutea develop but their
secretion does not maintain an endometrial reaction for more than 6 days
(Lacassagne and Gricouroff, 1941). In any event, an amount of irradia-
tion which destroys all developing follicles will cause failure of corpora
lutea to develop.

Although 600 r produces only minimal changes in the rat, most ova are
destroyed at 1600 to 1800 r, while sterilization requires 3000 r (Lacassagne
and Gricouroff, 1941). However, total-body irradiation of rats with
600 r produces great degenerative changes in the ovary, there being
practically no intact ova at any stage later than 2 days, although a rare
corpus luteum but no ova may be found after four months (W. Bloom,
1948d). The primitive ova of the mouse are exceptionally sensitive to
irradiation (Murray, 1931 ; Schugt, 1928). A dose of 54 r destroys many
follicles, and doses of 140-150 r produce histological sterilization.

Daily irradiation with 8.8 r of y rays per 8-hour day produces apparent
sterilization of the mouse ovary at four months, with 4.4 r per day at
eight months, while with 1 . 1 r per day there are still some growing follicles
at sixteen months (Eschenbrenner, 1946; Spargo et al., 1951).

The ovary is sensitive to only some of the radioactive isotopes, pre-
sumably because of the failure of many others to localize in the organ.
Radium and plutonium in small doses produce marked degeneration in
the ovaries of both mice and rats (W. Bloom, 1948d).

The columnar epithelium of the oviduct and uterus is much less sensi-
tive than the stratified squamous epithelium of the vagina. In uteri of
rabbits exposed to 2000 r shortly after birth, all coats of the uterus show
some degeneration in a few hours and the organ atrophies (Gricouroff,
1930). But starting with the fifth day, mitoses reappear and the organ
continues to grow like that of controls, although regression of the ovaries
persists.

It is noteworthy that the male keeps its secondary sex characters after
irradiation (Ancel and Bouin, 1907) whereas the whole female sex appa-

HISTOLOGICAL CHANGES AFTER IRRADIATION

1119

ratus atrophies. This occurs in practically all mammals except the
mouse_in which sterilization of all follicles in very young or adult mice
does not prevent later development of estrual cycles (Parkes, 1927).

SKIN

As might be expected from the different tissues that enter its structure,
the changes in the skin following irradiation vary considerably with the

m

%

lis *&*-■■■

v. *

tf iii r

% # W *

m$ ^

B

Fig. 17-6. (A) Section through ear of mouse 24 hours after 5000 rep of external /3 rays.
The epidermis is of normal thickness, the derma is slightly edematous; 325 X. (B)
Lumbar skin 56 days after 12,000 rep of external /3 rays. The epidermis is greatly
thickened and the derma still shows some infiltration with inflammatory cells. This
thickening of the epithelium is typical of regeneration after severe injury to skin.
Hair follicles are absent, although there is a portion of the smooth muscle of a hair in
the lower left-hand part of the section; 210 X. (After Snider, 1948.)

histological constitution of the several parts of the organ. One group of
changes occurs in the epidermis with its stratified squamous epithelium
and its derivatives, the hair and associated sebaceous glands, the sweat
glands, and the nails. In the other category of tissues are the vascular
connective tissues forming the derma and the subdermal tissues. The
smooth muscles of the hair may be included with this group. The
analysis of the changes in skin has been complicated by the long latent
period in the epidermis and by the fact that most of the human epidermis
is more sensitive and thicker than that of the laboratory mammals. This
probably explains its exceptional reaction to various chemical and

1120

RADIATION BIOLOGY

physical injuries; it forms blisters readily, in contrast to the skins of prac-
tically all other animals, which do not.

The gross changes in the skin extend from erythema and epilation
(and, in some cases, pigmentation in human skin), which result from
exposure to 500 to 800 r, to more marked changes with higher dosages,

which result in thin, fragile,
regenerating epidermis, sclerosis
of the derma, and necrosis and
ulceration. The extent of the
injury to the skin varies with the
amount of radiation absorbed
(the effect is also greater per
roentgen of soft X rays) , with the
animal species, with different
individuals of the same species,
and even with different areas on
the body of the same individual.
This last difference is important
when using erythema as a criterion
of dosage.

Epithelium. The first change
to be seen after irradiation with
doses up to several hundred
roentgens is a temporary cessa-
tion of mitosis in the lower layers
of the epithelium. It has even
been claimed that with 35 r the
mitoses in the epidermis of the
mouse ear are markedly decreased
(Knowlton and Hempelmann,
1949). Two to 6 or 8 days after
300 r in human skin, there are
clumped chromosomes and some
atypical mitoses and the debris of
mitoses. No more dividing cells
are found until about the twen-
tieth day. Usually there are only

Fig. 17-7. Section of base of crypt from a
duodenum of art exposed 28 hours pre-
viously to 600 r of 200-kv X rays, total-body
irradiation. There is still much debris
among the cells in the crypt epithelium and
several mitoses indicative of beginning
regeneration. There are also enormously
swollen cells considerably larger than the
normal cells. 1270X. (After Pierce, 1948.)
Drawn by Esther Bohlman.

a few binucleate cells. From 20
to 40 days there is a variable amount of nuclear polymorphism, with
large and small cells and occasional binucleate cells (Miescher, 1938).
The mitoses now are of normal appearance and the epidermis is not
thickened.

Exposure of human skin to 600 to 1200 r produces some degeneration of
cells of the germinative layer. With these larger doses the number of

HISTOLOGICAL CHANGES AFTER IRRADIATION 1121

mitoses becomes very small after 5 or 6 days and many of the dividing
cells are degenerate. During the second and third weeks many of the
cells seem shrunken. In the succeeding weeks the polymorphism of the
cells becomes greater, and many of the cells appear swollen and poorly
stained. The epidermis shrinks to two or three layers of cells beneath the
cornified layer. If the epidermis is not completely destroyed, a period of
mitotic activity may set in (fifth to eighth week). From this time on
the epidermis is composed of normal appearing cells. With still larger
doses this layer degenerates, and consequently after several weeks the
entire epidermis in the irradiated area has been destroyed. This epi-
dermal degeneration has been claimed to be the result of destruction of
dividing cells. More properly, it is the expression of injury to resting
cells which becomes apparent only when the cells divide some time later.
After complete degeneration of the epidermis, regeneration is said to
occur mainly, if not exclusively, by ingrowth of epithelium from the sur-
rounding uninjured epidermis. In laboratory animals exposed to intense
or prolonged doses of external (3 rays, regeneration is helped also by
proliferation of those hair follicles which are not destroyed by these
shallow penetrating rays. The regenerating epidermis is often thin and
very susceptible to mechanical and other injuries. This friable epi-
dermis may persist for some years. After long-continued irradiation and
a long latent period, it has been the source of malignant tumors, particu-
larly following the softer X rays used so extensively in the early days of
radiology.

Derma. A dose of 600 to 800 r total-body irradiation in laboratory
animals calls forth an exceedingly mild inflammatory reaction character-
ized by slight hyperemia, a very low-grade migration of heterophil
leukocytes and lymphocytes, some destruction of mast cells, and a barely
perceptible edema by 24 hours (Lacassagne and Gricouroff , 1941 ; Schinz
and Slotopolsky, 1925; Snider, 1948a).

In man an erythema due to dilatation of the blood vessels is elicited
with 600 to 700 r of hard X rays over relatively small areas. The
erythema appears in the first day or so, lasts for a few days, and dis-
appears, to reappear on or about the eleventh day. This second ery-
thema lasts three or four weeks and gradually merges into a stage of
pigmentation of the skin (Ellinger, 1941).

With larger doses the inflammatory changes are more marked and
persist longer. After repeated exposures, giant fibroblasts may be found
many months later (Maximow, 1923). Perhaps the most important
effects are on the blood vessels, which show marked arteriolar changes.
It is claimed that with doses up to 1200 r the vascular changes are reversi-
ble, but that with higher doses permanent damage to endothelium and
especially to the smooth muscle and elastic fibers results. The swollen
or vacuolated endothelial cells may form tufts extending into the lumen.

1122 RADIATION BIOLOGY

and endothelial proliferation may be so marked as to occlude the vessels
(Wolbach, 1909). Some of the small vessels may be closed by thrombi.
Multinucleated endothelial cells in the derma one year after an erythema
dose and arteriolar change a year and a half after the same dose in
human skin have been described. Some of the dilated vessels may
persist for years. An important observation is that some vessels always
seem undamaged. After injection of radium, the endothelial cells may
calcify and there may be calcification of the walls of small vessels
(Rhoades, 1948b). Although vascular radiation lesions are well estab-
lished, some believe that their importance as the cause of many of the
changes observed in various irradiated organs has been exaggerated
(Lacassagne and Gricouroff, 1941). These critics believe that the part
played by obliterating arteritis in radionecrosis has not been proved;
they hold, rather, that transformations of extracellular substances of the
connective tissue and associated infection are much more important
factors in the pathogenesis of this process.

Some of the changes in the derma, especially in laboratory animals,
may be secondary to infections resulting from denudation of the derma
with ulcer formation.

The late effect in the derma is progressive sclerosis, resulting from the
inflammatory changes and characterized by hyalinization of the collagen
and loss of elastic fibers. These changes, coupled with, and perhaps in
part responsible for, the long-persisting, incomplete regeneration of the
epidermis, leave the skin highly susceptible to further injury.

A quite different picture is called forth in the connective tissue by the
local injection of those radioactive isotopes which adhere to collagen, such
as plutonium and Y91. In such areas the inflammatory reaction persists
for weeks or months, but is atypical in that there is a continued emigra-
tion from the vessels of granular and nongranular leukocytes, which are
apparently killed shortly after leaving the vessels. Consequently, there
is no flooding of the field with small, medium-sized, and large inflamma-
tory macrophages, which in ordinary inflammations develop from the
nongranular leukocytes of the blood. The local macrophages (histio-
cytes), which are relatively few for an inflamed tissue, do not divide by
mitosis but become enormously swollen and very basophilic; they do not
degenerate. The local fibroblasts likewise become swollen but show no
evidence of death. It is undoubtedly the continued irradiation which is
responsible for the failure of a typical inflammatory reaction to develop
around radium needles.

EPIDERMAL DERIVATIVES

Hair. It has been known for a long time that irradiation of the skin
causes destruction of the hair follicles and falling out of the hair. In

HISTOLOGICAL CHANGES AFTER IRRADIATION 1123

man this may be temporary, appearing in about three weeks with doses
of 400 to 500 r of 200-kv X rays, but becoming permanent with 700 r or
more (Shields Warren, 1942, 1943). There are marked species differ-
ences in this respect, the laboratory mammals requiring about 2000 r or
more of X rays to produce permanent epilation (Ellinger, 1941).

After temporary epilation in animals the hair may grow back white in
regions where previously it was pigmented (Hance and Murphy, 1926).
It has also been reported that regenerating hairs in man may be darker.
After a dose of radiation producing permanent epilation, the hair follicles
show vacuolation, cessation of mitosis, and focal necrosis. This is some-
times associated with a slight polymorphonuclear and mononuclear infil-
tration, but there is generally no intense leukocytic infiltration.

During the first month the smooth muscles of the hairs increase in size
with some vacuolar degeneration (Wolbach, 1909), followed by increasing
atrophy, ultimately often disappearing completely.

Nails. Occasional damage to finger nails has been reported.

Skin Glands. The sebaceous glands are also highly sensitive, reduction
of their secretion beginning 5 or 6 days after exposure (Ellinger, 1941).
Usually one month after a dose of radiation causing permanent epilation
few sebaceous glands will be visible, and those persisting frequently show
some peripheral keratinization as well as accentuation of the basement
membrane. It is also claimed that the sebaceous glands are destroyed in
three to four weeks with a dose of 1200 r (Borak, 1936). The sebaceous
glands tend to persist longer than the hair follicles but not quite so long
as the sweat glands.

The sweat glands are said to be as sensitive as are the hair follicles
(Flaskamp, 1930), a dose that produces temporary epilation decreasing
temporarily or inhibiting completely the secretion of sweat. There are
other reports which indicate that 2500 r is required to destroy these
glands (Borak, 1936). In the first few days vacuolar degeneration of
the epithelium is prominent, sometimes leading to great distinctness of
so-called "myoepithelial cells." About a month after irradiation, the
basement membrane is doubled or quadrupled in thickness, the lumens of
the glands have practically disappeared, and the connective tissue
adjacent to the coils is increased in amount and hyalinized. Later the
sweat glands may disappear completely.

Mammary Gland. The duct system of the mammary gland, which
constitutes the epithelium of the resting organ, seems to be markedly
radioresistant. It has been reported that 2800 to 3600 r is necessary to
prevent the development of secretory acini in the rabbit (Turner and
Gomez, 1936). A dose of 720 r applied during the middle of pregnancy
inhibits the subsequent ability of the epithelium to secrete milk. Lacta-
tion is completely suppressed after 2800 r, and this will persist through
succeeding pregnancies.

1124 RADIATION BIOLOGY

GASTROINTESTINAL SYSTEM

There are marked variations in the radiosensitivity of the several
types of epithelium lining the various segments of the digestive canal,
reflecting differences in their histological constitution. The stratified
squamous epithelia lining hollow viscera are about as radiosensitive as
the stratified squamous epithelium of the skin (Lacassagne and Gricouroff,
1941). This is presumably due to injury of the cells in the germinative
layer of the epithelium with resulting failure to produce new cells to
replace those more superficial ones which will be desquamated with or
without previous cornification. Those covering epithelia of the digestive
system composed of columnar cells are decidedly more resistant than the
stratified squamous epithelia. However, this columnar epithelium is
provided with areas of less differentiated, germinative cells and these have
been found to be relatively radiosensitive.

Associated with the digestive tract are the large salivary glands, the
liver, and the pancreas. These show contrasts in reactivity, the liver and
pancreas being very difficult to injure by radiation while the salivary
glands are somewhat more sensitive.

Mouth, Nasopharynx, and Esophagus. Except in a few areas these
cavities are lined with stratified squamous epithelium, and they show,
after irradiation, changes similar to those shown by the epidermis. In
man complete desquamation and subsequent repair in these mucous
membranes occur somewhat earlier than in the skin after comparable
doses of radiation, and variations in sensitivity in the mouth, tongue,
inner surfaces of the cheeks, and nasopharynx have been described
(Coutard, 1922). With larger doses, necrosis of the epithelia and the
underlying tissues may result. As in the skin, fragility of the epithelium,
marked scarring, and permanently dilated vessels persist. There seem to
be no extensive studies on experimental animals.

Stomach and Intestines. Destructive changes in the gastrointestinal
tract are noted as early as 3^ hour after exposure to a moderately large
dose (Pierce, 1948), the damage being greater in the small intestine than
in the stomach or colon, greatest in the crypts, most pronounced at 8
hours, and repaired within four weeks. With smaller doses the injury
is not so severe and repair is completed earlier, while with larger doses
(2000 r to the abdomen of dogs), the intestine may be so greatly damaged
as to lead to severe hemorrhages and ulceration with death of the animals
(Regaud, Nogier, and Lacassagne, 1912; Stafford L. Warren and Whipple,
1922). There are considerable species differences and unfortunately
many of the experiments are not comparable as to dosage and species.

In contrast to the paucity of reports of studies on the upper portions of
the digestive tract, there is a voluminous literature on experimental and
therapeutic radiation of the stomach. The changes are of two main

HISTOLOGICAL CHANGES AFTER IRRADIATION 1125

types: (1) the functional and degenerative morphological changes and
subsequent repair occurring shortly after irradiation and (2) the develop-
ment of gastric ulcers some weeks later.

In rabbits, after total-body irradiation with 800 r of 200-kv X rays,
some degenerate cells and absence of mitoses are seen early in the necks of
the fundic glands. Mitoses reappear after 17 hours but are not normal
until 5 days later. The surface epithelium and the cardiac and pyloric
glands are much more resistant. In the fundic glands some of the
parietal cells show wrinkling of the nuclear membrane, or pyknosis, and
vacuolization of the cytoplasm from 2 hours to 5 days in some cells, but
only during the first day in the majority of them. The more severely
affected mucous and zymogenic cells show pyknosis for 5 days. Zymogen
granules are absent from 2 hours to 5 days, and are, for the most part,
irregular in size and number until 9 days (Pierce, 1948). Hemorrhages
may occur, especially in the lamina propria (Engelstad, 1938).

There is agreement that irradiation causes a marked decrease in gastric
secretion with a great decrease in acidity and relatively little change in
pepsin content (Desjardins, 1931). It is noteworthy that in dogs the
parietal cells, presumably the source of hydrochloric acid precursors, are
not greatly damaged, in contrast to the destructive changes and loss of
zymogen granules in the chief cells (Dawson, 1925). This decrease in
production of acid after irradiation, which may last for some weeks, is the
basis for the treatment of gastric ulcers by large doses of X rays (Palmer
and Templeton, 1939). Many but not all authors report a decrease in
secretion of gastric mucus.

The production of ulcers of the stomach by irradiation has been noted
in man and animals (Delbet et at., 1909; Engelstad, 1938; Rickettsial,
1949). They develop after a latent period of several weeks or more and
usually occur singly along the lesser curvature. They may be shallow or
may penetrate all the tunics of the organ and lead to perforation. Scar-
ring and healing of the ulcers may occur after many months. It has been
suggested that the failure to heal sooner is due to the inhibition of pro-
liferation of connective tissue cells resulting from irradiation.

There are minor differences of opinion as to the relative sensitivity of
the small intestine as compared with other tissues and organs (Friedman,
1942). Part of this absence of unanimity is due to our inability to com-
pare dosages used in much of the experimental material before 1930, to
differences in species, and to large gaps in the series of animals reported by
many investigators. Nevertheless, the majority of authors agree that
the small intestine is more sensitive than the stomach and that it is only
slightly more resistant than lymphatic tissue.

Parts of the epithelium of the duodenum of the rabbit, more sensitive to
radiation than any other part of the gastrointestinal tract, show marked
degenerative changes 30 minutes after exposure to 800 r of X rays (total

1126 RADIATION BIOLOGY

body) and these sometimes persist for three weeks or longer (Pierce,
1948). At 30 minutes mitosis has ceased and there are varying degrees
of nuclear swelling and clumping of the chromatin in all cells of the
villous, crypt, and Brunnerian epithelium. Collapsed goblet cells appear
to be more numerous than normal. The most extreme damage is in the
basal cells of the crypts of Lieberkuhn, where nuclear fragmentation and
karyolysis are present for some days. Mitotic figures reappear at 21
hours, but until 9 days degenerative changes can be detected in the epi-
thelium. At this time regenerative changes become dominant, as evi-
denced by disappearance of degenerating cells, the presence of the usual
number of mitotic figures, and crowding of the basal crypt cells. The
Paneth cells, which have lost their normal staining properties 2 hours
after irradiation, show irregular granulation until 9 days.

The epithelium of Brunner's glands, which is apparently more resistant
than the villous or crypt epithelium, does not show marked degenerative
changes with this dose until the 8-hour interval. However, recovery
seems to be less rapid ; in some animals the tubules are still distended at
21 days.

After 800 r (total-body) the destructive process in the epithelium of
the ileum is less severe but is qualitatively similar to that of the duodenum
and recovery is more rapid. Mitotic activity of the crypt epithelium,
absent 30 minutes after exposure, is restored at 3 days. Degenerative
changes are most marked in the basal crypt cells. There is only very
slight nuclear damage in the epithelial cells of the colon and rectum.
Increased mucus secretion, which is suggested, is difficult to evaluate
because of normal variations in this function. Stafford L. Warren
believes that the colon is about as sensitive as the stomach and that the
more resistant rectum is about as sensitive as the mouth.

Early effects in the lamina propria are edema and absence of inflam-
matory cellular reaction. At 2 hours the number of small lymphocytes is
reduced, and many of the lymphocytes show degenerative change. How-
ever, there is less destruction of lymphoid cells here than in the lymphatic
follicles, for many appear to survive. There is a return of lymphocytes
and plasma cells after 9 days. The effects on the lymphatic follicles of
the ileum and appendix are much like those described in the lymphatic

tissue.

After 400 r in the rabbit the damage is qualitatively similar to that
produced by 800 r but quantitatively less severe, with fewer cells affected
and earlier recovery. One hundred roentgens causes only a depression
of mitosis and occasional dead crypt cells. Doses of 50 r or lower produce
no effect.

Total-body exposure of rabbits and mice to fast neutrons and of mice
to slow neutrons has the same qualitative effect on the intestinal mucosa
as total-body X irradiation.

HISTOLOGICAL CHANGES AFTER IRRADIATION 1127

With 80 r of total-body X irradiation per day to mice there is a slight
degree of acquired radioresistance so that nuclear debris in the crypts of
Lieberkiihn, although prominent during the first week, is rarely found
from 15 to 35 days (Pierce, 1948). No changes are observed in the
intestines of mice exposed daily to 8.8 r of y rays from radium when
examined at from two to sixteen months.

With larger doses (2000 to 2500 r) over the abdomen of dogs much more
drastic changes are produced (Stafford L. Warren and Whipple, 1922).
Within a few days the entire epithelium of crypts and villi is destroyed
and sloughed off. The intestine is inflamed and hemorrhages are exten-
sive. There is bloody diarrhea and the animals soon die. Rats and
guinea pigs are slightly more sensitive than dogs, cats, or rabbits (Stafford
L. Warren, 1936). It has also been found that ulceration of the small
intestine may result after two or three weeks (Martin and Rogers, 1924).

Internally administered radioactive isotopes affect the gastrointestinal
epithelium in the same way as do X rays, although some differences are
notable (Pierce, 1948). Radium and plutonium seem to cause more
nuclear swelling than the /3 and 7 emitters. With most of the parenter-
al^ administered isotopes there is no massive destruction of the epi-
thelium, as after a single 800-r dose of X rays. However, Zr93-Cb93,
injected intracardially, causes marked destructive lesions throughout the
small intestine, resulting in extensive areas of sloughed epithelium.
Parenterally administered Na24 and Ba14n-La140 also cause severe damage
to the gastrointestinal mucosa, notably during the first two weeks
following treatment, while P32 (2.5 /xc/g) produces moderate damage,
Y91 (2.0 Mc/g) produces less, and Sr89 (3.6 juc/g) still less. Changes in
the gastrointestinal tract following administration of the a emitters occur
at later intervals and with lower doses than with the (3 emitters.

The effects of Y91, Sr89, and fission-products mixture, administered by
gavage, vary with the dose level and length of interval during which
the agent may act. Large doses (23 to 33 /xc/g) of fission-products mix-
ture cause extreme necrosis of the mucosa in rats, most marked in the
lower ileum and colon. Smaller single doses of fission-products mixture
or of Y91 or daily doses of Y91 as high as 2 /xc/g for three months cause
minimal changes.

Salivary Glands. Damage to salivary glands has resulted from thera-
peutic irradiation of surrounding tissues. Functionally this is manifested
by a marked decrease in secretion of saliva (Bergonie and Speder, 1911;
Ivy et al., 1923). Histologically it has been found that the serous cells
are more sensitive than the mucous (although the reverse is held by
Lazarus-Barlow to be the case after exposure to radium) and both of
these more sensitive than duct epithelium (Salis, 1924). After severe
irradiation there is interstitial fibrosis of the organs. Desjardins, 1928,
reviewing the literature, states that the salivary glands are exceptionally

1128 RADIATION BIOLOGY

sensitive to irradiation. Presumably this is in comparison with other
glands. Systematic studies in laboratory animals seem to be lacking.

Liver. The liver is markedly radioresistant, although there are reports
of minor cytological changes. The most extensive and closely spaced
work as to time intervals and dosage is by Pohle and Bunting who
exposed the livers of adult rats to 600 to 2500 r at 100 and at 140 kv and
studied them at daily intervals for a month. Necrosis was not seen in
any liver. Minute cytoplasmic hydropic and fatty changes have been
reported, including temporary atrophy of liver cells, but such changes
may not have been due to irradiation. Hepatic cells may become
necrotic after a dose of 1880 to 5250 r of unfiltered X rays to the exposed
liver of dogs (Bollinger and Inglis, 1933) and after intravenous thorotrast,
which lodges in Kupffer cells.

An unusual change results from the persistence of plutonium and
yttrium in liver (Rhoades, 1948a). Enormous liver cells with budding
nuclei are present after some months, and many of the dividing nuclei are
clearly polyploid. Irradiation of regenerating liver in rats likewise causes
marked chromosomal aberrations (Brues and Rietz, 1951).

Gall Bladder and Biliary Passages. No changes were found in the
biliary epithelium after 2500 r of X rays (Pohle and Bunting, 1932). The
few who have examined the gall bladder believe it is definitely more
resistant than the intestine.

Pancreas. Both exocrine and endocrine portions of the pancreas are
markedly radioresistant. The hypoglycemia reported after irradiation
of the pancreatic region is probably not due to effects on the pancreas.
The claim that irradiation of exteriorized portions of the pancreas in
rabbits causes degeneration of the organ with 750 r and scarring after
1000 r (Seino, 1937) is not consonant with the other reports in the litera-
ture. In our own experience with 800 r total-body irradiation (200 kv) in
rabbits and 1000 r in chickens there is no effect.

URINARY SYSTEM

Kidney. Total-body irradiation of rabbits and rats with 800 or 600 r
of 200-kv X rays does not produce visible renal changes (W. Bloom,
1948c), although there are contradictory reports in the literature. On
the whole, it seems that doses higher than 2000 r are necessary to produce
damage to the nephron. The proximal convolution is reported to be the
most sensitive portion, and with doses above 3000 r vascular damage
produces secondary renal effects (Stafford L. Warren and Whipple, 1922).
It is also claimed that the kidney is more sensitive than muscle (Des-
jardins, 1924).

Radioactive isotopes lodging in the kidney, with the possible exception
of radium, have not been found to produce renal damage.

HISTOLOGICAL CHANGES AFTER IRRADIATION 1129

Urinary Bladder. There are but few reports on the reaction of this
organ to irradiation. The epithelium is resistant, although with high
doses used in man for therapy of lesions in neighboring organs, desquama-
tion of epithelium has been observed. The most marked changes involve
the connective tissue in which there is an early transient erythema and a
second erythema occurring some three or four weeks later. Ulceration
of the mucosa has been reported. Experimental studies are needed.

RESPIRATORY SYSTEM

The minute structure of the respiratory portion of the normal mam-
malian (and avian) lung is one of the unsolved problems of normal
histology. The problem revolves around the delineation of the struc-
tures intervening between the vast network of blood vessels carrying
blood for aeration and the air spaces. Linked with these histological
uncertainties are the difficulties of analyzing mild changes resulting from
any irritation and the frequent occurrence of inflammation in the lungs
of laboratory animals and man. The analysis is further complicated by
the relative radioresistance of the organ, it being decidedly less sensitive
than any hematopoietic organ, and by the intervention of a latent period
of two or three weeks. Part of the divergence of opinion in the literature
has resulted from differences in the intervals between divided doses.

Just as an erythema dose delivered to the skin has a slight transient
effect, so an equivalent dose delivered to the lung has a definite, though
again a transient, effect. Severe damage to the lung, however, usually
occurs only after the repeated doses of radiation customary for therapy,
radiation pneumonitis being a frequent complication of such treatment.

In an extensive study on rabbits receiving 1500 to 9000 r Engelstad
(1934) described four main stages in the pulmonary reaction:

1. An initial period with acute degenerative changes in the lymphatic
tissue of the lung, increased mucus production in the conducting passages,
hyperemia, and some edema with mild leukocytic infiltration in the
stroma and respiratory portion. The first changes begin 2 hours after
irradiation.

2. A latent period of two to three weeks.

3. Principal reaction with degenerative changes in the bronchial epi-
thelium and acute bronchopneumonic changes about the bronchi and
blood vessels. There is also a great increase in alveolar macrophages,
often forming giant cells. This stage may last one to two months and
gradually regresses.

4. Predominantly regenerative stage with proliferation of connective
tissue, with sclerosis (at times with bone formation) and slight prolifera-
tion of bronchial epithelium.

1130 RADIATION BIOLOGY

The process may still be continuing six months after irradiation. With
subepidermicidal doses the changes are mild although some fibrosis is left.
Epidermicidal doses produce extensive sclerosis, often with bone forma-
tion. Such doses are often lethal. The time-intensity factor is impor-
tant, since with fractionation and protracted dosing the changes are less.
As the pulmonary vessels were found to react strongly to pharmaco-
dynamic drugs in these animals, Engelstad does not believe that the
primary action is on the blood vessels. Cartilage in the conducting
passages is radioresistant to these doses. The bronchial epithelium shows
great abnormalities, and often becomes stratified and even cornified.
The pleura is resistant.

These findings have been amplified by Shields Warren and Gates (1940)
in studies on a variety of species, including man. They describe the
first change as swelling of cells lining the alveoli and atria. These cells
may not become larger than macrophages although they may show some
anaplasia, especially in man. In man, but only infrequently in animals,
a hyaline membrane is prominent in influenzal and other pneumonic
lesions, including irritant gas poisoning. After irradiation this membrane
is usually seen close to the alveolar wall in alveoli which are distended
and usually free of exudate. The membrane is not fibrin ; it is probably
an increased and condensed amorphous ground substance. Later, some
splitting of elastic fibers occurs. Bronchial epithelium, in contrast to
that of alveoli, is relatively stable. There may be some increase in
mucous cells. The columnar cells become cuboidal, often with loss of
cilia, but do not become stratified columnar. Bronchiolar epithelium is
probably as sensitive as that of the alveolar wall. The increase in size
and fusion of epithelial cells noted in animals do not occur in man. These
investigators conclude: "Epithelial anaplasia, alveolar and bronchial,
ruptured and reduplicated elastica, and hyaline membrane lining alveoli,
combined, we have seen in no other disease process. The first two of this
triad are seen in irradiated skin. The third occurs rarely in a variety of
inflammatory conditions in the lung ..." (Shields Warren and Gates,

1940).

Both post-mortem and experimental data have proved that the
roentgenologic pictures and clinical symptoms usually represent transi-
tory changes, which are generally harmless. In the late reaction, thicken-
ing of alveolar walls, patchy atelectasis, and vascular change are out-
standing, but the more acute change may be present also. Fibrosis,
though often present, has not been established as a radiation effect inde-
pendent of intercurrent infection. There is some reason for thinking
that most pleural fibrosis is a result of infection rather than injury from
irradiation; the mesothelium is radioresistant. There is some clinical
and experimental evidence that inflammation of the lung may render its
tissues more sensitive to radiation (Shields Warren and Gates, 1940).

HISTOLOGICAL CHANGES AFTER IRRADIATION 1131

NERVOUS SYSTEM

The nervous tissue of adult animals seems to be very resistant to injury
by irradiation (Shields Warren, 1942, 1943; Stafford L. Warren, 1936).
As might be expected, the nervous system of embryos and of newborn
animals does suffer severe damage, even after the period of multiplication
of cells is over (Snider, 1948b). The sheath and satellite cells seem to be
more sensitive than the neurons in the autonomic nervous system of
young chicks.

EYE

Shortly after the discovery of X rays it was found that irradiation pro-
duces a severe inflammation of the conjunctiva, and by 1908 it was
reported that cataracts very probably could be caused by irradiation
(Birch-Hirschfeld, 1908). Since then many studies have been made on
this subject and those before 1942 have been thoroughly reviewed (Poppe,
1942). It has been shown that 250 r produces permanent damage to this
organ in the rabbit (Poppe, 1942). All authors agree on the long latent
period required at approximately this dosage — as long as five to ten years
in man and at least five months in rabbits. Neutrons seem to be espe-
cially effective in producing cataracts (Evans, 1947). All mice exposed
to long-continued external j8 rays become blind owing to opacities of the
eye, presumably in the lens (Zirkle, personal communication).

ENDOCRINE GLANDS

Adrenal. The adrenal glands are among the more resistant organs,
and the reports of experiments involving them are unusually contra-
dictory (Desjardins, 1928; Shields Warren, 1942, 1943). This is in part
due to the extreme variations in the structure of the gland from species
to species and from animal to animal, and even in the same animal in
different physiological states.

In rabbits no histological changes are found after 1500 r given at one
time, but degenerative changes in the cortex develop after higher doses.
It has been reported (Engelstad and Torgersen, 1937) that adrenals of
rabbits which received a single dose of 2200 to 2500 r of X rays locally
show marked degeneration in the cortex, particularly in the zona fascic-
ulata and the zona reticularis, but no irradiation effects in the medulla.
An initial transitory hyperemia between 1 and 3 days after treatment is
followed by a pronounced degeneration starting at 6 days and accom-
panied by hyperemia and often slight lymphocytic infiltration. The
changes are greatest and most constant three months after irradiation,
but are still found at six months. These authors point out a parallelism
between a visible skin reaction to X irradiation and noticeable effects on

1132 RADIATION BIOLOGY

the adrenal cortex. The latent period preceding degenerative changes in
both may also be significant. In these experiments the glomerular zone,
which is probably the main source of regeneration of the cortex, is not
damaged, the changes occurring mainly in the zona fasciculata and zona
reticularis. However, some investigators report no changes in the
adrenals of rabbits and guinea pigs (Strauss, 1921). In general the
medulla seems more resistant than the cortex.

No clear-cut changes have been observed after injection of radioactive
isotopes, although some of these, such as plutonium, radium, and
Zr93-Cb93, lodge in the organ. After insertion of radium needles, how-
ever, necrosis does occur.

Hypophysis. Most of the effects of irradiation in this organ seem to
center in the anterior lobe, as degenerative changes in the other portions
of the gland have been described only after very heavy irradiation. It is
claimed that 3000 to 10,000 r at 180 kv produces a loss of body weight
(Fehr, 1936).

The reported hypophyseal changes are very indefinite and are usually
described as swelling and increase or decrease of acidophils (Fehr, 1936;
Podljaschuk, 1928). Young rats exposed to 2000 to 2500 r have been
said to show a reduction of 50 to 75 per cent in these cells and numerous
pyknotic nuclei in the chromophils (Lawrence, Nelson, and Wilson, 1937).
However, other reports hold that there are no changes in the cells of the
hypophysis (Fehr, 1936; Lacassagne and Gricouroff, 1941). Irradiation
of the hypophysis of young animals seems to depress the growth and
development of the animals.

Larger doses administered over the whole head do not change the
histological structure of the anterior hypophysis but are said to produce
an increase in its content of thyrotropic and gonadotropic hormones
(Grumbrecht et al., 1938).

Thyroid. It is almost universally agreed that the thyroid is moderately
radioresistant. Radium implants caused hemorrhage and necrosis of
the thyroid (Bower and Clark, 1923). Repeated X irradiation produced
no changes with doses lower than those leading to lesions of the skin
(Walters, Anson, and Ivy, 1931). Some diminution of hyperplasia was
noted in opossums after irradiation (Walters, Anson, and Ivy, 1932).

However, very large amounts of I131 are markedly effective in cancer of
the thyroid and also in removing symptoms in some cases of hyper-
thyroidism.

Parathyroid. Little is known of the effects of radiation on the
parathyroid.

DISCUSSION

It is of considerable theoretical and practical importance to evaluate
the relative radiosensitivities of the different kinds of cells and organs in

HISTOLOGICAL CHANGES AFTER IRRADIATION 1133

the body. Many such attempts have been made but all of them are
deficient, owing mainly to extensive gaps in available data and to a
failure to compare sensitivities for the same end effect. In Table 17-1
are listed the reactions of the cells when compared only for cell death in
nearly grown rabbits exposed to 800 r of total-body 200-kv X rays. A
similar table for lower doses would show many fewer dead cells. Data are
not available for a complete list at double or treble this dose or for
localized irradiation of each organ. Such tables do not consider many
important questions, such as the time of onset and the rapidity of cell
death or the rate of regeneration.

As early as 1906, Bergonie and Tribondeau felt that enough was known
to enable them to make some generalizations as to the types of cells
readily affected by irradiation: "The greater the reproductive activity
of the cells, the longer their mitotic course ('leur devenir karyokinetique'),
and the less definitively fixed their morphology and functions, the more
intense is the activity of X rays on these cells." This so-called "law"
obviously does not apply to the relatively great radioresistance of reticular
cells which are more primitive than the free blood-cell-forming cells to
which they give rise. Nor does it hold for the radioresistance of primitive
ovarian follicular cells as compared with their derivatives, the growing
follicular cells which are much more radiosensitive. There are numerous
other instances in which this generalization does not hold. In our
opinion it should be remembered only as an early attempt to focus atten-
tion on the greater susceptibility of growing organs and tissues containing
dividing cells as against static ones.

Although it is true that many strains of cells which are actively in
division at the time of irradiation are readily and quickly destroyed, and
while it is also true that in some cell strains the destructive effects of
irradiation become apparent only when the cells later undergo division,
the destructive effects of ionizing radiations are not always connected
with cellular reproduction. Thus, the small lymphocytes, which prac-
tically never divide, are among the most sensitive cells in the body. If
those theories of hematopoiesis which hold that lymphocytes are undiffer-
entiated cells are found to be true, then the sensitivity of these cells might
be a reflection of their mitotic potential. On the other hand, plasma
cells, which are recognized by all to be slightly changed lymphocytes, are
exceedingly resistant. Again, spermatocytes are in continuous division
(meiosis) and yet are not visibly affected by doses of radiation which will
destroy all spermatogonia. It is thus clear that mitosis per se is not a
prime factor determining radiosensitivity.

As is well known, small amounts of radiation may have no visible effect
on the mature sex cells, and yet striking developmental changes may
appear in the progeny. These effects are usually lethal mutations or
malformations. Perhaps similar results may occur in somatic cells in

1134

RADIATION BIOLOGY

Table 17-1. Viability of Cells of Nearly Grown Rabbit after Total-body
Irradiation with 800 r of 200-kv X Rays

All or Most Cells Die Many or Some Cells Die

No Cells Die

Erythroblasts
Lymphocytes (all sizes)
Myeloblasts (all sizes)
Myelocytes
Thymocytes
Megakaryocytes

Leukocytes (?)
Monocytes (?)

Mast cells

Lens fibers

Osteoblasts and osteocytes

of growing bone
Cells of growing cartilage

Spermatogonia

Primitive ova

Young growing ova

Mature ova

Growing follicular cells

Cells of gastric pits
Crypt cells of small intes-
tine

Erythrocytes

Fibroblasts

Reticular cells

Macrophages

Pigment cells

Fat cells

Plasma cells

Endothelium

Mesothelium

Striated muscle

Cardiac muscle

Smooth muscle

Neurons and neuroglia

Retina

Osteocytes of mature bone

Osteoclasts

Colls of resting cartilage

Sertoli cells

Spermatocytes (?)

Spermatids

Spermia

Interstitial cells of testis and
ovary

Follicular epithelium of primi-
tive ova

Lutein cells

Fallopian tube

Endometrium

Vagina

Cornea

Epidermis

Hair

Sebaceous gland

Sudoriferous gland

Mammary gland

Esophagus

Bronchial epithelium

Surface epithelium of stomach

Surface epithelium of intestine

Hepatic and biliary epithelium

Pancreatic cells

Specific cells of all endocrine
glands

Kidney

Urinary bladder

HISTOLOGICAL CHANGES AFTER IRRADIATION 1135

which no visible changes occur while they are in the resting state, but
which do appear as abnormal, often multipolar, mitoses when the cells
undergo division, frequently leading to death of the cells. Somatic
lethal mutations or chromosome aberrations may perhaps explain the
death of cells long after irradiation of their parent cells (Muller, 1950).
This may be the explanation of the failure to persist of early regeneration
of erythroblasts and myelocytes in the beginning recovery of marrow
made aplastic by irradiation. In the latter case it may be that the
progeny of the irradiated, but not visibly damaged, reticular cells are
destined to develop somatic lethal mutations. Against this suggestion is
the fact that attempts at regeneration in this tissue seem to occur in
waves, whereas, on the proposed explanation, it should be a continuous
process with gradual predominance of those myelopoietic cells which had
not been damaged.

The normal hepatic epithelial cell undergoes mitosis only rarely. If a
large part of the liver is removed, the remaining liver cells undergo such
tremendous mitotic activity that in a few days a large part of the liver is
regenerated — this process, as might be expected, being faster in young
rats than in older ones. The liver cells normally are markedly resistant
to radiation. But the rapidly dividing cells of regenerating liver are
easily damaged by radiation, which calls forth great numbers of chromo-
some aberrations and cells with multipolar mitoses (Brues and Rietz,
1951). It is quite probable that if the liver cells were irradiated suffici-
ently intensely before partial hepatectomy, the irradiation damage which
would become manifest in the regenerating liver would be so extensive as
to interfere with or even prohibit the regenerative process.

One cannot speak of the absolute sensitivity of a particular cell type in
a complex organism, for the cell in question cannot be dissociated from its
environment and its functional state at a given moment. Changes in
environment affect its response to irradiation just as changes in its
activity also modify its reaction. Examples are seen in the greater
sensitivity of lymphocytes in lymphatic nodules compared to those
scattered in the lamina propria, or in bone cells of the metaphysis as
opposed to the more resistant cells of compact bone. Furthermore, every
cell strain becomes more radioresistant in vitro.

The radiosensitivity of an organ is not simply a reflection of the radio-
sensitivity of certain of its cells, for one organ may have tremendous
regenerative capacities while another may not. For instance, blood cell
formation may be wiped out from the bone marrow of a femur in a few
days after irradiation, but this marrow can regenerate completely from
its more radioresistant reticular cells. However, with the same amount
of radiation, an area of skin may require weeks to manifest its complete
damage and regeneration will come only from surrounding nonirradiated
skin; furthermore, the friable skin may persist for years. It is difficult to

1136 RADIATION BIOLOGY

decide which of these two tissues is the more radiosensitive unless it
is stipulated whether one is concerned with acute or chronic injury.
Another type of difference is shown after irradiation of ovary and testis.
The ovary may be permanently sterilized for the production of ova and
thus indirectly of ovarian hormones, while in the testis spermatogenesis
can return after complete but temporary sterilization, and the production
of androgen is at no time diminished. In fact, in the testis, spermato-
cytogenesis is relatively radiosensitive and spermiogenesis is radio-
resistant, but the organ as a whole is potentially radioresistant for both
external and internal secretion. A third kind of reaction is that of the
lens with its lack of acute damage after moderate irradiation, but which,
after a long latent period, may show irreversible cataract formation from
such a dose. Thus we must distinguish between acute and chronic
radiation effects, and it must be determined in each case whether there
is a potential for recovery or not.

In the gonads there is clear-cut evidence of the cumulative effect of
repeated small doses of irradiation. A similar but less intense effect has
been demonstrated in other organs, as the damage which results from
localized radioactive isotopes and as the damage in spleen, thymus, and
bone marrow after long-continued (eight to sixteen months) daily doses
of 4.4 or 8.8 r per day. Variations on these dose-time relations, as well
as the length of recovery periods between exposures, have not been
thoroughly studied. In the crypt epithelium of the mouse, repeated
daily exposure to 60 or 80 r per day of total-body X rays leads to a definite
but small degree of acquired radioresistance (M. A. Bloom, 1950).

There is considerable evidence in favor of an indirect effect of irradia-
tion. The symptoms of "radiation sickness" substantiate strongly the
existence of such an effect. From morphological data it would seem
that the best evidence rests on the following observations: (1) Radiation
elicits a greater response in a particular organ when the same amount is
applied to the whole body than when it is applied locally; and (2) local
irradiation sometimes produces destructive changes in distant tissues.
These conclusions are based on innumerable investigations carried out to
study other aspects of radiation, and also on specific studies (Halber-
staedter and Ickowicz, 1947; Jolles, 1950; Ssipowsky, 1934-35; Van
Dyke and Huff, 1949). For example, in the thymus, pyknosis of the
cells is more intense after total-body irradiation than after irradiation of
either the superior or inferior part of the body (in each instance, 2000 r of
X rays) (Halberstaedter and Ickowicz, 1947). Atrophy of the lead-
protected thymus and spleen of rats takes place when other regions of
the body are irradiated with several thousand roentgens of X rays
(Leblond and Segal, 1942). However, when rats have been adrenal-
ectomized several days before irradiation, this indirect effect on thymus
and spleen does not occur. These findings have been corroborated

HISTOLOGICAL CHANGES AFTER IRRADIATION 1137

(Halberstaedter and Ickowicz, 1947). Adrenal hypertrophy and fatty
infiltration of the liver, not reported by careful observers after direct
irradiation of either the adrenals or liver, are observed as a secondary
toxic effect of roentgen rays (Leblond and Segal, 1942).

The importance of the spleen (and other lymphatic organs) in radiation
injury is emphasized by experiments in which the LD50 for mice was found
to be nearly twice as great when the spleen was protected with lead during
total-body irradiation as when it was not (Jacobson et al., 1949).
Such protection of the spleen during irradiation of the rest of the body
not only lengthens the survival time of these mice, but also lessens
the injury to their irradiated lymphatic tissues and bone marrow; these
show greater powers of regeneration than when the whole body is
irradiated (Jacobson et al, 1950)— further evidence of the fact that,
whereas the hematopoietic process is very radiosensitive, the hemato-
poietic organs are potentially radioresistant.

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1140 RADIATION BIOLOGY

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HISTOLOGICAL CHANGES AFTER IRRADIATION 1141

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1142 RADIATION BIOLOGY

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HISTOLOGICAL CHANGES AFTER IRRADIATION 1143

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Manuscript received by the editor Apr. 6, 1951

CHAPTER 18

Carcinogenesis by Ionizing Radiations

Jacob Furth1
Biology Division, Oak Ridge National Laboratory

AND EGON LORENZ

National Cancer Institute, Bethesda, Md.

Introduction. Physical aspects of carcinogenesis by ionizing radiations. Historical.
Neoplasia induced by X and gamma radiation. Neoplasia induced by radioactive sub-
stances: Artificial radioactive isotopes. Neutrons. Organ susceptibility to tumor forma-
tion. Leukemia: Leukemia among radiologists — Experimental induction of leukemia.
Endocrine organs: Gonads — Pituitary tumors — Thyroid — Parathyroid. Mammary
gland. Uterus. Skin. Lung: Lung tumors among miners of Schneeberg and Jdchymov
— Experimental induction of lung tumors in radioactive mines — Experimental induction
of lung tumors by X or gamma radiation. Bone: Bone tumors in radium dial painters —
Experimental production of bone tumors. Liver. Gastrointestinal tract. Brain.
General comments: Cocarcinogenesis by irradiation. Mechanism of carcinogenesis.
References.

INTRODUCTION

During the past fifty years a mass of data has been accumulated on the
induction of cancer in man and many species of animals by various types
of ionizing radiations disclosing numerous factors. The need is felt to
review these in historical perspective. Recent developments are being
widely disseminated by scientists and their institutions, but little is
known of past achievements in which much recent work is rooted.
Excellent work by pioneers, many of whom died with cancer induced by
irradiation, paved the way to safety measures enabling now the handling
of large amounts of radioactive substances with perfect safety. Good
monographs on the subject in French by Lacassagne (1935a, b) are avail-
able, but neither these nor the work of their author is widely known, even
though the originality of his contributions is hardly matched in depth by
more recent work. Among the older monographs on the subject of
irradiation injuries in man and animals, those of Colwell and Russ (1924,
1934) and Colwell (1935) are outstanding. Excellent recent reviews on

1 Work at Oak Ridge and preparation of manuscript were performed under Con-
tract No. W-7405-eng-26 for the Atomic Energy Commission.

1145

1146 RADIATION BIOLOGY

this subject are those of Brues (1951) and several chapters in "Medical
Physics," of Glasser (1950). In locating and interpreting the enormous
literature we have made much use of these publications.

The induction of neoplasia by ionizing radiations was noted accidentally
soon after discovery of X radiation and radioactive substances and the
clinical observations were soon verified by experimental studies in
animals. Exposure to ionizing radiations arose as a new industrial and
medical hazard and also as a tool of experimental carcinogenesis and
cancer therapy. The artificial production of radioactive substances has
increased tremendously the potentialities of application of ionizing radia-
tions to varied fields of science and industry, but accidental or incidental
exposure to large quantities of radiation has also greatly multiplied the
hazards of neoplasia induction.

The use of ionizing radiations in science and industry is now in rapid
ascendency; knowledge of the carcinogenic potencies of radioactive sub-
stances is still much limited. The present review aims to survey the pub-
lished observations and to point to deficiencies of knowledge concerning
carcinogenesis by ionizing radiations.

PHYSICAL ASPECTS OF CARCINOGENESIS BY IONIZING RADIATIONS

The interaction of ionizing radiations with biologic matter is discussed
and the term of specific ionization or linear energy transfer is defined in
other chapters. The specific ionization in biologic matter varies from a
theoretical 6.3 ion pairs per micron (Gray, 1946) approached by the X
radiation in the multimillion-volt range to several thousand for the a par-
ticles and many thousand for the heavy fission particles. This difference
in specific ionization causes marked quantitative differences in the
biologic effects on small biologic entities. Similar quantitative differ-
ences may not exist as concerns the acute lethal syndrome in animals.
When calculated for the same absorbed energy with equal dose rates, the
30-day LD50 for mice is approximately the same after X irradiation as
after internal a irradiation from intravenously injected radon in equi-
librium with its short-lived decay products (Hollcroft and Lorenz, 1952).
The slight difference observed might be explained by the nonuniformity
in distribution of the a emitters. On the other hand, Zirkle (Chap. 6)
has shown that cyclotron neutrons are approximately five times and
X radiation 1.5 times as effective as y radiation in killing mice (30-day
LD50). The application of such findings to radiation carcinogenesis is,
however, merely speculative. The scant data in the literature (Henshaw
et at., 1947, and Upton and Furth, 1951, unpublished data) indicate no
marked differences in tumor induction by neutrons as compared to X
or 7 radiation.

The following generalizations apply to the process of carcinogenesis:
If the tumor induction is caused by a direct action of the radiation on

CARCINOGENESIS BY IONIZING RADIATIONS 1147

cells and if a single ionization event can cause a carcinogenic change, the
carcinogenic action should be independent of dose rate and dependent on
a minimum dose, and X radiation will be more effective than a particles
or neutrons. On the other hand, if several ionization events are necessary
for the carcinogenic change, the action of the radiation might depend on
the dose rate as concerns particles of low specific ionization (such as hard
X or 7 radiation), but would be independent of dose rate for heavily
ionizing particles. This may apply to irradiation of small volumes of
highly susceptible cells, e.g., the induction of bone tumors by localized
a emitters. The situation becomes complicated, however, after whole-
body irradiation when the systemic secondary reactions caused by the
sum of the primary effects are dominant over the primary injury, as may
be the case with induction of leukemia or ovarian tumors.

In order to arrive at a quantitative relationship, the roentgen as a dose
unit is adequate for irradiation with 7 or X radiation up to a few Mev,
and enables a correlation of data for whole- or part-body irradiation,
acute fractionated, or long-continued irradiations. When dealing with
corpuscular reactions, the roentgen is of little value, especially if the
radiation is localized and the particles have path lengths in tissues rang-
ing from several n to several mm. However, the number of particles
absorbed and their average energy can be determined, and this energy can
be converted to rep. The energy absorbed in 1 g of tissue by 1 r being
approximately 93 ergs, the unit rep (roentgen equivalent physical) is
defined as that amount of absorbed energy of corpuscular radiation that
equals 93 ergs per g of tissue (Lea, 1947). Specific ionization does not
enter into the determination of the tissue dose in rep. Since data
on the relative efficiency of radiations of different specific ionizations on
mammals are scant, but striking differences are observed on small
biologic objects, the unit of rem (roentgen equivalent mammal) has been
introduced. One rem equals 1 rep divided by an arbitrary factor that
expresses the greater biologic effectiveness of corpuscular radiation of
high ion density over X or 7 radiation, e.g., as concerns fast neutrons this

L rep
factor is estimated to be about 5. Hence, 1 rem = — ^ — = 0.2 rep.

This brief outline of the physical aspects of ionizing radiations points to
difficulties encountered in obtaining quantitative data on the carcinogenic
effects. While the problem of obtaining such data for X or 7 radiation
is relatively simple, comparison of such data with those from radiations
of different specific ionizations is exceedingly difficult, because a uniform
tissue dose cannot be obtained easily by total-body irradiation with
corpuscular radiation. When bone tumors are produced by X radiation,
the determination of the tissue dose is simple. Bone tumors are more
easily produced by /3 or a emitters. The amount of radioactive material
deposited in the bone can be measured, but no quantitative comparison
can be made with X radiation on the basis of an average computed dose,

1148 RADIATION BIOLOGY

because the radioactive material is not uniformly distributed and neo-
plastic transformation may occur at points of greatest density of the
radioactive material.

The maximum permissible amount of a radioactive element in micro-
curies fixed in the body of man as revised in "International Recommenda-
tions on Radiological Protection" (1951) is as follows: Ra226, 0.1; Pu239,
0.04; Sr89, 2.0; Sr90 ( + Y90), 1.0; Po210, 0.005; H3, 10; C14 (as carbon
dioxide in air), none; Na24, 15; P32, 10; Co60, 1; I131, 0.3 (0.18 in thyroid).
Additional data on these elements, such as effective mean life, permissible
daily deposition in body, proportion absorbed via lungs and retained in
the body, proportion retained after absorption from the gut, and maxi-
mum permissible level in liquid media and in air are listed.

HISTORICAL

The first published account of the injurious effects of X irradiation,
such as dermatitis and alopecia, was made in 1896 by Marcuse. Soon it
became apparent that these skin lesions are excruciatingly painful, slow
to heal, and likely to break down and terminate in cancer.

The dramatic sequence of events leading to carcinoma from exposure to
X radiation is well recorded by Hall-Edwards in his autobiographical
notes (1904, 1906) which will be read with interest and respect by those
interested in the history of science.

Hall-Edwards commenced his clinical research a few weeks after
publication of Roentgen's discovery. In 1896 he gave a series of demon-
strations in fluoroscopy, on each occasion exposing his hands for several
hours. A primitive X-ray tube in operation is shown in Fig. 18-1 and the
wide-open exposure of the fluoroscopist is evident. Some two or three
weeks after exposure, Hall-Edwards noticed that the skin around the
roots of the nails was red and painful. Thus commenced the lesions
described objectively in their full tragedy in his first communication in
1904. During the subsequent two years he did "not experience a
moment's freedom from pain . . . and was rendered absolutely incapable
of work, either mental or otherwise." On the back of each hand fifty to
sixty warts appeared, many of them confluent.. In 1908 his forearm was
amputated, and he died in 1926. "As far as I know of all that early band
of workers who persisted in practicing Radiography, he is the only one
who escaped death from malignant disease" (Barling, 1926).

The first cancer arising in radiologists was described by Frieben (1902)
and Sick (1903), who made the diagnosis, and the next by Lloyd (1903).

The first American martyr of X irradiation was Clarence Madison
Dally, an assistant of Thomas A. Edison in research on the fluoroscope.
Seven years after the first exposure in 1896 he developed a cancer at the
site of an X-ray ulcer of the hand. This was not controlled by amputa-

CARCINOGENESIS BY IONIZING RADIATIONS

1149

tion, and he died with metastases (Brown, 1936). Elizabeth Fleischman
Aschheim was one of the first, if not the first, clinical fmoroscopist. Her
exposure to X radiation began in 1897, and she died in 1905 with squam-
ous carcinoma of the fingers (Brown, 1936). Another early American
martyr of irradiation was Kassabian. He wrote sixteen papers on effects
of X irradiation and a textbook of roentgenology; the latter reached a
second edition in the year of his death (1910).

Porter and White (1907) reviewed the first eleven verified fatal cases
of X-irradiation cancers in man. The sequence of events in their first
patient is characteristic. Although he stopped all X-radiation work for a

Fig. 18-1. A pioneer roentgenologist fully exposing himself to X rays while examining
a patient and testing the setup by observing the opacity of bones of his hand {after
Brown, 1936).

year, ulcers developed upon his fingers, and after the first demonstration
of cancer in 1902 malignant degeneration occurred in eight different
areas. Porter and White were impressed by the multiplicity of cancers
produced by X irradiation.

When Rowntree (1909) gave the Hunterian lecture on cancer induction
by X irradiation, it was already well established that dermatitis caused
by repeated exposures to small doses of X radiation was frequently
followed by the development of squamous cell carcinoma (Rowntree,
1908). X-radiation burns usually began with blistering of the skin, after
which the exposure was discontinued temporarily. In the course of time
warts appeared, some of which became larger and their base thickened.
Some warts ulcerated and fell off, but at their base malignant cells
extended into the surrounding tissue. Nearly all such cancers were
multiple and occurred on both hands. Rowntree pointed out that chronic
irritations can be caused by many other agents, but these are seldom

1150

RADIATION BIOLOGY

complicated by cancers; malignant growths can, however, be caused also
by ultraviolet radiation.

A monograph by Hesse (1911) records fifty-four cases of roentgen
cancers. The pioneering spirit finds its expression in the large number of
cases reported from the United States of America (26) ; from Germany and
England each, 13 cases were recorded. Twenty-six of the fifty-four
patients were physicians and twenty-four, X-ray technicians.

Feygin (1914) surveyed a number of cases of radiation-induced cancers
in relation to latent periods and duration of exposure as follows:

No. of years after first irradiation

14

13

12
4

11
4

10

9

8

7

6

5

4

3

2

1

No. of cases

2

4

5

4

6

5

5

3

2

No. of years after radiodermatitis

11

10

9

8

7

6

5

4

3

2

1

No. of cases

3

6

3

3

3

9

4

1

2

0

1

The observations in man were soon verified by experimental studies in
animals by Marie et al. (1910, 1912) and his student, Clunet (1910), who
produced sarcomas in rats by repeated administration of X radiation.
Imitating the sequence of events in man, they waited until ulcers formed
and healed and then administered more X radiation.

The similarity between the cutaneous changes caused by X irradiation
was recognized in 1900 by Walkhoff. The first casualty (1901) from
overexposure to radium radiations was Henri Becquerel himself. He
carried in his vest pocket radium enclosed in a glass tube. Erythema of
the skin followed by ulceration marked the site of the exposure. The
ulcer healed completely a month afterward (see Colwell and Russ, 1934).
It is now well established that a single local exposure is likely to cause a
neoplasm only under exceptional conditions, but a single massive expo-
sure to either X or y radiation over the entire body may often cause
cancers in some internal organs.

Wolbach (1909) and Ordway (1915), working under the Cancer Com-
mission of Harvard University, studied in detail the injurious effects in
handlers of radioactive substances. The very first changes consisted
chiefly of flattening of the characteristic ridges, thickening and scaling
of the superficial layers of the skin, atrophy and intractable ulcerations.
These objective changes were slight compared with the subjective persist-
ing symptoms such as paresthesia, anesthesia, tenderness, throbbing,
and pain. Wolbach concluded that not a single trauma but long-
continued progressive lesions of connective tissue supporting the epi-
thelium are responsible for the acquisition of malignant properties.

CARCINOGENESIS BY IONIZING RADIATIONS 1151

The first two cases of carcinomas of the skin from exposure to radia-
tions from radium occurred in handlers of radium tubes and applicators
and were described in 1923 by MacNeal and Willis and in 1927 by
Wakely. Both patients had years of repeated exposures. The lesions
began with numbness in finger tips, roughness of the skin of the hand,
and tingling sensation; later the nails were friable and fissured, the skin
atrophic and thin but not ulcerated. Telangiectasias appeared and light
brown pigmentations and hyperkeratoses; one of the last showed, on
section, a squamous-cell carcinoma. These, as many other patients
suffering from neoplasms induced by irradiation, were treated with
radium, X radiation, and sunlight, which are carcinogenic (Colwell and
Russ, 1934). This paradox is frequently discussed. Treatment with
ionizing radiation causes regression of neoplasia within days or weeks
but is carcinogenic only after several years and only under special
circumstances.

Lazarus-Barlow (1918, 1922) was the first to induce cancer of the skin
by radioactive substances. He introduced radium sulfate or silicate into
the subcutaneous tissue of mice and observed the development of five
malignant tumors of the skin in sixty-seven animals surviving six months.
He was also believed to have induced cancer of the gallbladder by insert-
ing radium in cholesterol stones and placing the stones in the gallbladder,
but this was disputed by others. The character of an epithelial prolifera-
tion is often difficult to assess and extensive hyperplasia may mimic neo-
plasia, notably in the gallbladder or uterus, where, without being neo-
plastic, nests of epithelial cells may be established in deeper layers.

Slowly, clinical reports multiplied on induction of neoplasms in man by
therapeutic irradiations. On the other hand, experimental work on this
subject slackened until World War II, when developments in nuclear
physics introduced the use of ionizing radiation in warfare and expanded
tremendously its use in medicine and industry.

NEOPLASIA INDUCED BY X AND GAMMA RADIATION

Since 1902 when skin tumors were first observed on hands of persons
who developed radiodermatitis, clinical reports appeared in steadily
increasing numbers, at first on the development of cutaneous and later
on internal tumors. The prophecy "La radiotherapie ne donne pas les
cancers" (Belot, 1910) is now a reminder of the danger of drawing such
negative conclusions. In 1911 Hesse surveyed a total of ninety-four
cases, to which 126 cases were added by Krause in 1930. The clinical
aspects of malignant growths induced in man are described in over thirty
publications which cannot be cited here. Most of these deal with
tumors of the skin and only a few with those of deep-seated organs, e.g.,
uterus, ovary, and larynx, and it became a matter of dispute whether

1152

RADIATION BIOLOGY

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CARCINOGENESIS BY IONIZING RADIATIONS 1153

the latter were actually produced by X irradiation. No such doubt
exists as concerns tumors in bones which are particularly susceptible,
notably if inflamed; but even apparently normal bone, when subjected to
massive irradiation, may undergo a neoplastic change (Spitz and Higin-
botham, 1951).

Experimental induction of neoplasms in animals by X irradiation
began eight years after the first observations in man. The procedures
used imitated the conditions of exposure in man. The early literature is
surveyed in Table 18-1. In his monograph, "Les Cancers Produits par
les Rayonnements Electromagnetiques," Lacassagne (1945b) discusses
extensively the early literature which will not be fully reviewed here.

Marie et al. (1910, 1912) and Clunet (1910) gave repeated doses to rats
estimated by Lacassagne to be about 600 to 2000 r during four to twelve
months and so induced spindle-celled sarcomas. For many years remark-
ably little use was made of this novel type of carcinogenesis. Bloch
(1923, 1924) and Schurch (1930) induced carcinoma in the rabbit by
repeated exposures to X radiation. Subsequent work on the induction
of cutaneous sarcomas in guinea pigs (Goebel and Gerard, 1925), cutane-
ous carcinomas in mice, and bone tumors in rabbits is surveyed in Table
18-1.

The tumors of the skin were induced by relatively large doses of ion-
izing radiations at the sites of exposure. It is doubtful if any species
would resist this type of carcinogenesis. The next important observa-
tion was that, following total-body exposures, neoplasms, leukemias,
ovarian, mammary, and other tumors appeared in different organs.
Mammary tumors were also induced in rats and rabbits. Subsequent
research aimed to elucidate the pathogenesis and morphogenesis of the
induction of internal neoplasms and to determine the relevance of
observations made in one species to the animal kingdom at large. At
first, single or a few large doses of X radiation were used in these studies ;
later, also 7 radiation and chronic exposures. A large-scale chronic
exposure study in mice, guinea pigs, and rabbits using 7 radiation of
radium was carried out by Lorenz et al. (1947). In mice there was a
marked increase in the incidence of leukemia in the females of the groups
exposed to 4.4 and 8.8 r (8 hours) daily (Fig. 18-2), an even greater
increase in the incidence of ovarian tumors, and a slight increase in
incidence of breast and lung tumors. In guinea pigs, there was an
increase of mammary tumors, and in rabbits of uterine tumors. This
and other recent work are surveyed in the sections which follow.

NEOPLASIA INDUCED BY RADIOACTIVE SUBSTANCES

While experimental induction of cancers by X irradiation followed, that
by radioactive substances preceded the observations on man. Lazarus-

1154

RADIATION BIOLOGY

Barlow (1918) was the first to report on the experimental induction of
carcinomas by means of radium. Although doubt was expressed as to
the neoplastic nature of the epithelial proliferative changes described by
him, his experimental pattern was followed by others who fully confirmed
his claim. The first "radium carcinoma" was described by Wakely
(1927) on the thumb of a man handling radium salts. In his first studies,
Lazarus-Barlow produced carcinomas of the skin in mice and rats (1918).
Later (1922) he reported on the induction of carcinoma of the gallbladder
by inserting radium into gallstones and placing such stones in the gall-
bladder. Investigations performed during the next two decades on the

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Fig. 18-2. The incidence of leukemia in female mice after chronic total-body exposure
to 7 rays of radium (after Lorenz et ah, 1947).

carcinogenic effects of radium, radon, mesothorium, and thorium are
surveyed in Table 18-2.

Daels (1925) inserted radium into the subcutaneous tissue of mice and
rats, inducing sarcomas after seven to twelve months. About a year
later he described the induction of carcinomas by the same procedure.
Daels and Biltris (1931) soaked silk strings or tissues in radium chloride
solution, coated these with collodion, and inserted them in various tissues
of rats and guinea pigs, inducing sarcomas of the kidney, carcinomas of
the liver, and intracranial sarcomas after twelve to thirty months. The
similarity of action of this carcinogen to that of chemical carcinogens such
as methylcholanthrene, discovered later, is noteworthy. The type and
readiness of neoplasia induction seem to depend on the tissue, species,
and other factors, not on the carcinogen. When inserted in the kidney,
both chemical (e.g., methylcholanthrene) and physical carcinogens induce

CARCINOGENESIS BY IONIZING RADIATIONS

1155

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1156

RADIATION BIOLOGY

sarcomas but do not render the renal epithelium neoplastic. The inci-
dence of induced tumors (Daels and Biltris, 1931) was low, many animals
dying too early of intercurrent disease. Biltris (1933) using the above
radium-string technique, in which the carcinogenic agents were /3 and y
radiations, induced in guinea pigs sarcomas of the meninges, metastasiz-
ing carcinomas of the gallbladder, and malignant tumors of the spleen
after a latency of eight to twenty-four months. In their last publication
Daels and Biltris (1937) described the induction of osteosarcoma in two
chickens by the collodion-string technique after a latency period of

nearly five years.

Following the work of Schurch and
Uehlinger (1931, 1947) (Fig. 18-3), numer-
ous articles appeared on the induction of
bone tumors in rabbits. These investiga-
tors implanted 1 mg of radium sulfate sub-
cutaneously in the jaw of a rabbit and left
it there for 20 days. The tumor appeared
one and one-half years afterward. Later
(1935) they placed vaseline impregnated
with 2-5 mg of mesothorium in the femora
of rabbits, inducing osteo- and poly-
morphous sarcomas. Sabin et al. (1932)
induced bone tumors in rabbits by intrave-
nous injection of radium chloride (approxi-
mately 5.1 mg) and mesothorium (approxi-
mately 7.7 mg) in eleven to nineteen
months.

Ross (1936) implanted in the thoracic
wall of rabbits 0.1 mg of radium enclosed
in platinum tubes 1 cm long and with a wall thickness of 0.5 mm.
Earlier, the experimental induction of neoplasms had been successful
only when unscreened radioactive substance was used, with the exception
of some work of Schurch and Uehlinger, and the opinion was held that
a radiation was the principal carcinogenic factor. However, six of the
nine experimental animals of Ross developed a tumor adjacent to the
tubes ; one had an osteosarcoma, the others nonosseous neoplasms, indi-
cating that 7 radiation also induced tumors. These and earlier studies
are surveyed in Table 18-2.

Petrov and Krotkina (1933, 1947), repeating the disputed work of
Lazarus-Barlow, implanted radium needles in the gallbladder of guinea
pigs and induced carcinomas of the biliary tract. The agent causing
spontaneous carcinoma of the gallbladder in man is not known; it is not
likely to be a radioactive substance. Therefore, such studies do not
contribute to the knowledge of spontaneous tumors or those likely to be

Fig. 18-3. Osteogenic sarcoma
induced in a rabbit by injections
of thorotrast (after Schurch and
Uehlinger, 1935.)

«

CARCINOGENESIS BY IONIZING RADIATIONS 1157

caused in man by radioactivity, but they may disclose sensitivities to
neoplastic changes.

Roussy et al. (1934) were first to report on the experimental induction
of tumors by thorium dioxide (thorotrast) by intraperitoneal injections of
this substance into rats. Subsequently, several workers induced sar-
comas by subcutaneous injection of thorium dioxide in rats and mice
(Roussy et al., 1936; Selbie, 1936; Miyamota, 1939; Prussia, 1936;
Bogliolo, 1937, 1938; and Selbie, 1938. The tumors so induced by
Selbie were spindle-cell sarcomas (six), osteosarcomas (three), and one
hemangioendothelioma. This recalls the extraosseous bone tumors
induced in man by X irradiation and the hemangiosarcoma induced by
radium. Bogliolo induced tumors in all of twenty-three rats that had
been given subcutaneous injections of thorotrast. In guinea pigs
Foulds (1939) induced sarcomas and a carcinoma by repeated injections
of thorotrast in the nipple after a latency of three years. Andervont
and Shimkin (1940) confirmed the induction of sarcoma and hemangioma
in mice by subcutaneous injection of this substance and called attention
to the absence of lung tumors in their mice, even though radioactive
particles were demonstrated in the lungs throughout the course of the
experiment and the strain of mice used was highly susceptible to lung
tumors. Dunlap et al. (1944) fed rats each with 100 ^g of radium and
produced osteosarcomas in nine of thirteen animals.

The statements of Roussy and Guerin (1941), and later of Willis
(1948), that tumors may appear in human beings who had been given
thorotrast for diagnostic purposes, proved valid. The use of thorotrast
is relatively recent and many years may elapse before tumors develop.
An endothelial sarcoma developing in the liver of a seventy-year-old
woman, following thorotrast injection, was described by MacMahon
et al. (1947). Thorotrast taken up by macrophages was demonstrated
microscopically in tissue sections, and this phagocytic activity was cor-
related with radioactivity. In spite of the wide use of thorotrast for
diagnostic purposes, no other case of endothelial sarcoma of the liver
was reported, even though thorotrast is retained by reticuloendothelial
cells. Hence, the possibility is considered that this tumor was spon-
taneous. All cells of the liver are relatively resistant to induction of
neoplasia. The observations of MacMahon and those made earlier by
Ross suggest that endothelial cells are the most susceptible elements of
this organ. Many radioactive heavy metals entering the body are
retained in the liver, notably those given in suspensions or colloids.
Tissue containing 1 per cent thorotrast by weight is estimated to receive
approximately 3000 rep in ten years (Evans, 1950a, b).

Sarcoma of the maxilla developing nine years after intravenous injec-
tion of mesothorium was described by Gricouroff et al. (1943), and a
solitary plasmocytoma of the humerus in a mesothorium worker by

1158

RADIATION BIOLOGY

Lumb (1950). One patient developed a carcinoma of the eyelid thirty-
five years after local injection of thorium dioxide (Rudolphi, 1950).

Maisin and Dupuis (1929) induced embryomas in chickens by intra-
venous injection of ionium in combination with injection of embryonic
extract into the pectoral muscle. Embryonic extract alone did not induce
tumors. The role of the radioactivity of ionium in inducing these
embryomas may be doubtful because its half life is 7.6 X 104 years.
Mottram (1935) induced mesoblastic tumors in fowls by exposure to
radium.

SKULL LESIONS DUE
TO RADIATION
OSTEITIS

PULMONARY METASTASES
FROM ORBITAL SARCOMA
BY WAY OF OPHTHAL-
MIC VEIN

RADIATION
OSTEITIS

PRIMARY OSTEOGENIC
SARCOMA OF ORBIT
(Anoplostic type)

SARCOMA OF PLEURA
BY CONTINUITY
FROM LUNGS

PRIMARY OSTEOGENIC
SARCOMA OF ILIUM
(Anoplostic type)

REGENERATIVE MAR-
ROW OF MEGALOBLAS-
TIC TYPE
( Panmyelosis ? )

Fig. 18-4. Multicentric osteosarcoma in a dial painter with secondary tumor deposits
in the lungs and foci of the precancerous radiation osteitis {after Martland, 1931).

The accidental induction of malignant bone tumors by radioactive
substances in luminous watch dial painters (Martland and Humphries,
1929, and Martland, 1931) (Figs. 18-4 to 18-6), and skin tumors following
therapeutic use of radon seeds (Evans, 1950b) will be described more
fully. N0rgaard (1939) described the development of fibrosarcoma in a
man following therapeutic intra-articular injection of radium chloride.

Artificial Radioactive Isotopes. Some radioactive isotopes are now
standard therapeutic agents while others are being used experimentally.
These include P32, I131, Au198, Mn52, Na24, Ga67, Sr89"90, and many others
(Siri, 1949). The doses employed in cancer therapy are in the carcino-
genic range, but so far no patient receiving such therapy has been known
to develop a neoplasm. As many as ten to thirty years may elapse
before tumors appear.

CARCINOGENESIS BY IONIZING RADIATIONS

1159

Experimental studies on the carcinogenicity of radioisotopes obtained
from fission products were begun by investigators of the Manhattan
Project. The following observations were made by Lisco et at. (1947):
Tumors of the colon followed ingestion of an insoluble Y91 compound by

Fig. 18-5. X-ray picture of the pelvis of a dial painter with primary osteosarcoma of
the pubic bone (after Martland, 1931).

Fig. 18-6. Radiation osteitis. In the bone marrow, areas of hyperplasia alternate
with areas of fibrosis. There is an increased osteoblastic activity (after Martland 1931).

rats. When Y91 was injected subcutaneously into mice as phosphate,
epidermoid carcinoma and fibrosarcoma were induced. When yttrium
phosphate was injected as a suspension intravenously, it induced osteo-
genic sarcoma in the long bones of rats. Plutonium as nitrate or citrate
injected subcutaneously into mice induced fibrosarcoma at the site of
injection and osteogenic tumors in distant bones. When plutonium was

1160

RADIATION BIOLOGY

implanted as metal subcutaneously (Lisco and Kisieleski, 1953) the
majority of the animals appeared to suffer no ill effects from the local
implants or from the absorbed plutonium. When plutonium was injected
as citrate or phosphate intramuscularly or intravenously into rats, it
induced osteogenic sarcoma. When injected intraperitoneally into
rabbits, it induced osteogenic sarcoma at distant sites. When adminis-
tered intravenously, it induced, in addition to bone sarcomas, massive

liver damage with secondary cirrho-
sis; whether this will be followed by
carcinoma has not been established.
Plutonium as citrate or nitrate
injected intratracheally or pluto-
nium oxide and cerium oxide pro-
duced as aerosols by burning,
inhaled by rats, caused squamous
and medullary carcinomas of the
lung (Lisco et al., 1947; Lisco and
Finkel, 1949).

Strontium-89 given intraperito-
neally induced in mice heman-
gioendothelioma of the femur and,
more commonly, osteogenic tumors
in different bones. Radium, stron-
tium, and the rare earth and
transuranic elements are all "bone
seekers" which means that from
the primary sites of injection they
are likely to be translocated to bones.
Large doses of P32 (Koletsky et
al., 1950), injected in rats either in
single or repeated doses, caused the development of osteogenic sarcomas
(Fig. 18-7) in many bones and squamous cell carcinomas in about 47 per
cent of the animals. Radiophosphorus, as well as many other fission
products, was deposited heavily in the skeleton. The carcinomas in
these rats originated in structures close to bone, as in the roof of the
mouth. The dose used in rats is estimated to be about twenty times the
usual therapeutic dose in man. When applied to the skin, the 0 radiation
of P32 caused multicentric tumors in every rat exposed (Raper et al., 1946;
Raper, 1947; Henshaw et al., 1947).

Fig. 18-7. Osteogenic sarcoma in a rat
tibia produced by P32 {after Koletsky et al.
1950).

NEUTRONS

Henshaw et al. (1947) studied the effect of fast neutrons from the Oak
Ridge reactor and noted a marked increase in the incidence of leukemia
and ovarian neoplasms. The incidence of leukemia following a single

CARCINOGENESIS BY IONIZING RADIATIONS 1161

exposure to fast neutrons was 22 to 32 per cent in groups of mice exposed
to 26 and 90 N as compared to 14 per cent in controls. The variations in
relation to dose seemed not significant. In parallel experiments they
studied the leukemia incidence following a single dose of 7 radiation and
found this to be 67 per cent in mice exposed to 500 r and 64 per cent in
mice exposed to 700 r.

The effects on neoplasia incidence of slow neutron exposure are being
studied currently in the biological tunnel of the Oak Ridge reactor.
Analyses about one year after exposure indicated a five- to tenfold
increase in leukemias in the dose ranges at or moderately below the LD50
dose. When compared on the basis of LD50, no preferential effect was
noted between slow neutrons and X radiation.

The chronic effects of neutrons from cyclotrons were studied by a few
investigators, but unfortunately either the animals were not observed
long enough or no distinction was made between leukemias and leukemoid
reactions. Leitch (1947) noted an increased incidence of mammary
tumors in rats exposed to cyclotron neutrons.

In exploring the possible role of cosmic rays in carcinogenesis, Figge
(1947, 1949) claimed that the potency of chemical carcinogens may be
altered by slight variations in the intensity of penetrating environmental
radiations. No data are available to indicate that cosmic radiation is
carcinogenic. No such effect was found by Franks and Meek (1950) in a
carefully controlled experiment.

ORGAN SUSCEPTIBILITY TO TUMOR FORMATION

It is probable that all cells capable of multiplication are susceptible to
the induction of neoplasia by irradiation. The incidence and sites of
neoplasia depend on the amount of energy absorbed by the cells, the
functional state and radiosensitivity of the cells, and other factors.
There are ample observations to indicate that hemopoietic cells, notably
lymphocytes, are highly radiosensitive, while macrophages (histiocytes,
reticuloendothelial cells) are highly resistant. Specialized cells, as osteo-
blasts, are highly radiosensitive; muscle, liver, and kidney cells are
more resistant. The most radiosensitive organ appears to be the
mouse ovary.

Studies published up to 1942 on the effects of irradiations on normal
tissues have been reviewed in a series of papers by Shields Warren and
associates (1942-1943) and the histological changes also by Bloom (1948).
It is difficult to compare the relative sensitivity of different normal cells
on the basis of total-body irradiation, since they are not in a comparable
state of the reproductive cycle or under identical physiological and
environmental influences. The differences in response of different types
of cells and of different cells of the same type to similar ionizing radiations
are truly remarkable (Lacassagne, 1936). As he phrased it:

1162 RADIATION BIOLOGY

Why does a ray passing through a certain region cause the death of one kind, e.g.,
the germinative cells of the gonads or certain cells of the lymphatic nodes, delayed
death of certain others, e.g., those of the epidermis, simple transient alterations
such as are noted in connective tissue, or no reaction at all, as in the case of nerve
cells. . . . Among apparently identical cells receiving equal doses of radiation,
some are killed, others show various lesions, still others are but slightly damaged
and are capable of recovery. This difference in radiation effects explains the
repair following irradiation, such as that of irradiated epidermis, or reversible
blood changes, or the reconstitution of the germinal epithelium. It explains
also the recurrences of cancer treated by irradiation. The traditional explanation
of this phenomenon is biological, viz., that the cells, in their life cycle pass through
stages of varying sensitivity corresponding to varying physiological conditions.

The variable radiation sensitivity of the cell can be explained, and a means of
establishing this sensitivity furnished, by the discontinuous nature of radiation.
The sensitivity exhibited by each cell type appears to be a function of two main
factors: (1) the histological structure, which determines the size of the various
zones corresponding to each lesion, and (2) the physicochemical composition,
which determines the number of quanta necessary to produce ionization sufficient
to cause disintegration of a sufficient number of constituent molecules of the
intracellular organelle in question.

These considerations apply to the induction of neoplasms at the site of
irradiation. Leblond et al. (1951) noted some correlation between the
incidence of spontaneous neoplasms of different organs and rate of repro-
duction of their cells. Organs with high mitotic activity appear radio-
sensitive, but this is not a general rule. A different mechanism is in
play when tumors arise at distant sites, as will be discussed.

LEUKEMIA

The increased incidence of leukemia among radiologists now seems to
be well established, and there are numerous experimental studies elucidat-
ing the conditions of leukemia induction in mice.

Leukemia among Radiologists. As early as 1911 Jagie et al. described
four cases of leukemia among radiologists and one in a radium worker and
stated that long-continued exposure to X radiation may cause this
disease. In 1912 Aubertin stated that in nineteen years he had seen five
cases of leukemia among radiologists, a then relatively small fraction of
physicians, and none among other physicians. In 1925 Emile-Weil and
Lacassagne reported on myeloid leukemia caused by handling radioactive
substances. These reports were soon amply confirmed. Henshaw and
Hawkins (1944) found that during the period of 1933-1942 leukemia was
1.7 times more common among American white physicians than among
the general male white population (Dublin and Spiegelman, 1947) and
cited similar evidence from England and Wales. Ulrich (1946) estimated
that leukemia was eight times more common among American radiolo-

CARCINOGENESIS BY IONIZING RADIATIONS 1163

gists than among other physicians; between 1935 and 1944, according to
him, 3.9 per cent of 205 radiologists died of leukemia as compared to 0.44
per cent of 34,626 physicians other than radiologists. According to
March (1944, 1950) the greatest excess in the mortality of physicians
was found from leukemia; the death rate from this disease was 175 per
cent of that for white males in the general population. The increased
incidence in leukemia among physicians other than radiologists was 133
per cent of that of the general population. The incidence of leukemia in
radiologists was over ten times as great when compared with other
physicians, and this difference according to March was statistically
significant.

Analyzing the conditions of irradiation under which leukemia developed
in radiologists and was induced in mice, the following differences are
noted: Chronic irradiation produces leukemia in mice only when young
animals are exposed over the entire body at a dose rate far exceeding
that now encountered by radiologists. Radioscopy involves exposure of
only part of the body and such exposures in mice are not leukemogenic.
Furthermore, radiologists are exposed to a relatively small average tissue
dose. This may explain why the increase of leukemia incidence among
radiologists is relatively slight.

Explosions of atomic bombs may cause leukemia predominantly by
virtue of 7 radiation of high energy, and the results in man (Folley et al.,
1952) and experimental animals (Upton et al., unpublished data) are
similar. In both, there is ample evidence that a massive instantaneous
single exposure increases the leukemia incidence. In Hiroshima and
Nagasaki the increase in incidence of leukemia during the past three years
is highly significant in subjects exposed at distances less than 2000 meters
and the magnitude of increase is inversely related to the distance from
the hypocenter, as indicated by the following very approximate figures:

Exposure Distance from Hypocenter, m Death Rate from Leukemia per 106 Living
Up to 1000 500

1000-1500 210

1500-2000 87

Over 2000 32

Leukemia Deaths per 104 Total Dead
In all Japan (1943-1949) 13

United States (1940) 39

The disease occurred mostly in the early and intermediate age groups and
the predominant types were acute myelocytic leukemias (Folley et al.,
1952).

Experimental Induction of Leukemia. Carefully controlled experi-
mental studies with mice exposed to radiations of an atomic bomb con-
firm the preceding observations. Ten months after exposure over 6 per
cent of a large surviving population of mice exposed to a dose above the

1164 RADIATION BIOLOGY

LD50 died of leukemia as compared to 0 per cent of the controls. Expo-
sures below that dose have caused only few leukemias thus far (Upton
et al., unpublished data).

While depression of hemopoiesis immediately after exposure to ionizing
irradiation is reversible, the liability of late changes of which leukemia is
most important appears irreversible. This varies greatly with species,
strains of animals, mode and dose of exposure, and is influenced by
several factors. In mice the smallest dose of X radiation which depresses
lymphopoiesis is about 25 r and the smallest dose which causes leukemia
is somewhat below 200 r. Rats are possibly susceptible to the induction
of leukemia by radiation (Metcalf and Inda, 1951) ; rabbits and guinea
pigs, and probably also dogs, are refractory, but no large-scale studies in
which animals were observed during their entire life span have been
reported in species other than these.

The leukemogenic potency of X radiation was first demonstrated
experimentally in mice by Krebs et al. (1930). Of 5500 mice exposed to
sublethal doses of X radiation 3.5 per thousand developed lymphoid
leukemia as compared to 0.6 per thousand of the controls. Furth and
Furth (1936) exposed large numbers of mice of three unrelated strains to
one or a few doses of 300-400 r of X radiation and found a sevenfold
increase in lymphoid leukemia, mainly of mediastinal type, and an eight-
fold increase of myeloid leukemia. Subsequent work by several investi-
gators disclosed the following: (1) Repeated, well-timed exposures are
more leukemogenic than single exposures. (2) The leukemia induction
rate is increased with the dose. (3) All strains of mice that were carefully
tested proved susceptible although there are marked strain and sex differ-
ences in susceptibility. (4) In no species other than the mouse, and
possibly the rat, has leukemia been induced experimentally, but adequate
follow-up of the exposed animals has been done only in mice, rats, rabbits,
and guinea pigs. (5) Gamma radiation of radium and fast and slow neu-
trons of the atomic reactor likewise increased the incidence of leukemia
in mice. (6) Leukemia induction by radioactive substances other than
P32 has not been reported in animals; it has been described, however, in
handlers of radioactive substances.

In most strains of mice spontaneous leukemias are thymic in origin.
Operative removal of this organ (Furth, 1946) or its involution brought
about by cortisone (Upton and Furth, unpublished data) diminish the
incidence of spontaneous leukemias or their induction by irradiation
(Kaplan, 1950b; Kaplan et al., 1951). Hueper (1934) exposed mice bear-
ing spontaneous mammary tumors to repeated doses of 30-80 r of X
radiation once a week over a period up to six weeks. His treatment
caused extensive myeloid hyperplasia with extensive myelopoiesis of the
spleen; the greatly increased incidence of leukemia in Hueper's experi-
ments suggested that irradiation caused a leukemic transformation

CARCINOGENESIS BY IONIZING RADIATIONS 1165

in these hyperplastic but nonmalignant myelopoietic and lympho-
poietic tissues. Conversely, removal of the adrenals increases suscepti-
bility to the induction of leukemia by X radiation (Kaplan et al., 1951).
Desoxycorticosterone is without effect. The inhibitory effect of cortisone
is still manifest when its administration is deferred until six weeks after
irradiation (Kaplan et al., 1951).

There are few experiments bearing on the pathogenesis of leukemia
induction by ionizing irradiation. Kaplan (1949) found that, in contrast
to whole-body exposure, local irradiation of mice failed to increase
consistently the incidence of lymphoid tumors. He concluded that the
induction of these tumors is not solely the result of a direct action of
radiation upon susceptible cells or their ancestors. In C57 black mice
irradiation of the upper half of the body yielded leukemia in 4 per cent
of the mice, lower half 2 per cent, whole body irradiation 64 per cent,
when a total of ten treatments were given in twelve consecutive weeks.
However, by alternating upper and lower body exposures within 24 hours
many lymphoid tumors were produced (Kaplan and Brown, 1951), sug-
gesting again that an indirect systemic mechanism is involved in leukemo-
genesis. Similarly, Lorenz and Eschenbrenner (unpublished data)
observed the development of lymphomas in mice of strain A irradiated
over the entire body with the thorax shielded, but the increase over con-
trols was somewhat less than after total-body irradiation and exposure of
the thymus alone did not increase the incidence. On the other hand,
Kaplan (1949) found that shielding the thymic region of C57 black mice
did not increase the incidence of leukemia and shielding the leg had a
protective effect. Similarly, direct irradiation of the thymic region with
the rest of the body shielded did not produce leukemias (Kaplan, 1949,
1951).

These observations led to the view that the induction of leukemia is by
some indirect mechanism. Prolonged depression of hemopoietic activity
may also be a factor in eliciting lymphoid tumors (Lorenz et al., 1953).
Kirschbaum and Mixer (1951) found that in estrogen-treated mice
thymic lymphosarcoma could be induced by irradiation of the entire
body except the thymus and concluded that the actual leukemogen, as
Kaplan and others believe, is a humoral substance. Whether the
humoral agents protecting the body from early radiation death will also
lessen the likelihood of leukemia development remains to be seen.

Little attention has been given thus far to the determination of the
types of experimentally induced leukemias. Some radioisotopes, as
Au198 colloids, are selectively deposited in macrophages and the leuke-
mogenic effect of such exposures remains to be studied. Aubertin (1931)
and Emile-Weil and Lacassagne (1925) reported an increase of myeloid
leukemias among radiologists but, in subsequent analyses, all types of
leukemias are considered together. In mice, lymphoid tumors induced

1166 RADIATION BIOLOGY

by irradiation are predominant. Furth and Furth (1936) mention also
an increase of myeloid leukemias, but this lacks confirmation. The type
of leukemia induced by irradiation may be influenced by the state of
the hemopoietic organs at the time of irradiations. This is suggested by
the observations of Hueper (1934).

The factor of age was studied by Kaplan (1948). The maximum
incidence of leukemia occurred in mice of strain A when irradiated at
one month (29.0 per cent), with a sharp decrease at two months (2.8 per
cent) and later (at 3 months 10.9 per cent, at 4 months 6.2 per cent, at
6 months 0 per cent). In mice irradiated at two weeks of age the
leukemia incidence was 14.3 per cent. The variability of values is prob-
ably due to the small number of animals used. These observations were
confirmed by him with C57 black mice.

The influence of age and dose rate on leukemia induction was investi-
gated by Brues et al. (1949). Throughout most of the life, the suscepti-
bility to irradiation-induced lymphoma approximately doubled at inter-
vals of 80 days. After 700 days of age no further increase in morbidity
rate was observed. Using different patterns of X-radiation treatment, it
was found that 400 r total-body X radiation is more effective if divided
over a 10-day period than if given either as a single dose or divided over
40 days. Increasing doses are more effective until a saturation value is
reached (Brues et al, 1949). There is also evidence for a threshold dose
or dose rate below which leukemia is not induced.

Lorenz et al. (1947), investigating the effect of chronic exposure to 7
radiation of radium, induced malignant lymphomas in mice at a rate that
was roughly dependent on the dose rate. The greater the dose rate,
the earlier the appearance of the tumors. Their results are illustrated by
Fig. 18-1. This shows that long-continued daily exposure of female
LAFi mice to 8.8 r daily (8 hours) to the 7 radiation of radium greatly
increased the incidence of leukemia and hastened its onset. The total
dose received by these animals over a period of twenty-three months was
5900 r. Daily exposure to 4.4 r had a similar but less marked effect,
while smaller doses (2.2 r daily or less) did not increase the incidence of
leukemia in comparison to untreated controls. The strain used had a
very high incidence of spontaneous leukemia appearing mainly late in

life.

Rats were exposed by Metcalf and Inda (1951) to doses of 0.1-10 r per
day during a period of two years. Their findings suggest that leukemia
may develop in this species as a result of such irradiations.

P32 has carcinogenic properties comparable to those of X radiation
(Brues et al, 1945). The rate of tumor production with both is roughly
proportional to the corresponding lethal doses. P32 is effective whether
given in a single dose or in monthly divided doses.

The carcinogenic effect of slow neutrons in the thermal column of the

CARCINOGENESIS BY IONIZING RADIATIONS 1167

Oak Ridge reactor is being studied currently. When analyzed six to
sixteen months after exposure, it was found that doses of 128 r of X
radiation and equivalent doses of slow neutrons increase the leukemia
incidence in mice about fourfold; 512 r and an equivalent dose of slow
neutrons, about sixfold. A preferential effect of neutrons as found for
cataract induction was not evident for leukemia induction. This slow
neutron flux has negligible quantities of fast neutrons, but it contains
much 7 radiation of high energy of about 5-6 r/minute (Darden et al,
1951); thus an LD50 exposure would include exposure to 400-480 r of y
radiation and would account for about half the biological effectiveness as
concerns neoplasia induction.

Earlier, Henshaw et al. (1947) compared the effects of fast neutrons and
X radiation. The incidence of malignant lymphoma following exposure
to a single dose of y radiation was 64 and 67 per cent in groups receiving
single doses of 700 and 500 r and 22-32 per cent in groups exposed to
26-90 n of fast neutrons, as compared to 14 per cent in the controls. The
r-n ratio of y radiation to fast neutrons for the different effects studied
varied from 8:1 to 35 : 1.

The production of reticulum cell sarcoma in a rat by administration of
thorotrast was described by Onufrio (1938). This tumor is not uncom-
mon in the rat so this report is inconclusive, as are our observations on
similar neoplasms in the only two mice surviving the administration of
large quantities of Au198 colloid.

ENDOCRINE ORGANS

The induction of neoplasia in endocrine organs by ionizing radiations is
of unusual interest for the following reasons: In the genesis of endocrine
tumors the specific (direct) irradiation effect is minor. This carcino-
genesis is reproducible in almost all mice in the two organs thus far
studied extensively (ovary and pituitary). The tumor incidence is not
dose-dependent in acute exposures; if a threshold dose is exceeded, almost
all animals develop the neoplasm. This threshold dose may be very
small, being below 32 r in the case of ovarian tumor induction by single
total-body exposure (Lorenz et al, 1947; Furth, 1949) or a total of 90 r
by long-continued y radiation (Lorenz et al, 1947) or, in the case of
pituitary tumors, many thousand rep to the thyroid (300 /ic of I131 per
mouse) (Gorbman, 1949). This dose may correspond to that required to
destroy the functional capacity of this organ. The importance of
hormonal imbalance in carcinogenesis has been reviewed by Gardner
(1948). The ovaries are much more susceptible to irradiation than the
thyroid gland. After sterilization of the mouse, subsequent irradiation
does not modify the sequence of the carcinogenic process. The radiation
effect can readily be counteracted or minimized by endocrine therapy;

1168 RADIATION BIOLOGY

this is proved in the case of ovarian tumors (Li and Gardner, 1949;
Gardner, 1950; Kaplan, 1950a) and pituitary growths (Furth, 1951,
unpublished data).

Gonads. Many studies of the early effects of radiations on the gonads
have been reviewed by Warren (1942-1943). It is uncertain whether
irradiation within the permissible dose would cause tumors of the testes
and whether localized massive irradiation of the testis would give rise to
specific tumors; on the other hand, the ovary of the mouse is highly sensi-
tive to neoplasia induction.

In a series of excellent papers, Brambell et al. (1927-1929) described the
changes in the ovary of the mouse for a period of approximately six
months after irradiation. The possibility that the regenerative changes
lead to neoplasia was then unsuspected. Irradiated mice may undergo
several pregnancies, yet develop ovarian neoplasms a year and a half
after irradiation (Furth, 1949; Deringer et al., 1953). The induction
period of the tumors is invariably long. While regenerative changes
begin, depending on total dose after four to six months, tumorlike
growths do not appear before about seven months; large tumors are
infrequent and metastases are rare.

In the first study describing the induction of ovarian tumors by X
radiation (Furth and Furth, 1936) mice of three different strains were
exposed to a single dose or repeated large doses of X radiation at the age
of approximately five to twelve weeks. Irradiation caused a fifteenfold
increase of ovarian tumors over that of controls. Following irradiation
of four- to six- weeks-old mice with single doses of 87, 175, or 350 r of X
radiation (Furth and Boon, 1943), ovarian tumors began to appear when
the mice were about eleven months of age. The frequency of these neo-
plasms increased with time and almost every mouse that lived seventeen
months developed a unilateral or bilateral ovarian growth, irrespective of
the dose of radiation.

The observation that total-body irradiation in female mice is fre-
quently, if not inevitably, followed by the development of ovarian tumors
has been described by numerous investigators (Bali and Furth, 1949).
Lorenz et al. (1947) pointed out that, in mice chronically exposed to 0.1 r
daily (8 hours), there is a significant increase in ovarian tumor incidence
over that of controls. The maximum accumulated dose was 164 r.
There is no significant decrease in mean survival time. Thus the ovarian
tumor-inducing dose is cumulative and the smallest dose rate inducing
ovarian tumors in mice is about 0.1 r daily, with an average total dose of
110 r.

Exposure of ovaries to X radiation causes the development of tumors
(Fig. 18-8) of different types. These neoplasms are multicentric, occur
frequently in both ovaries, and are of diverse histological type, each of
which can be isolated as a pure line by serial transplantation. Most

CARCINOGENESIS BY IONIZING RADIATIONS

1169

tumors arising in irradiated ovaries are complex (Bali and Furth, 1949).
Most transplantable neoplasms are of the granulosa cell type; a small
number are luteomas and tubular adenomas. All these types are fre-
quently present in the same irradiated ovary. Less commonly encoun-
tered neoplasms were hemangiomas and endotheliomas (two of which
resembled chorioepitheliomas) and sarcomas. Only two types of cells
thus far transplanted were found to be associated with hormone produc-
tion: granulosa cell tumors causing morphological changes indicative of
the production of estrogens, and luteomas with changes indicative of
progestin production. It is not known whether all small tumorlike
nodules in the ovary of X-irradiated mice are autonomous growths, but

Fig. 18-8. Unilateral ovarian tumor (lacteoma) in a mouse induced by total-body
exposure to X rays with hormonal hyperplasia of the uterine horns (after Furth and
Butterworth, 1936).

five types have been grafted on normal hosts : granulosa tumors, luteomas,
tubular adenomas, sarcomas, and angioendotheliomas. The granulosa
tumors occur in a wide range of morphological forms similar to those seen
in woman and simulate so many different types of neoplasms that their
identification on a morphological basis alone is often not possible. Their
common denominator is the ability to produce or initiate production of
estrogens and plethorins (a hypothetical substance that raises blood
volume— Sobel and Furth, 1948) although not all tumor-bearing hosts
show the effects of these substances.

Investigators who have attempted to describe the morphogenesis of
ovarian neoplasms have arrived at contradictory conclusions (Bali and
Furth, 1949). Some trace their origin to the covering germinal epi-
thelium, some to stromal cells (ovariocytes). The chief pacemakers of
the ovarian cycle are the pituitary gland and the ova. The latter gradu-

1170 RADIATION BIOLOGY

ally die after irradiation by even small doses, and a sequence of changes
ensues, leading to the development of neoplasms. Neither the morpho-
genesis nor the pathogenesis of this process is well understood. Some
endocrine disturbance is doubtless at play, but the term "endocrine
imbalance" merely covers our ignorance.

When a threshold dose is exceeded, the onset of ovarian tumors and
their course follow a similar pattern; increasing the dose of radiation
beyond the sterilization dose (Lorenz, 1950) does not shorten the incuba-
tion period. This is unlike the situation found with leukemia or bone
tumor induction by irradiation. The induced ovarian neoplasms are
always slow to grow, hardly interfere with the life span of the animals,
and seldom metastasize. These findings suggest that there is some
trigger mechanism causing an all-or-none response, and this is best
explained on the basis of hormonal derangements initiated by the deple-
tion of follicles of the normal ovary by irradiation (Gardner, 1950).
Neither the virus nor the mutation theory can adequately explain ovarian
tumorigenesis by irradiation.

Lick et at. (1949) have studied the pathogenesis of ovarian tumor induc-
tion. They found that X radiation did not induce tumors in an irradiated
ovary if the animal's second ovary remained unirradiated and functional.
Irradiation of a single ovary induced ovarian tumors only if the second
ovary was extirpated. Although a second functioning ovary inhibited
ovarian tumor development following unilateral contact irradiation,
bilateral contact irradiation resulted in tumor development. Irradiated
ovaries implanted intramuscularly into irradiated and nonirradiated
groups of spayed LAFi hybrid mice gave rise to many granulosa-cell
tumors, luteomas, and related neoplasms (Kaplan, 1950a). No such
tumors occurred when irradiated ovaries were implanted into nonirradi-
ated, nonovariectomized mice. In concluding that intact ovarian endo-
crine function inhibits the development of tumors in irradiated ovarian
grafts, Kaplan confirmed the observations of Lick et al. (1949) and sug-
gested that both a direct and an indirect mechanism may be involved in
the induction of ovarian tumors by irradiation.

The induction of ovarian tumors in mice by exposure to ionizing radia-
tions, even though it is of unusual interest, may represent a special case.
Thus far (fifteen years after the original observations were made), no
other species has been found to be susceptible to ovarian tumor induction
by irradiation. This carcinogenesis requires no more than a single
exposure, once the threshold is exceeded; it is not dose-dependent, has a
long tumor-development time (approximately one year), and occurs in
almost all exposed mice after about one and a half years.

Sterilization of women by X radiation was practiced some years ago.
Whether X irradiation has increased the frequency of carcinoma in the
reproductive organs of women has been the subject of much debate. The

CARCINOGENESIS BY IONIZING RADIATIONS 1171

available information is certainly not impressive. The assumed period of
latency was so short in many reported cases that many neoplasms may
have been present at the time of irradiation. However, the denial of the
theoretical possibility of such an event (Dehler, 1927) certainly lost its
wisdom in the light of later experimental findings on the induction of
neoplasms in the reproductive organs of animals. Nevertheless, the
clinical observations of Depenthal (1919), Bumm (1923), Vogt (1926),
and Dehler (1927) should be regarded as possibilities rather than proof
of the induction of neoplasm in reproductive organs of women (uterus,
ovary, vulva).

Pituitary Tumors. Large quantities of I131 (200-400 nc in mice) com-
pletely destroy the thyroid gland. Persistent lack of thyroid hormone
causes an over-stimulation of the pituitary gland and the hyperplasia of
the pituitary cells, which secrete thyrotropic hormones, gradually
terminates in neoplastic growth.

The original findings of Gorbman (1949) that the destruction of the
thyroid by I131 gives rise in mice to such growths have been fully con-
firmed, every mouse receiving doses of I131 large enough to destroy the
thyroid and surviving such treatment longer than thirteen months
developed pituitary tumors (Furth and Burnett, 1951). In line with this
interpretation are the few observations that long-continued administra-
tion of antithyroid compounds may cause the development of similar
tumors.

The pituitary growths induced by doses of I131 destructive to the
thyroid are readily transplantable in mice, the thyroid glands of which
have been similarly destroyed, but not in normal mice, with rare excep-
tions. Therefore, these growths are not fully autonomous but merely
conditioned neoplasms even though they may metastasize. Pituitary
enlargement after thyroid destruction by I131 can be prevented by admin-
tration of 0.15 per cent USP desiccated thyroid in diet (thyroxine) (Gold-
berg and Chaikoff, 1951b); similarly, thyroid hormone restrains the
growths of the grafted pituitary tumors. These tumors discharge thyro-
tropic and possibly gonadotropic hormones and, when grafted in young
mice, cause a tremendous cystic dilatation of the extrahepatic biliary
tract (Furth et al., 1952). While a study of these tumors is of great
interest in both endocrinology and oncology, it is not likely to become a
human problem. When patients with thyroid carcinoma are given
thyroid-destructive doses of I131, and the symptoms of hypothyroidism
become pronounced, thyroid hormone is administered, which counteracts
the stimulation of the pituitary gland.

Thyroid. Administration of I131 localizes predominantly in the thyroid
gland, and thus 300-400 /zc given to mice causes complete destruction of
the gland. Somewhat lower doses (200-300 /*c) destroy the gland, leav-
ing a few atypical acini which do not seem to respond to excessive amounts

1172 RADIATION BIOLOGY

of thyroid-stimulating hormone circulating in the blood. Some of these
thyroid remnants exhibit the morphological appearance of a precancerous
lesion. Doniach (1950) administered 32 /xc of I131 either alone or in
combination with methylthiouracil to rats, and thereby increased the
rate of formation of thyroid adenomas and, in one rat, a thyroid tumor
was observed. Similarly, Goldberg and Chaikoff (1951a) observed multi-
ple adenomas in two of ten rats that had been given a single injection of
400 juc of radioactive iodine.

In a later work, Goldberg and Chaikoff (1952) observed malignant
thyroid tumors in seven of twenty-five rats given a single injection of
400 juc of radioactive iodine. Metastases occurred in all seven rats, the
organs invaded being lung, adrenal, lymph nodes, skin, and bone.

Invasion of blood vessels, and even lung metastases, were seen by
Morris and Green (1951) following long-continued administration of
thiouracil to mice. Many of these tumors ultimately became autono-
mous in the sense that they grew in normal mice (Morris et al., 1951).

The very definition of malignancy is now controversial. It is no longer
possible to determine on the basis of gross or microscopic studies whether
such lesions, so readily induced in different endocrine organs by irradia-
tion, possess autonomy, or are nonneoplastic growths caused and main-
tained by derangement of the normal hormonal balance. Further
studies on the induction of neoplasia of the thyroid gland by I131 alone
or in combination with goitrogens are desirable. On the basis of related
experience, one might expect induction of thyroid tumors in a high per-
centage of the experimental animals by a dose below that which com-
pletely destroys this organ. The origin of autonomy in conditioned
growth has been demonstrated in several organs, including the thyroid
and the pituitary (Furth et al., 1952).

In man, Kindler (1943) described the development of thyroid carcinoma
and that of the hypopharynx following massive prolonged irradiation of
the neck. After discussing the literature, he concludes that "Uberdosier-
ung erzeugt mit Sichercheit Krebs."

Parathyroid. This appears to be resistant to radiation. No tumor has
ever been described in this organ, either after total-body or local irradia-
tion. When mice are radiothyroidectomized with I131, this organ must
also receive large quantities of ionizing radiations. It usually decreases
in size and often little of it is found in serial sections; yet neither neo-
plastic nor preneoplastic changes have been noted in this organ (Furth,
unpublished data).

MAMMARY GLAND

There are numerous reports on the induction of mammary tumors by
ionizing irradiations in several species of animals (mice, guinea pigs, and
rats). The induction of this tumor appears to take place by an indirect

CARCINOGENESIS BY IONIZING RADIATIONS 1173

mechanism; in several experiments estrogens seem to be clearly
implicated.

The relation of mammary tumors to irradiation has been reviewed by
Lorenz ct al. (1951). Acute massive whole-body roentgen irradiation of
mice reduced the over-all incidence of these tumors in comparison with
that of the controls, but the incidence of this neoplasm was high in those
irradiated mice which developed granulosa-cell tumors of the ovary.
These findings were attributed to the well-known decrease in estrogen
production following irradiation of the ovaries and excessive estrogen
production by the granulosa-cell tumors (Furth and Butterworth, 1936).
Lorenz ct al. (1951) exposed C3H mice for ten to fifteen months to doses
of y radiation of 0.044-4.4 r daily. Although the females became sterile,
this type of chronic exposure had no influence upon the incidence of
mammary tumors.

In another series of experiments, Lorenz ct al. (1951) irradiated mature
LAFi mice with doses of y radiation ranging from 0.11 to 8.8 r daily
(8 hours) during the entire life span of the animals. LAFi mice which
do not carry the milk agent have a low spontaneous incidence of mam-
mary tumors. All experimental groups of female mice, with the excep-
tion of that exposed to 0.11 r daily, showed an increased incidence of
mammary carcinoma ranging from 4-14 per cent. Moreover, in all but
the 8.8-r group irradiated during the entire life span, sarcomas of the
mammary gland developed, the incidence ranging from 13-25 per cent.
Most breast tumors were associated with granulosa-cell tumors of the
ovary.

In their most recent paper, Lorenz ct al. (1951) describe the effects of
chronic massive y irradiation on the mammary tumor incidence in the
milk factor-free C3Hb mice. Forty-seven per cent of the females
developed mammary carcinomas and sarcomas, and 88 per cent, ovarian
tumors. These workers exclude the possibility that direct irradiation
alone was responsible for the production of these tumors and postulate a
combination of factors such as hormonal stimulation, direct effect on the
mammary gland, and other unknown indirect effects of systemic origin.
It is noteworthy that the mammary sarcomas like the mammary carci-
nomas are associated with granulosa-cell tumors. Histologic evidence of
estrogen secretion is often lacking in hosts bearing primary tumors, but is
invariably present in the subpassages of such tumors in young hosts.
Obviously, atrophic target organs may not respond to the hormones.

It is noteworthy that estrogen induces carcinomas in mice and fibro-
adenomas in rats, indicating that both epithelial and connective tissue
elements of the breast are under endocrine influence. This can also be
demonstrated by hormonal stimulation of grafted fibroadenomas. Of
twenty-eight rats living longer than 150 days after repeated doses of
cyclotron neutrons, eleven had malignant tumors of which seven were

1174 RADIATION BIOLOGY

mammary carcinomas (Leitch, 1947). The incidence of benign mammary
tumors following X irradiation (Metcalf and Inda, 1951) was 10.8-34.6
per cent in female rats exposed to 0.1-10 r daily during a period of two
years and 8.3 per cent in the controls. There was a direct relation
between the daily dose of radiation and the number of benign tumors,
the smallest increase occurring among rats exposed to 0.1 r and the great-
est among those exposed to 10 r. Ovarian tumors were not seen in rats.
It is not likely that in the same species X radiation would induce benign
tumors, and neutrons, malignant tumors; the discrepancy is probably due
to differences in diagnoses.

In guinea pigs of both sexes chronically exposed to 1.1 r for 8 hours
daily, mammary carcinomas occurred three to five times as often as
among the controls (Lorenz, 1950). These tumors were likewise not
associated with ovarian neoplasms. Thus the pathogenesis of the induc-
tion of mammary tumors in rats and guinea pigs requires an explanation.

Those who practice "prophylactic" irradiation to prevent recurrence of
tumors after their surgical removal may find experimental support in the
work of Owen and Williams (1940), who exposed C3H mice, which have
a high spontaneous incidence of breast carcinoma, to 100-400 r at approxi-
mately four months of age. They observed a decrease of cancer incidence
from 65 per cent in the controls to 11-39 per cent in the irradiated mice,
the decrease in cancer incidence being a function of the dose. These
experiments are subject to different interpretations, and arguments can
be presented in favor of both indirect (hormonal) and local mechanisms
causing this change. Unfortunately, the animals were sacrificed too
soon. An increase in mammary tumor incidence occurs after a longer
period of time, when in the ovary the proliferation of estrogen-secreting
granulosa cells supersedes the degenerative changes (Furth and Furth,
1936).

UTERUS

Chronic irradiation significantly increased the incidence of uterine
carcinomas in rabbits chronically exposed to 7 radiation of radium
(Lorenz, 1950) ; the higher the dose rate, the earlier the tumor development
time. The tumors did not metastasize in the controls but did so in most
experimental animals, and it is possible that chronic irradiation may have
enhanced the dissemination process. All but one of twelve rabbits
exposed to 1.1-8.8 r daily developed a uterine carcinoma and seven of
these had metastases. Two of six controls had similar tumors but none
metastasized. None of three rabbits exposed to 0.11 r daily had tumors
because they died early of intercurrent disease. Kaplan and Murphy
(1949) described in mice a transplanted mammary tumor, which usually
does not metastasize but will do so when irradiated with doses insufficient
to destroy the tumor. Similarly, one of us (Furth, 1935) found that a

CARCINOGENESIS BY IONIZING RADIATIONS 1175

transplantable leukemic tumor which tends to remain localized in normal
hosts at the site of injection will disseminate in irradiated hosts, causing
generalized leukemia.

SKIN

This organ is more resistant to induction of neoplasia by irradiation
than most internal organs. Single or repeated exposures to X or 7 radia-
tion which are harmless to the skin may cause neoplasms of internal
organs; leukemia in man; leukemia, ovarian, mammary, and lung tumors
in mice; mammary tumors in guinea pigs; and uterine tumors in rabbits.
Nevertheless cutaneous tumors were the first noted in man and repro-
duced in animals, mainly because of the relatively soft type of X radiation
used earlier and due to lack of adequate filtration. While penetrating
radiations, irrespective of type, produce internal tumors and leukemias,
the carcinogenic effect of nonpenetrating radiations is limited to the
skin because this is the site of greatest absorption (see also Blum, Vol.
II, this series, in preparation).

During the first decades following the discovery of X radiation, radia-
tion cancer of the skin occurred among those professionally exposed, as
has been described, but in recent years therapeutic exposure has been the
more common cause (Saunders and Montgomery, 1938). Self-produced
radiodermatitis has not vanished, however. In one large hospital alone,
115 physicians were treated for radiodermatitis, and 39 for skin cancers.
Carelessness and ignorance are the two main causes of these preventable
conditions (Leddy and Rigos, 1941). Skin tumors do not develop follow-
ing irradiation without an attendant reversible inflammation. Chronic
radiodermatitis is encountered particularly as a result of the injudicious
irradiation of various benign dermatoses. The principle that the more
extensive the injury, the more likely the superimposition of cancer, is well
established. Epitheliomas apparently develop with equal frequency
from either keratoses or ulcerations (Saunders and Montgomery, 1938).

In man the neoplasms arising in the epidermis usually commence with
a warty growth (papilloma) and terminate in squamous-cell carcinoma.
Ulceration usually but not invariably precedes the development of
■carcinoma. Rarely, tumors (sarcomas) arise in the connective tissue.
The first experimental tumors induced in rats were sarcomas, in the
rabbit carcinomas, and this may be due to differences in the texture and
thickness of the skin. The studies of Wolbach (1909) on the early histo-
logical changes in skin leading to carcinoma are now classical.

The earliest changes recognized by Wolbach were in the collagen. The
most conspicuous and constant change in connective tissue is rarefaction
immediately beneath the epidermis and a greater density in deeper layers
(Saunders and Montgomery, 1938). There is homogenization of the
collagen with formation of dense sclerotic areas taking on a bluish color
with hematoxylin and eosin stain. Wolbach found degenerative changes

1176 RADIATION BIOLOGY

in smooth muscle, obliteration of capillaries by proliferation of the endo-
thelium, and telangiectasia of other capillaries. The obliterative changes
in the veins and arteries were manifested chiefly by a great increase in
the connective tissue beneath the endothelium and a marked thickening
of the media. Hypertrophy of the epidermis was a constant finding.
Complete absence of hair follicles and of sebaceous and coil glands was
the rule in lesions of long duration. In no case was there evidence of
proliferation of any of the dermal appendages. Coil glands were often
found in regions in which there was a total absence of hair follicles and
sebaceous glands.

Saunders and Montgomery (1938) described the characteristic histo-
pathological picture of radiodermatitis as follows: The epidermis is
hyperkeratotic and acanthotic, and usually there is an associated increase
in the stratum granulosum. Necrosis and ulceration of the epidermis are
frequently encountered. The formation of abscesses and spaces (Liicken)
in the epidermis are seen infrequently. Destruction of the elastic tissues
occurs in severe cases. Sometimes a few fine fibers of newly formed
elastic tissue can be demonstrated. There is new formation of capil-
laries arising from thickened vessels in the upper cutis. The larger
vessels show varying degrees of thickening of the adventitia and media
and proliferation of the intima to the point of partial or complete occlu-
sion. The infiltrate in the cutis is not consistent or characteristic, poly-
morphonuclear leukocytes predominating in necrotic connective tissue in
areas of ulceration, and lymphocytes predominating elsewhere. The
sebaceous glands are almost invariably destroyed. Next, and depending
on the severity of the radio 'ermatitis, the hair follicles become involved
and, in the case of third-degree injuries, the sweat glands are usually
atrophic or completely missing. Hyperpigmentation, a feature of acute
radiodermatitis, is not evident, very little melanin being demonstrable
microscopically.

All epitheliomas originated in the epidermis and none in the hair
follicles or sweat ducts. Most epitheliomas studied show phenomena of
individual cell keratinization ; many showed giant epithelial cells, repre-
senting amitotic cell division. Roentgen epitheliomas simulate and even
duplicate the histologic picture of epitheliomas arising from senile and
arsenic keratoses, and they tend to begin as epitheliomas in situ, with
the various phenomena of individual cell keratinization.

Gamma radiation, from radon seeds, was widely used for the removal
of hemangiomas. In a review of all the case histories of a large metro-
politan tumor clinic, four cases have been found in which late radiation
dermatitis or a skin carcinoma followed in thirteen to sixteen years. Two
patients received much larger doses (about 1 10,000 r) than did the radium
poisoning cases, yet the tumor development time is comparable (Evans,
see Brues, 1951). The tumors so induced arise either in the epidermis or
subjacent connective tissue.

CARCINOGENESIS BY IONIZING RADIATIONS 1177

The carcinogenicity of a or 0 emitters can be tested conveniently by
direct introduction into the subcutaneous tissue. The tumors induced
arise either in the epidermis or in subjacent connective tissue. Andervont
and Shimkin (1940) induced subcutaneous tumors by injection of col-
loidal thorium dioxide and Lisco ct al. (1947) by plutonium and colloidal
yttrium phosphate. In the former experiment the effect of metallic
thorium has not been excluded. Lacassagne and Rudali (1942) caused
the regression of papillomas by irradiation in five rabbits, but in one
rabbit a new tumor, histologically a rhabdomyosarcoma, developed at
the site of the originally benign growth twenty-eight months after the
irradiation. A similar sequence of events has been observed in man.

Beta radiations from P3'2 are highly carcinogenic to the skin when
applied directly. The energy absorption from P32 is limited almost
entirely to the skin (Raper ct al, 1946, and Raper, 1947). In experiments
of Raper ct al, all rats receiving single doses of 4000-5000 rep developed
skin tumors (see also Henshaw et al, 1946, 1947) and the number of loci
of tumors arising in some rabbits exceeded fifty. The lethal dose of
external 0 radiation varied with the size of the animal, being approxi-
mately 4500 rep for mice and 7000 for rats.

LUNG

Lung Tumors among Miners of Schneeberg and Jachymov. There is no
doubt that ionizing radiations will induce lung tumors in experimental
animals, but it is still controversial whether radiations emitted by radon,
either inhaled as radon-contaminated air or present in the expired air
from radium deposits in the body, will induce lung tumors in man. Many
complicating factors make an evaluation of the problem difficult (Lorenz,
1944). With the extension of mining activity of radioactive deposits
throughout the world, exposure to inhalation of radioactive dust and gases
will constitute an increasing hazard, and this problem will therefore be
fully reviewed.

The main information on the induction of lung tumors in man comes
from the study of this disease in the miners of the uranium mines of
Schneeberg and Jachymov (Joachimsthal) . The history of a strange
disease called Bergkrankheit among the miners of Schneeberg goes back to
the sixteenth century (Sikl, 1950). However, not until 1879 was it
recognized (Haerting and Hesse) that this disease originated in the lungs
and that about 75 per cent of all deaths of the miners was due to a neo-
plasm diagnosed erroneously as lymphosarcoma. These findings were
confirmed by Arnstein (1913) who corrected the diagnosis to squamous-
cell carcinoma and attributed 40 per cent of all deaths of miners during
the period of 1875 to 1912 to this tumor. The incidence of pulmonary
tumors in miners of both Schneeberg and Jachymov was reviewed by
Peller in 1939, who gave the mortality statistics contained in the accom-

1178

RADIATION BIOLOGY

panying tabulation. More than 87 per cent of primary tumors in these
miners originated in the lung.

Cancer Mortality per 1000

Jdchymov
(1929-1938)

Schneeberg

Vienna males,
15-79 years
(1932-1936)

(1895-1897)

(1895-1912)

Lung cancer

Cancer of other organs

9.8 ± 1.5
0.7 ± 0.4

12.7 + 1
2.4 ± 0.4

16.5 + 1.5
2.1 ± 0.6

0.34
2.1

Early investigators attributed the high incidence of pulmonary tumors
to inhalation of arsenic dust. Besides pitchblende, the mines contain
silver, cobalt, arsenic, and nickel. Silicosis is as frequent as carcinoma,
but no relationship could be established between the two diseases. The
average time spent in mines before manifestations of carcinoma is thirteen
to seventeen years. The sites in the lung, the types and the biological
behavior of carcinoma are identical with those occurring in this organ
throughout the world (Behounek et al., 1937; Sikl, 1950).

The literature on this miner's disease is too voluminous to be fully
reviewed. After the discovery of the carcinogenic properties of radio-
active substances, radon was blamed for the high incidence of lung cancer.
In a detailed investigation, Rajewsky et al. (1942) found that the radon
content of air in the mines of Schneeberg and Jachymov varied from
7 X 10~12 to 7 X 10~9 curie per liter. Measurements of the radioactive
content of lungs of miners gave radium equivalent values not different
from that of non-miners. Rajewsky et al. found an average radon con-
tent of the air of the mines to be 3 X 10~9 curie per liter and assumed a
permissible dose of 1 X 10-8 to 1 X 10~9 curie per liter of air and con-
cluded that radon is possibly one of the causes of the lung cancer of the
miners.

A similar permissible dose was established by Read and Mottram
(1939). Evans and Goodman (1940) arrived at a daily dose of 1 X 10~n
curie per liter as follows: 1 jug of radium stored in the body will produce
1.1 X 10_u curie of exhaled radon per liter. Three dial painters devel-
oped carcinoma of the ethmoids and antrum (Martland, personal com-
munication). One of these had 2 ng of stored radium in the body and
exhaled 2 X 10-11 curie per liter. Typical air measurements in these
mines show ten to two hundred times this value. Krebs et al. (1930)
measured the body content of radium of eighteen persons who died with
no known exposure to radium and found a range of less than 1 X 10~9 to
4 X 10-8 radium equivalent. Hursh and Gates (1950) made similar
measurements and arrived at one hundred times smaller values, which

CARCINOGENESIS BY IONIZING RADIATIONS 1179

are 50,000 times lower than those that were found in people with bone
sarcoma induced by radium.

Assuming an average radon content of 3 X 10~9 curie per liter of mine
air, Evans and Goodman (1940) estimated an hourly exposure of dry
lung to be equivalent to 0.17 erg per gram, and the corresponding daily
dose on the upper bronchial epithelium of the order of magnitude of 0.1
rep per day. Assuming seventeen years as an average tumor induction
time, the total dose would be 600 rep (estimated to equal 3000 rem; Evans,
see Brues, 1951). The dose effective in initiating the tumor would
obviously be much smaller. It is not known whether a particles are
more effective than X radiation for the same total dose. On the basis of
blood analyses Woldrich (1931) concluded that cancer of the bronchi is
caused by inhalation of radium emanation.

Induction of lung tumors in mice requires either a comparatively
massive single dose or considerably higher chronic doses, as will be
detailed. In comparison to the mouse, man has a low incidence of spon-
taneous lung tumors; most tumors in mice are benign, while most of
those in man are malignant. In chronic exposures of man to radon in
the Schneeberg and Jachymov mines much smaller total doses are
involved than are necessary to induce lung tumors in mice. In breathing
air containing radon, the tissue dose of a radiation delivered to the
epithelium is, however, greatest in the larger bronchi where the primary
neoplasms are frequently located (Evans, see Brues, 1951). Radon gas
which emits a radiation becomes an atom of the solid radioactive sub-
stance RaA, decays into a series of radioactive elements of varying half
lives, which may adhere to the bronchial epithelium. Among other
factors which may contribute to the induction of these human tumors are
pneumoconiosis produced by the mine dust, and the cobalt and arsenic
content of the dust. Less likely is a hereditary susceptibility due to
inbreeding, mentioned by several investigators, or the carcinogenicity of
nonradioactive cobalt, chromium, and arsenic (see also Schinz and
Uehlinger, 1942; Hueper, personal communication).

Abrahamson et al. (1950) described the development of bilateral
alveolar lung carcinoma in a patient who, sixteen years previously, had
received an intravenous injection of 75 cc of thorotrast. The liver and
spleen of the patient still contained much radioactivity at time of death.
The multifocal origin of the lung tumors in this patient is cited as evi-
dence for their induction by irradiation.

Experimental Induction of Lung Tumors in Radioactive Mines. This has
been carried out either on a very small scale or without adequate con-
trols. Dohnert (1938) exposed mice in the Schneeberg mines at places
where the miners worked, but his uncontrolled experiments were incon-
clusive. Hereck (1939) reported on histologic changes in 110 mice
similarly exposed, of which five had adenomas of the lungs. Unfortu-

1180 RADIATION BIOLOGY

nately, no controls were observed. Lorenz (1944) injected eleven mice of
a lung tumor-susceptible stock four times at weekly intervals with an
aqueous radon solution totaling 1.2 mc. Some animals lived up to eight
and one-half months, but no increase in lung tumor incidence was
observed in the injected animals over that of controls. Rajewsky et al.
(1943) exposed mice continuously to various doses of radon in an emana-
torium. Mice exposed to 1.16 X 10~6 curie per liter lived from 161 to
453 days. Of the twelve experimental animals, ten had adenomas of the
lung, one an adenocarcinoma originating from a small bronchus, while
only one of the control animals had an adenoma.

Thus it is questionable whether radon can induce lung tumors in
experimental animals. The work of Rajewsky et al. suggests this, but a
problem as important as this deserves a sounder experimental foundation.

Experimental Induction of Lung Tumors by X or Gamma Radiation.
This has been carried out on a much larger scale. The induction rate of
lung tumors in mice by total-body irradiation is very low and the tumor
development time is long.

Furth and Furth (1936) gave evidence that massive doses of X radia-
tion given to three strains of lung tumor-susceptible mice induces lung
tumors. The increase in lung tumor incidence observed in females was
slight but statistically significant. Lorenz et al. (1946) exposed mice to
chronic y radiation giving 8.8 r daily (8 hours) and a total of approxi-
mately 2400 r over a period of ten months and found an increase in lung
tumor incidence of 50 per cent over controls. In later experiments,
Lorenz (unpublished data) has shown that the lung tumors are induced
by a direct action of the radiation upon the lungs. The lung tumors
induced in mice were alveolar in origin and benign, as were those in the
controls.

Lisco and Finkel (1949) induced bronchiogenic carcinomas in rats by
inhalation of an unknown quantity of radioactive cerium oxide; this
compound covers the bronchi tenaciously.

Gorbman (1949) described the development of tumors of the trachea in
mice that had been injected with large doses of I131. This was not seen
by one of us in more than thirty mice of the same strain similarly radio-
thyroidectomized by I131 and observed for over a year. The changes
seen in the trachea were destructive and proliferative but not neoplastic.
Cancer of the larynx was described in a woman whose thyroid received
prolonged X radiation (Jacques, 1935).

BONE

The development of a malignant tumor in a normal bone or in a benign
tumor that had been exposed to heavy doses of X or y radiation was first
described by Beck in 1922, and in 1945 Hatcher could collect from the

CARCINOGENESIS BY IONIZING RADIATIONS

1181

literature twenty-four cases (Fig. 18-9). In all except six the irradiation
was given for chronic joint infection. In three cases bone sarcoma
resulted from irradiation given for the treatment of other tumors. In all
but one case a large amount of radiation was administered in fractional
doses over a long period. The median interval between irradiation and
recognition of sarcoma was six years; the shortest interval was three
years; the longest, eleven years. Chondrosarcoma occurred more fre-
quently among irradiation-produced sarcomas than among other spon-
taneous bone tumors. The degree of bone formation in the tumors was
variable; those in which none was found were labeled as polymorphous
or spindle-celled sarcomas.

mm

f,

"*S\

Yj^ - ;^Hs^^

r

, ^" , aAljjS;^.

■ "ifcMi^'

iHEfc

<" '* -J

jtnpMjMIM

«

Fig. 18-9. Osteogenic sarcoma induced in man by X rays (after Hatcher, 1945).

Cahan et at. (1948) collected from the literature seventeen cases of
bone sarcomas following radium or roentgen radiation therapy reported
until 1948 (exclusive of Mainland's cases of radium dial painters) and
described eleven additional cases observed by them. The first thirteen
cases noted between 1922 and 1937 occurred in patients receiving thera-
peutic irradiation for bone tuberculosis. The patients were eleven to
forty years old at the onset of the neoplasm. Three to ten years elapsed
between irradiation and the discovery of the tumor. The bone sarcomas
were variously described as spindle-celled, pleomorphic, giant-celled,
chondro-fibro-osteosarcoma, and osteo-chondro-myxosarcoma. The sub-
sequent four cases described between 1939-45 occurred in patients
twenty-three to fifty-six years old receiving treatment for chronic
arthritis, benign giant-cell tumor, chondroblastoma, or as a "prophy-
lactic" measure following removal of carcinoma of the breast. It was
subsequently pointed out by several investigators that the irradiation of
benign giant-cell tumors is hazardous, as it is likely to cause a malignant
transformation of this neoplasm. Jaffe and Selin (1951) noted an appar-
ent increase in the number of malignant metastasizing giant-cell tumors
since the beginning of the era of radiation therapy of this tumor.

The eleven cases of bone tumor following X irradiation, well studied

1182 RADIATION BIOLOGY

by Cahan et al. (1948), occurred in patients from nine to fifty-nine years
of age. Some were treated for nonmalignant bone disease, such as
ossifying fibroma, bone cyst, osteoid osteoma, fibrous dysplasia, and
benign giant-cell tumor, or for nonosseous malignant tumor, as retino-
blastoma, or for an inflammatory process, as "cloudiness" of antrum.
This recalls earlier observations of bone tumor induction by X-radiation
treatment given for joint tuberculosis and the experimental work of
Lacassagne (1936), who induced varied neoplasms by irradiation of
chronic inflammatory tissue. The estimated tumor dose in the patients
of Cahan et al. varied between approximately 1550 to 16,000 r and the
interval between the last irradiation and onset of bone tumor varied
from six to twenty-two years. To one of the eleven patients, the X-
radiation treatment was administered to the site of a bone fracture follow-
ing radical mastectomy. It seems probable that osteoblastic activity at
the time of irradiation facilitated the induction of these neoplasms ; osteo-
blasts are highly sensitive to irradiation.

Most internal tumors in man exposed to either X radiation or radio-
active substances arose in bones. Auerbach et al. (1951) reported an
extraskeletal osteogenic sarcoma originating in soft tissues of the back.
This tumor arose directly beneath the skin in an area that four years
previously had received a skin dose of 4000 r of X radiation delivered in
ten exposures over a period of 70 days. The rare occurrence of extra-
osseous bone tumors following irradiation can be explained by osteogenic
metaplasia at sites of chronic inflammation.

The characteristic sequence of changes in bone following irradiation
and preceding the carcinogenic transformation has been described by
Ewing (1926), who named this change "radiation osteitis." This is
essentially a degenerative change with necrosis and reactive atypical new
bone formation and is similar following all types of irradiations. (For
recent references, see Koletsky et al., 1950.)

Bone Tumors in Radium Dial Painters. An industrial hazard occur-
ring in New Jersey, well followed by Martland (1931), claimed forty-one
victims during a period of some twenty years. Between 1916 and 1925
dial painters moistened, by licking, brushes which had been dipped into
the luminous compound containing radium and mesothorium. This
material ingested over many years lodged in the bones, causing severe
anemia and bone tumors. Death occurred within four to six years after
ingestion ceased. Autopsy revealed the outstanding changes were
necrosis of the jaw and aplastic anemia. The bone changes first noted
were those of radiation osteitis (Martland et al., 1925). Later (1931),
bone tumors developed in many foci, and death was caused from metas-
tases of one of these growths. At autopsy, Martland found other primary
bone tumors and transitional changes from radiation osteitis to malignant
growth. Thus far (Martland, 1951, personal communication), fourteen

CARCINOGENESIS BY IONIZING RADIATIONS 1183

girls had osteogenic sarcomas, all of whom died before 1930; four are still
known to be alive, suffering from a crippling bone lesion, probably radia-
tion osteitis with pathological fractures; and three died from epidermoid
carcinoma that started in the mucosa of the accessory sinuses of the head.
Since this type of carcinoma is rare and no other carcinoma was found in
this small group, their relation to the ingested radioactive materials
seems highly probable. Mainland's observations gave impetus to the
study of experimental induction of bone tumors, and to establish factors
governing the induction of this tumor, such as the dose levels of tumor
induction, permissible maximum exposure to radiation, and methods to
increase the elimination of radioactive substances from the body.

In persons with chronic radium poisoning who have developed osteo-
genic sarcomas, the radium content varied from a few to over 20 ^g.
Analysis of the radium content of bone of a patient with a fibrosarcoma
made after approximately fifteen years of exposure showed an average of
1.5 X 10-9 g of radium per g of dry bone, delivering approximately 1 rep
per day per g of bone (Evans et al., 1944; Evans, 1950a). Average dose
values are of little significance unless the distribution pattern is known to
be uniform. This is not the case, however, since the radium is distributed
at discrete points throughout the bone.

Recent investigations of Evans (1950a) showed that persons who
had received radium only and who contained from 0.3 to 22 ^g of stored
radium were free of bone tumors up to thirty-four years. A clinical pic-
ture similar to that of the luminous dial painters was produced in these
patients by approximately five to ten times as much radioactive material.
The compound ingested by the dial painters contained mesothorium, an a
emitter with a half life of 6.7 years. When Martland's cases were
investigated, mesothorium had already decayed many half lives and
only the stored radium was determined. Mesothorium may have been
involved in all cases of irradiation injury in which the stored radium was
less than 5 ^g- Accordingly, the maximum permissible value of 0.1 ng
of radium may have a safety factor of 50. The permissible doses of the
various elements given were established before these findings were known.

Experimental Production of Bone Tumors. The first bone tumors in
man and experimental animals were induced inadvertently. Lacassagne
and Vinzent (1929), irradiating chronic inflammatory lesions of rabbits,
produced an osteosarcoma and a periosteal sarcoma. Lacassagne and
Nyka (1937), attempting to induce tumors of the hypophysis with radon,
caused the development of bone tumors of the sella turcica. The
sequence of changes noted has been necrosis, revascularization, callus
formation, and cancerization. The numerous studies made subse-
quently, including those with fission products, have been detailed in the
section on corpuscular irradiations. It is evident from the observations
described that osteoblasts are highly susceptible to neoplasia induction.

1184 RADIATION BIOLOGY

Marrow cells adjacent to radioactive bone received large doses of radia-
tion and humans so exposed frequently developed anemia, yet only a
few myeloid leukemias have been reported among them.

LIVER

Malignant liver-cell tumors resulting from exposure to ionizing radia-
tions have not been described in man or animals, but there is some evi-
dence that massive doses of ionizing radiations may cause endothelial
sarcomas in man and benign hepatomas in mice.

Ross (1936) described an illustrative example of the consequence of
prolonged exposure to 7 radiation in a localized area of human liver, in
which a hemangioma developed subjacent to a radium needle. Metas-
tases were present in the lung and bone marrow. The origin of the tumor
was traced to branches of the portal vein. Similarly, MacMahon et al.
(1947) described an endothelial cell sarcoma of the liver following thoro-
trast injections.

Lorenz (1950, and unpublished data) noted an increased incidence of
hepatomas in mice following chronic 7 irradiation. The direct relation
of these benign nodules to irradiation remains to be demonstrated. Au198
colloid had been given to man and animals, delivering to this organ many
times the LD50 total-body dose, but thus far no local tumors have been
produced. Liver damage with some cirrhosis was seen following Au198
therapy (Hahn et al., 1951). The observations thus far are sketchy, how-
ever. It seems that, in man, the endothelial cells of the liver are sus-
ceptible to neoplasia induction by irradiation while liver cells are resist-
ant. Chronic exposure studies with Au198 colloid are most desirable,
since this colloid selectively localizes in the Kupffer cells, exposing the
liver to thousands of rep without causing early death of the animal. The
above observations suggest the possibility of neoplasia induction from its
use. Radiogold colloid is currently used in the experimental therapy of
human cancers. (Concerning cirrhosis in animals receiving thorium, see
Hugenin et al., 1931.)

Direct introduction of radium into the gallbladder will cause carcinoma
of this organ (Biltris, 1933).

GASTROINTESTINAL TRACT

Lisco et al. (1947) produced carcinoma of the colon by feeding rats with
radioactive yttrium. These tumors were associated with polypoid hyper-
plasia of the colon and ulcerative colitis. The pathogenesis of these
tumors deserves further study, as little is known about the etiology of the
common tumors of the large intestine of man, and such tumors cannot be
easily induced by chemical carcinogens.

CARCINOGENESIS BY IONIZING RADIATIONS. 1185

BRAIN

This organ is resistant to irradiation and thus far no human or experi-
mental brain tumor caused by ionizing radiation is on record. Glia cells
of the brain are susceptible to chemical carcinogens, and one might expect
the formation of gliomas in brains exposed to large doses of ionizing radia-
tion. Yet thus far only sarcomas of the connective tissue and bone
tumors have resulted from such exposures (Jentzer, 1937; Lacassagne and
Nyka, 1937).

Wilson et al. (1951) exposed rat embryos to X radiation on the ninth
day of gestation and noted the development of discrete tumorlike growths
in and around the brain and related these directly to X irradiation. They
first appeared on the second day after irradiation and thereafter exhibited
varying capacities for growth and differentiation. Some grew for 1 or 2
days, then disappeared as a result of dispersal of the cells; others grew
rapidly until the fifth or sixth postirradiation day, then became atrophic;
and still others continued to grow slowly until the seventh or eighth day,
then became static or underwent atrophy and regression. It seems to us
that these developmental anomalies should not be designated as
neoplasms.

GENERAL COMMENTS

Ionizing radiations were the first type of carcinogens thoroughly
studied and much of what has been done subsequently with chemical car-
cinogens is a duplication of the work with these physical carcinogens. In
carcinogenic potency, ionizing radiations are second to none, as indicated
by the certainty and frequency with which they induce neoplasms. In
variety of neoplasms induced, precision in quantitation, and as a tool in
cancer research they match any other carcinogen. Nevertheless, after
the hazards of radiations were discovered and adequately controlled,
research on this type of carcinogenesis waned until recent developments
have made this area again a highly important field of investigation.

The dose-time relationship after administration of a single dose of a
radioisotope was well worked out by Brues (1949) with the use of Sr89,
which causes neoplasms at the site of deposition. His findings are
illustrated in Fig. 18-10. Each quantity of Sr89 absorbed by bone confers
a given probability of bone tumor formation, the tumor development
time decreasing and tumor incidence increasing with the dosage. It is
not known whether a threshold dose exists for such effects, the limiting
factor being the survival time. The daily tumor morbidity will con-
stantly increase as long as further irradiation occurs.

Carcinogenesis by a bone-seeking radioisotope cannot be accurately
computed on body-weight basis. The concentration in the area of sus-
ceptible cells, rate of elimination from the area, and degree or character of

1186

RADIATION BIOLOGY

the specific ionization may be important. Brues (1949) found that the
development period of bone tumors is comparable in mice, rats, rabbits,
and dogs. His findings are illustrated in Fig. 18-1. The ordinate repre-
sents the probability that a mouse in a given dosage group will develop a
tumor or possess it in a microscopic state on a given day.

Species differences doubtless exist, as illustrated by leukemia and
ovarian tumor induction in mice, and difficulties of their induction in
other species of animals; however, not enough work has been done in
other species and the tumor development time may not have been given
adequate consideration. The species differences are probably the same
for all types of carcinogens, e.g., leukemias are readily induced in mice

0.020

0015 -

0010

0005

100 200 300 400 500 600 700 800
DAYS
Fig. 18-10. The relation of dose of Sr89 to rate of induction and latency period of bone
tumors {after Brues, 1949).

also by chemical carcinogens and hormones, but not in other species.
Work in this area is, however, inadequate.

Cocarcinogenesis by Irradiation. Many observations made on the
induction of neoplasms in man or animals subjected to ionizing irradia-
tion are best explained by postulating cofactors as being operative, before,
during, or after irradiation.

Observations made in the course of therapeutic use of X radiation in
the control of chronic inflammatory lesions in man induced Lacassagne
and Vinzent (1929) to study the promoting factor of inflammation in
carcinogenesis. In a series of studies, Lacassagne et at. (1927-1933) have
conclusively demonstrated that, in the course of inflammation, many
types of cells are potentiated to the carcinogenic effects of X radiation.
Streptobacillus caviae introduced into the thigh of the rabbit produced a
suppurative inflammation which was often fatal but could be cured by
X radiation. From six months to four years later infiltrative metastasiz-
ing neoplasms appeared in several rabbits so treated. Two malignant
tumors (an osteoblastoma and a fibromyosarcoma) were obtained in the
pectoral region of rabbits in which abscess was produced with diatomace-

CARCINOGENESIS BY IONIZING RADIATIONS 1187

ous earth, followed 6 days later by X irradiation. The first tumor
appeared six, the second thirteen months after irradiation.

A third tumor similarly induced was atypical bat also of bony origin.
A fourth tumor was a periosteal sarcoma, the fifth a myxosarcoma, and
the sixth an intracanalicular epithelioma. Tumors were not encountered
in rabbits treated either with this bacillus or X radiation alone. The dose
of X radiation was smaller than that required to produce radiodermatitis
(600 r) and in three rabbits it even failed to cause epilation.

The great variety of the histological types of the tumors (osseous,
periosteal, aponeurotic, loose connective tissue, muscular and ductal-
epithelial) are attributed to the preparatory intervention of the inflam-
matory process, raising the sensitivity of different cells to X radiation.

Burrows et at. (1937) fully confirmed the work of Lacassagne. They
produced a focus of inflammation in the groin in each of twelve rabbits by
injections of kaolin and of finely powdered silica suspended in olive oil.
These foci were exposed to X radiation, each receiving a single dose of
600 r. Among nine rabbits thus treated and surviving for two years or
longer, tumors appeared in the irradiated tissues in six. In four of these
rabbits the tumors were sarcomas that had produced metastases.

On the one hand, X radiation has been employed in the treatment of
inflammation; on the other hand, there are numerous observations indi-
cating that inflamed tissues are more readily rendered cancerous than
normal tissues. Most of these observations were made in the course of
therapeutic irradiation of tuberculous joints, and the bone tumors devel-
oped in the area of irradiation. The predisposing factor of inflammation
frequently present in the female genital tract has been blamed for cancers
in the vulva, cervix, and uterus (Bumm, 1923).

A cocarcinogenic effect of methylcholanthrene and externally applied
j8 radiation from P32 was reported by Hamilton and Passonneau (1949).
Each of these agents alone caused skin tumors in 13.3 per cent of the
mice and, when combined in the same doses, in 53.3 per cent.

Mottram (1937) observed the cocarcinogenic effects of benzopyrene
and ionizing irradiations. Painting the skin of mice twice weekly with
3,4-benzopyrene for six weeks caused some reaction of the skin but no
tumors. However, with continued painting or additional application of
a single dose of /3 or y radiation, an appreciable number of tumors was
induced. Similar results were obtained by Mayneord and Parsons
(1937), who used X radiation in combination with benzopyrene or
dibenzanthracene.

An enhancement of the leukemogenic action of methylcholanthrene by
X irradiation was described by Furth and Boon (1943). Destruction of
hemopoietic tissues by X irradiation is followed by regeneration, and
during this phase mitotic figures are seen in abundance in sections of
blood-forming organs. It seemed probable that, during this phase, the

1188 RADIATION BIOLOGY

blood-forming organs would be particularly susceptible to leukemogenic
irritants. Experiments performed to test this assumption have shown
that the leukemogenic action of small doses of methylcholanthrene, which
alone rarely produces leukemia, is greatly enhanced by preirradiation
with doses of X radiation. It is obvious that proper timing and dosage
are necessary to obtain such results. A similar coleukemogenic effect
of these two agents was also noted by Mixer and Kirschbaum (1948)
and a similar synergistic effect was found in inducing thymic tumors by
estrogenic hormones and X irradiation (Kirschbaum et at., 1949; Kirsch-
baum and Mixer, 1951). Agents such as cortisone, which cause involu-
tion of the thymus and depress lymphopoiesis, will probably lessen the
liability of leukemia induction by X irradiation (Kaplan, 1951). Thus
the reduced incidence of leukemia may be explained when pneumonia
or other diseases, which cause accidental involution of the thymus, are
prevalent in the animal colony.

The complexities of carcinogenesis are best illustrated by those of the
mammary tumor of mice which has three major causes — genetic, hor-
monal, and viral; if powerful, either of the latter two may cause mammary
tumors provided the genetic factor is present (Bittner, 1946-1947). Ion-
izing irradiation may produce mammary tumors in an agent-free strain
(Lorenz et at., 1951) ; it is not likely that this is accomplished by a direct
mutagenic action on cells of the mammary gland. The genes are no
doubt ever-present modifiers of all extrinsic agents but relatively little is
known on genetic factors in carcinogenesis by ionizing irradiations. The
uniformity of response of different strains of the same species to the same
dose of ionizing radiations administered under comparable conditions is
impressive. However, not enough work has been done on this subject.

Certain heavy metals not possessing radioactivity have proved to be
carcinogenic under certain experimental conditions. The first experi-
ments pointing this out were performed by Schinz and Uehlinger (1942).
Hueper (personal communication) injected powdered uranium into the
bone of rats, 36 per cent of which developed tumors within six months.
Some of these tumors originated from the periosteum, others from
adjacent muscle.

MECHANISM OF CARCINOGENESIS

Electromagnetic as well as corpuscular radiations can be considered
together, since no qualitative differences have been disclosed between the
two. The release of energy in cells by absorption of ionizing electro-
magnetic radiation is accompanied by liberation of secondary electrons
which form ions along their path. These ions may be unstable, undergo
chemical changes, and interact with molecules of the tissues. The greater
part of the ionization will occur in the water of the tissues, resulting in

CARCINOGENESIS BY IONIZING RADIATIONS 1189

the formation of OH, H, and other radicals. These radicals will interact
with different cellular constituents. This fundamental ionization process
is identical for all ionizing radiations whether electromagnetic or cor-
puscular. A biologic system can recover at least partially if sufficient
time elapses between successive ionization events.

Concerning views and findings relative to the mode of action of radia-
tions on living cells, the reader is referred to Scott (1937), Lea (1947),
Tobias (1951) and Chap. 6 of this book, by Zirkle.

If a single ionization event can cause a carcinogenic change by a direct
action of the radiation on the cell, the carcinogenic action will be inde-
pendent of dose rate, but a minimum dose will be required to induce a
macroscopic tumor during the life span of the animal. On the other
hand, if several ionization events are necessary for the carcinogenic
change, the action of the radiation may depend on the dose rate as is the
case with particles of low specific ionization, such as penetrating X or y
rays.

A survey of the facts gathered indicates a multiplicity of mechanisms
by which neoplasia is produced by ionizing radiations. Instances of both
direct and indirect effects are evident, although they are not always
clearly identified. The immediate cause of the neoplastic change is, how-
ever, still hidden.

In the genesis of tumors of endocrine organs, such as pituitary tumors
which arise following thyroid destruction by I131, radiation may do
barely more than destroy the thyroid. Similarly, an endocrine "imbal-
ance" brought about by ovarian irradiation appears to be the major if
not the sole force in the genesis of ovarian tumors. It is possible that
thyroid tumors arising after administration of I131 are caused by the
combination of a local effect on the cells and excessive output of thyroid-
stimulating hormones by the pituitary.

The "bone seekers" such as Sr8990, mesothorium, and radium, on the
contrary, seem to exert their effect directly on osteogenic cells with
which they come in contact. Injury and sustained regenerative efforts
are characteristic events in the course of evolution of both bone and skin
tumors.

If it is assumed that neoplasia induction requires a new type of cell
with permanently altered reproduction, carcinogenesis is best looked upon
either as a special type of mutation or that of abnormal differentiation
(Waddington, 1947; Henshaw, 1945). Waddington relates the problem
of the origin of cancer to that of cellular variations, in particular to those
that are discontinuous and irreversible, but he considers carcinogenesis to
be of a special type. Henshaw considers mutation a plausible explana-
tion of neoplasia if this term is applied to irreversible changes in extra-
chromosomal as well as chromosomal constituents of cells. Mutation in
this sense, however, does not differ from differentiation (Henshaw, 1945).

1190 RADIATION BIOLOGY

Mutagens are numerous, and ionizing radiation acting directly on a
"sensitive" part of the cell is one of them. Others may be endogenous
in origin.

Experiments were reported showing that cholesterol exposed to massive
doses of X radiation (about 60,000 r) acquires carcinogenic power (Bur-
rows and Mayneord, 1937), but this finding has not been confirmed.
This and other ideas such as the transformation of an endogenous steroid
into a carcinogenic hydrocarbon and the meaning of the presence of
carcinogens in tissue extracts, are fully discussed by Lacassagne in his
monograph on cancers produced by endogenous substances (1950).
Concerning problems common with carcinogenesis by ultraviolet rays,
see Rusch (1949) and Blum (1953).

The assumption of indirect carcinogenesis by endogenous carcinogens
is also speculative, as no one has yet proved that endogenous carcinogens
isolated from the body exist in that form and are not the products of the
chemical techniques used for their isolation. Suggestive evidence for
the mutation theory is furnished by studies on the genetic character of
the neoplastic cells (Furth et al., 1944), by demonstration of mutagenic
power of some carcinogens (Demerec, 1948, and Latarjet et al., 1949), the
carcinogenic power of some mutagens (Boyland and Horning, 1949;
Burdette, 1950; and Heston, 1950), and by the character of mitotic
abnormalities, but direct demonstration of a chromosonal change is
lacking. Mottram (1931) tested the validity of the somatic mutation
hypothesis by exposing cells in vitro to high concentration of carbon
dioxide and 0 rays which, in Drosophila, gives rise to derangements of
chromosomes. The irradiated cells were reimplanted in the animal
but only one testicular tumor was produced. Such experiments are
cumbersome, yet are worth pursuing to find out whether the thesis, that
mutation is proportional to the dose, applies to carcinogenesis.

That tissues which are subjected to an increased physiological or patho-
logical regenerative process are more susceptible to neoplasia induction
than normal tissues has long been demonstrated by several investigators.
This may, in part, be due to the presence of an increased number of cells
in a sensitive (mitotic) phase. The mere fact that ionizing radiation is
one of the most powerful agents causing genetic alterations (Muller,
1938; Catcheside, 1948) lends strong support to the somatic mutation
theory of cancer.

Based on studies of Polytoma uvella, Lacassagne (1936) postulates five
types of lesions in the cell: (1) temporary suppression of growth due to
absorption of energy in the cytoplasm (associated with a reparable injury
to mitochondria and other cytoplasmic structures), (2) destruction of
parts of chromatin resulting in abortive anomalies of division, (3) sup-
pression of reproduction due to injury of centrosomes, (4) suppression of
motility due to injury of motor centers, and (5) immediate death of cell.

CARCINOGENESIS BY IONIZING RADIATIONS 1191

The neoplastic change is explained as an anomaly of cell division. Lacas-
sagne distinguishes between equipotential mitosis, from which arise
similar daughter cells, and differentiated mitosis, which yields dissimilar
daughter cells. The malignant transformation is related to alteration of
the reproductive apparatus of the cells, fixing their descendants and
causing preponderance of equipotential mitosis.

The tumors arising in the endocrine organs (e.g., pituitary) represent
another extreme type of growth in which little if any change in character
of the cell has to be postulated, since multiplication of these tumor cells,
like that of their normal homologues, is caused by a sustained endocrine
stimulus. Since they metastasize, they behave as malignant growths,
yet they may be checked by correction of the hormonal disturbance
(Furth et at., unpublished data). It may be disputed that such condi-
tioned growths could be classified as tumors. On the other nand,
ignorance of the processes on which the origin and sustained growth of
common neoplasms may depend is no guarantee that such processes do
not exist. In any event the known dependent growths gradually or
suddenly lose the dependency and sooner or later become autonomous.

The statement that irradiation may hasten the aging process and thus
bring about an earlier appearance of spontaneously occurring tumors does
not seem to hold for all neoplasms. Leukemia can be induced in young
animals by irradiations and other carcinogens with high frequency in
strains of mice in which this disease is exceedingly rare (e.g., in strain
C57 black). Many neoplasms occur in young people and are of types
not caused by ionizing irradiations. On the other hand, many tumors
observed induced by irradiations are of the same type as found
spontaneously.

The induction of neoplasms by ionizing irradiation is a fascinating
chapter in the history of science. It is full of problems of increasing
practical importance awaiting solution. As a tool in cancer research, it
is unequaled.

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Most of the older literature, well reviewed in the monographs of Lacassagne
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clinical reports, only those of historical interest and the more recent ones are listed,
notably those which cite the older literature. Of the publications of authors with
numerous articles, only the first, the last, and the more comprehensive ones are listed.

(Information on availability of government reports indicated by an asterisk may
be obtained from the Office of Technical Services, Department of Commerce,

Washington, D.C.)

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1192 RADIATION BIOLOGY

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Versammlung von Vergiftungsfallen, Vogel, Berlin, 2: 79-80.
Zirkle, R. E. (1953) The radiobiological importance of linear energy transfer.

Chap. 6, this volume.

Manuscript received by the editor Apr. 29, 1952

Addendum

For more recent publications see:

Furth, J., and A. C. Upton (1953) Histopathologic and carcinogenic effects of

ionizing irradiation. Ann. Rev. Nuclear Sci., 3: 303-338.
and (1954) Late effects of experimental nuclear detonation in mice.

Radiology, in press.

NAME INDEX

Page numbers in boldface type denote bibliographical references
A

Abderhalden, R., 1000, 1008
Abele, R. K., 160, 176, 189
Abelson, P. H., 998, 1008
Abrams, H. L., 926, 933, 947
Abrams, R., 305, 308, 962, 983, 1008
Abrahamson, L., 1179, 1191
Abrahamson, M. L., 1179, 1191
Adair, R. K., 115, 141, 161, 185
Adamova, N., 1155, 1193
Adams, G. D., 112, 141
Adams, W. G., 794, 795
Adams, W. S., 966, 1008, 1035
Aebersold, P. C., 161, 185, 320, 330, 333,

336, 337, 339, 343n., 344, 347, 348,

350, 786, 824, 827, 830, 833, 838, 859,

923, 947, 1041, 1042
Ahlstrom, L., 276, 278, 305, 306, 308, 926,

947
Ahmed, I. A. R. S., 662, 678, 704
Albers, D., 299, 308
Albers-Schonberg, 1114
Alberti, M., 776, 782, 799, 804, 811, 816,

817, 822
Alberti, W., 554, 602, 628, 629, 696, 1097
Albot, G., 1184, 1195
Allen, A. C., 926, 948, 982, 985, 1010
Allen, A. O., 328, 344, 559, 602, 750, 751,

753, 756
Allen, B. M., 682, 708, 935, 936, 947
Allen, E., 906, 914
Allen, J. G., 926, 947, 966-969, 1003,

1005, 1008, 1017, 1018, 1025, 1031,

1035, 1044-1047, 1072, 1082
Allison, S. K., 68, 107, 142
Allsopp, C. B., 272, 274, 278, 655, 696,

750, 756, 757
Almy, G. M., 112, 141
Alper, T., 275, 278
Altenburg, E., 362, 389, 391, 393, 395,

405, 412, 430, 446,462, 469, 500, 520,

531, 534-538, 545, 602, 618, 629, 635,

706
Altenburg, L. S., 534-538, 545, 602
Altenburger, K., 131, 141

Altman, K. I., 962, 1002, 1008

Amato, A., 554, 602, 627, 696

Ames, F. B., 686, 708, 831, 837, 841, 842,

845, 846, 859
Amoroso, E. C., 832, 834, 835, 837, 856
Amory, H. I., 1182, 1192
Ancel, P., 935, 947, 1118
Anderson, C. D., 40
Anderson, E. A., 933, 958, 984, 1008
Anderson, E. G., 457, 462, 644, 696, 697
Anderson, E. H., 509, 561, 563, 564, 579,

580, 595, 602, 611, 746, 758, 856, 858
Anderson, E. K., 1004, 1011
Anderson, R. S., 287, 288, 308, 940, 947
Andervont, H. B., 1157, 1177, 1192
Andrews, M. B., 414, 466
Ane, J. N., 971, 978, 1008
Ansari, M. Y., 415, 466
Anslow, G. A., 161, 185
Anson, B. J., 1132
Anthony, E., 968, 1016
Apolant, H., 553, 603
Appleyard, R. K., 166, 185
Araratian, A. G., 363, 467, 480, 615, 799,

821
Archangelsky, B. A., 900, 914
Ardao, M. I., 304, 309
Arnason, T. J., 584, 603
Arnberg, B., 509, 584, 604
Arnold, G., 554, 603, 1114
Arnow, L. E., 696, 697
Arnstein, A., 1177, 1192
Aron, W. A., 87, 141
Aschheim, E. F., 1149
Ashler, F. M., 692, 707
Astaurov, B. L., 510, 603
Aub, J. C., 998, 1024, 1155, 1157, 1193
Aubertin, C., 1030, 1031, 1034, 1035,

1061, 1162, 1165, 1192
Auerbach, C., 136, 141, 363, 415, 417,

422, 458, 463, 507, 512, 514, 551, 577,

578, 600, 603, 656, 657, 686, 697,

704, 843, 858
Auerbach, O., 1182, 1192
Auger, P., 34, 47
Axelrod, Dorothy, 330, 339, 344, 1112

1203

1204

RADIATION BIOLOGY

B

Babcock, E. B., 582, 603

Bachem, A., 990, 1014

Bacher, R. F., 198, 252

Back, A., 305, 311, 961, 1002, 1015, 1097

Back, L., 682, 701

Baclesse, F., 946, 947, 1157, 1195

Bacq, Z. M., 131, 141, 275, 280, 655, 656,

697, 941, 945, 946, 947, 948, 951,

1067
Baeten, G., 1155
Bagg, H. J., 390, 467, 864, 865, 869, 870,

879, 893-896, 898, 914, 915
Baidens, A. von, 994, 1009
Baily, N., 167, 176, 186
Baker, C. P., 5, 144
Baker, R., 537
Baker, W. K, 509, 561, 563, 565-568,

579, 580, 603, 611, 663, 674, 675,

697, 746, 758, 856, 858, 904, 936, 940,

948
Bakker, C. J., 66, 87, 141
Bali, Talia, 1168, 1169, 1192
Bancroft, F. W., 390, 467
Bardeen, C. R., 554, 603
Barling, G., 1148, 1192
Barnard, R. D., 985, 1009
Barnes, B. T., 23, 142
Barnes, K. K., 319, 333, 348, 1030, 1058
Barnes, W. A., 926, 948, 960, 1000, 1009
Barnett, J. C, 925, 949
Barratt, J. O., 554, 603, 1114
Barron, E. S. G., 245, 257, 262, 265, 278,

286-289, 293, 295, 298, 299, 301-308,

310, 312, 313, 551, 580, 599, 603, 922,

937, 941, 948, 962, 973, 984, 986, 987,

989, 990, 1001, 1009, 1011, 1021,

1067
Barrow, J., 979, 988, 1009
Bass, H., 176, 185
Bate, R. C., 650, 703
Bateman, A. J., 584, 603
Bates, M. I., 1157, 1184, 1198
Battacharya, P., 514, 515, 603
Bauer, H., 379, 380, 463, 481, 482, 495,

604, 628, 630, 646, 650, 655, 662 664,

667, 670, 673, 674, 677-679, 681, 687,

697
Baur, E., 415, 463, 511
Baxendale, J. H., 291, 309
Beadle, G. W., 373, 414, 461, 463, 472
Bean, W. B., 985, 1009
Beattie, J. W., 146, 187
Beattv, A. V., 559, 561, 568, 609, 620,

742-744, 746, 748, 749, 752, 753, 758,

759, 941, 950

Beaujard, E., 1030, 1031, 1034, 1035,

1061
Beck, 1180

Becker, R. M., 1003, 1009
Becquerel, H., 1150
Bedichek, S., 455, 470, 641, 707
Bediirftig, G., 980, 1009
Behounek, F., 1178
Beiler, J. M., 979, 1009
Belgovsky, M. L., 363, 380, 463, 480,

493, 553, 604
Bell, A. L., 910, 914
Belling, J., 368
Belot, J., 1151, 1192
Belser, N. O., 268, 280, 703
Bender, A. E., 992, 1009
Benedek, A. L., 966, 968, 974, 1024
Benjamin, E., 1002, 1009, 1080
Bennett, L. R., 940, 941, 944, 945, 949,

971, 974, 985, 986, 1003, 1009, 1021,

1037, 1066, 1067
Bennett, V. C, 971, 974, 1009
Berg, M., 926, 955
Berg, R. L., 480
Bergendahl, J., 538, 602
Bergonie, J., 553, 604, 627, 681, 697, 927,

948, 1114, 1127, 1133
Berman, Z. I., 409, 463
Bertalanffy, F. D., 1162, 1197
Bertani, G., 417, 422, 464
Bethard, W. F., 926, 952, 983, 1002, 1018,

1045, 1057, 1065, 1070, 1072, 1079,

1080
Bethe, H. A., 66, 87, 89, 141, 143, 165,

166, 185
Betz, H., 941, 945, 951
Beutel, A., 971, 1009
Bevan, E. S., 290, 310
Bigelow, R. R., 966, 968, 971, 978, 979,

1009, 1039
Billen, D., 580

Biltris, R., 1154-1156, 1184, 1192, 1193
Bink, N., 984, 985, 1001, 1025
Binks, W., 153, 187
Birch-Hirschfeld, A., 1131
Bird, M. J., 416, 424, 463
Birkina, B. N., 414, 463
Bischoff, F., 999, 1009
Bisgard, J. D., 1112
Bishop, C. J., 660, 697, 722, 723, 757
Bishop, D. W., 628, 665, 697
Bishop, F. W., 932, 948
Bishop, M., 532, 540, 606, 677, 697
Bittner, J. J., 1188, 1192
Blackford, M. E., 937, 938, 942, 946,

954
Blackwood, O., 525, 530, 604
Blakeslee, A. F., 368, 391, 465

NAME INDEX

1205

Blanc, J., 1114

Blanchard, C. H., 92, 141

Blau, M., 131, 141

Bloch, B., 1152, 1153, 1192

Block, M. H., 930, 935, 952, 963, 965, 967,

1017, 1018, 1025, 1059
Blocker, W., 97, 141
Blomfield, G. W., 164, 185
Bloom, M. A., 935, 948, 962, 983, 997-
999, 1009, 1010, 1031, 1036, 1037,
1099, 1100, 1101, 1112
Bloom, W., 929-932, 943, 948, 960-964,
988, 989, 997-999, 1010, 1022, 1023,
1026, 1031, 1033, 1035, 1036, 1064,
1093-1096, 1100, 1101, 1110, 1112,
1118, 1128, 1161, 1192
Blount, H. C, Jr., 935, 946, 948, 956, 993,

1010
Blum, H. F., 597, 604, 811, 812, 817,

1175, 1190
Blumel, J., 682, 697
Blumenthal, G., 601, 616, 928, 953
Boche, R. D., 687, 705, 932, 948
Bodemann, E., 849, 859
Boffil, J., 926, 948, 963, 1010
Bogliolo, L., 1157, 1192
Bohn, G., 553, 554, 604
Bohr, N., 51

Bollinger, A., 1000, 1013, 1128
Bolomey, R. A., 584, 609, 733, 758
Bond, V. P., 926, 934, 948, 949, 956, 961,
963, 965, 982, 985, 1005, 1010, 1011,
1016, 1025, 1072
Bonet-Maury, P., 274, 278, 286, 309, 328,
344, 567, 604, 744, 750, 751, 757, 938,
948
Bonham, K., 448, 463, 465, 710
Boni, A., 589, 609
Bonner, D. M., 461, 462, 593, 604
Bonnier, G., 477, 509, 510, 513, 514, 604,

644, 697
Bonte, F. J., 1160, 1182, 1196
Bonzell, V., 287, 289, 309
Boon, M. C, 1168, 1187, 1190, 1194
Borak, J., 976, 977, 1010, 1123
Borchert, R., 542, 615
Borges, W., 1163, 1194
Borisoff, 480
Born, H. J., 574, 605
Borowskaja, D., 1004, 1010
Bortner, T. E., 176, 185
Bose, I., 642, 708
Bostian, C. H., 388, 473, 683, 710
Bothe, W., 40, 72, 82, 83, 92, 113, 141,

142
Bouin, P., 1118
Bouricius, J. K., 848, 856

Boveri, T., 394, 464

Bowen, T., 166, 185

Bower, J. O., 1132

Bowers, J. Z., 929, 957, 966, 1005, 1010,

1027
Boyd, W., 589, 609
Boyland, E., 424, 464, 600, 605, 1190,

1192
Boys, F., 967, 1010
Bozeman, M. L., 665, 679, 688-690, 697,

705
Braasch, N. K., 977, 1010
Bradley, Muriel, 338, 347, 684, 705, 798,

821
Bragg, W. H., 87, 88, 91, 93, 157, 185
Brambell, F. W. R., 1168, 1192
Brandt, C. L., 304, 310
Brandt, E. L., 424, 465
Brar, S. S., 168-170n., 185
Braun, R., 180, 187
Brawner, H. P., 1001, 1010
Bream, H., 1003, 1004, 1024
Brecher, G., 926, 942, 943, 948, 949, 967,
1001, 1010, 1067, 1076

Breed, H. E., 112, 141

Brennan, J. T., 943, 948, 1001, 1011

Brenneke, H., 686, 697, 826, 827, 832-
835, 856, 858, 999, 1010

Brent, R. L., 888, 916, 918, 1185, 1201

Bretscher, E., 139, 143, 299, 301, 312,
330, 331, 347

Brewster, W., 689, 707

Brick, I. V., 986, 1010

Bridges, C. B., 358, 368, 393, 464, 495,
496, 605, 638, 643, 644, 650, 664, 698

Bridges, P. N., 532

Bright, E. M., 936, 955

Brinckerhoff, R. F., 167, 188

Brinkhouse, K. M., 966, 968, 1022

Brohult, S., 296, 313

Bromley, D., 147, 185

Brookhaven National Laboratory, 630,
698

Brooks, B., 1112

Brooks, H., 116, 143

Brooks, J. W., 934, 948, 1005, 1010

Brooks, R. E., 979, 1024

Brown, C. S., 972, 974, 1010

Brown, H. M., 555, 609

Brown, M. B., 991, 1005, 1018, 1164,
1165, 1196

Brown, M. G., 303, 314

Brown, M. S., 650, 707

Brown, P., 1149, 1192

Brown, R. F., 994, 1018

Brown, W. H., 985, 1018

Brownscombe, E. R., 328, 345

1206

RADIATION BIOLOGY

Brues, A. M., 303, 310, 925, 929-932, 948,
955, 956, 961, 964, 965, 967, 969-975,
980, 981, 984, 987, 989, 995, 1000,
1001, 1005, 1010, 1011, 1022, 1023,
1026, 1027, 1031, 1074, 1128, 1135,
1146, 1159, 1166, 1176, 1177, 1179,
1184-1186, 1192, 1197

Brumrield, R. T., 484, 566, 578, 605, 621,
725, 737, 757, 760

Brunschwig, A., 346

Brunst, V. V., 924, 948

Bryson, V., 553, 592, 605

Buchanan, D. L., 293, 310, 973, 1011

Buchmann, W., 414, 464, 574, 605

Buchsbaum, R., 938, 948, 961, 972, 1011

Buchwald, K. W., 986, 1011

Buechner, W. W., 25, 141

Bumm, E., 1171, 1187, 1192

Bunker, J. W. M., 1183, 1194

Bunting, C. H., 987, 1022, 1128

Burch, G. E., 971, 978, 1008

Burckhard, G., 865, 866, 893, 894, 914

Burdette, W. J., 423, 464, 650, 698, 1190,
1192

Burhop, E. H. S., 193, 202n., 205, 206,
208, 211, 236, 237, 239, 243, 253

Burnett, W. T., Jr., 579, 605, 611, 1171,
1172, 1194

Burr, B. E., 993, 1001, 1022

Burrill, E. A., 25, 141

Burrows, H., 1187, 1190, 1192, 1193

Burrows, W., 1003, 1004, 1011

-Burstone, M. S., 864, 894, 899, 914

Burton, M., 193, 253

Busch, E., 966, 1011

Buschke, W., 811, 812, 816, 817, 818

Bush, F., 168, 169, 185

Bushland, R. C, 441, 464

Bushnell, R. J., 589, 625

Bushnell, R. S., 737, 761

Butler, C. L., 987, 1014

Butler, E. G., 776, 817

Butler, G. C., 289, 313, 694, 698, 708,

754, 757

Butler, J. A. V., 277, 278, 290, 310, 754,

755, 757

Butterworth, J. S., 1169, 1173, 1194
Buu-Hoi, 1190, 1197
Buzzati-Traverso, A. A., 130, 141, 437,
443, 464, 630, 698

C

Cade, S., 778, 817

Cahan, W. G., 1181, 1182, 1193

Caillot, T., 274, 280

Caldecott, R. S., 568, 571, 572, 605

Camerino, B., 289, 311

Campbell, B., 995, 1011

Campbell, I. L., 964, 1011

Cannon, C. V., 147, 160, 185

Canti, R. G., 773, 776, 778, 782, 788, 790,
817

Capron, P. E., 163, 185

Carlson, J. G., 125, 141, 484, 605, 628-
630, 632, 643, 645, 659, 661, 666, 667,
679, 682, 698, 748, 757, 763, 765, 768,
769, 773-780, 782, 789, 791-794,
799-801, 804-816, 817, 818, 1094

Carothers, E. E., 628, 698

Carr, J. G., 415, 463

Carson, G., 403, 427, 472

Carter, C. E., 300, 310

Carter, R. E., 943, 948, 966, 1011

Carter, T. C, 907, 918

Carty, J. R., 936, 940, 945, 948

Carvajal-Forero, J. de, 986, 1017

Caspari, E., 131, 141, 476, 605, 630, 656,
658, 659, 666, 701

Cassen, B., 148, 185

Catcheside, D. G., 135, 138-140, 141, 143,
342, 344, 379, 382, 464, 480, 482, 487,
488, 490, 491, 499, 517, 521, 526-528,
530, 566, 567, 605, 615, 630, 632, 646,
647, 652, 653, 655, 658, 660, 663, 666,
667, 675, 678, 687, 696, 698, 704, 713,
714, 716, 717, 720, 721, 724, 726-729,
731, 732, 735, 741, 755, 756, 767,
769, 825, 856, 1190, 1193

Catsch, A., 481, 483, 484, 519, 535, 605,
621, 669-671, 684, 698, 699

Cattley, R., 803, 818

Cavalli, L. L., 130, 141, 630, 698

Cavallo, L., 160, 189

Chadwick, J., 42, 66, 143

Chaikoff, I. L., 993, 1014, 1015, 1171,
1172, 1195

Chambers, F. W., 264, 278

Chambers, F. W., Jr., 308, 310, 932, 935,

942, 943, 945, 949, 979, 988, 1009
Chambers, R. J., 939, 949
Chanutin, A., 300, 312, 973, 975, 987,

1015, 1020
Chapman, A. B., 848, 859
Chapman, W. H., 264, 278, 934, 935, 942,

943, 945, 949, 1067
Chapman, W. J., 308, 310

Charles, D. R., 484, 616, 727, 759, 844,
847, 848, 850, 851, 855, 856

Charleton, E. E., 12, 112, 141

Chase, H. B., 136, 142, 997, 998, 1011

Chase, H. Y., 811, 818

Chase, J. H., 991, 1004, 1013

Chastain, L. R., 1066

Chastain, S. M., 940, 941, 944, 945, 949,
986, 1009

NAME INDEX

1207

Chesley, L. C, 300, 310

Chew, G. F., 12, 142

Christian, E. J. B., 925, 956, 989, 1026

Christie, J. H., 924, 926, 950, 955, 1002,

1003, 1019
Chrom, S. A., 982, 988, 1002, 1011
Cipollara, A. C, 925, 953
Claesson, L., 994, 1009
Clark, A. M., 388, 464, 683, 699, 929, 949
Clark, G. L., 287, 310
Clark, J. B., 303, 314, 417, 473, 537, 544,

545, 548-552, 562, 606, 610, 623, 625,

626, 751, 761
Clark, J. H., 1132
Clark, R. K., 160, 167, 168, 170n., 176,

185, 186, 188, 923, 955
Clark, W. G., 932, 949, 969, 1011
Clarke, A. M., 93, 144
Clarke, G., 993, 1014
Clarkson, J. R., 1030, 1032, 1061
Clayton, F. E., 561, 562, 566, 567, 610
Cleland, G. H., 417, 464, 550, 607, 752,

757
Clemmesen, J., 1004, 1011
Cloud, R. W., 93, 144, 998, 1024
Clunet, J., 1150, 1152, 1153, 1193, 1198
Cocchi, U., 685, 699
Cochran, K, 937, 949, 987, 988, 1013
Cockcroft, A. L., 165, 166, 185
Coe, W. S., 287, 310
Cogan, D. G., 998, 1011, 1024
Coghill, R. D., 444, 466, 467, 471, 540,

611
Cohen, I., 769, 771, 784, 790, 820
Cohn, M., 869, 876, 894, 900, 914
Cohn, S. H., 968, 1011, 1035, 1044
Cold Spring Harbor Symposia, 630, 699
Cole, J. W., 966, 968, 1017
Cole, L. J., 926, 949, 963, 965, 1011
Coley, B. L., 1181, 1182, 1193
Collins, J. L., 582, 603
Collinson, E., 300, 310
Colwell, H. A., 1094, 1145, 1150, 1151,

1193
Committee on the Standardization of

X-ray Measurements, 150, 185
Compton, A. H., 37, 38, 44, 69, 71, 102-

105, 107, 137, 142
Conard, R. A., 985, 986, 1011
Congdon, C. C, 964, 965, 1020, 1075,

1165, 1198
Conger, A. D., 73, 139, 142, 341, 345, 386,

417, 464, 562, 563, 573, 588, 606, 726,

734, 737, 745, 748, 757
Conlon, P., 1182, 1198
Conn, E. E., 308, 310
Conway, B. E., 755, 757
Cook, E. V., 935, 949

Coolidge, A. S., 216, 252

Coon, J. M., 914, 915, 957

Cooper, K. W., 520, 606, 653, 654, 699

Cope, O., 998, 1024

Copp, D. H., 1112

Corey, R. B., 295, 313

Corscaden, J. A., 908, 916

Corson, D. R., 5, 144

Cottet, P., 577, 606

Coulson, C. A., 319, 320, 347

Coulter, M., 1002, 1003, 1017

Coutard, H., 893, 898, 915, 1124

Cowing, R. F., 829, 837, 857, 999, 1014

Crabb, E. D., 967, 1018

Crabtree, H. G., 275, 278, 305, 306, 310,

319, 339, 344, 558, 606, 936, 939-941,

945, 946, 949, 1001, 1010
Craddock, C. G., Jr., 970, 978, 991, 1003,

1004, 1012, 1019, 1027
Cramer, W., 275, 278, 558, 606, 936, 939-

941, 945, 946, 949
Craver, B. N., 991, 1012
Creighton, M., 628, 642, 660, 699, 776,

818
Crevecoeur, E., 163, 185
Crone, H. G., 227, 253
Cronkite, E. P., 264, 278, 308, 310, 926,

934, 935, 942, 943, 948, 949, 966, 968,

969, 991, 1005, 1011, 1022, 1031,

1035, 1044-1046, 1067, 1076
Cross, C. F., 290, 310
Grouse, H. V., 665, 679, 687, 699
Crow, H. E., 799, 818
Crowell, J., 811, 812, 818
Crowther, J. A., 283, 310, 525, 606
Cummings, E., 584, 603
Cupp, M. N., 968, 969, 979, 1019
Curie, M., 134, 142
Curl, H., 978, 1023
Curran, S. C, 165, 166, 185
Curtis, H. J., 933, 935, 956, 958, 1005,

1026
Curtis, L. R., 148, 185
Czepa, A., 1030

D

Daels, F., 1154-1156, 1193

Dainton, F. S., 236, 252, 257, 270, 278,

279, 291, 300, 310
Dakin, H. D., 297, 310
Dalcq, A., 643, 699
Dale, W. M., 257-263, 265, 267-269, 271,

272, 275, 279, 293, 296, 297, 299, 300,

307, 310, 329, 344, 599, 656, 699, 938,

949
Daily, C. M., 1148
Dalton, A. J., 1172, 1199

1208

RADIATION BIOLOGY

D'Amato, F., 275, 279, 556, 563, 575, 576,

578, 590, 606
Dancoff, S. M., 112, 141
Daniel, J., 919, 949
Daniels, D. S., 488, 614
Darden, E. B., 1167, 1193
Darlington, C. D., 566, 578, 606, 660,

694, 699, 715, 717, 721, 723, 741, 749,

757, 784, 794, 799-801, 818
Darwin, C, 453
Dauphin, J., 390, 464
Davidoff, L. M., 995, 1012
Davidson, H., 592, 605
Davidson, N., 148, 186
Davies, J. V., 260, 262, 263, 265, 267-

269, 279, 296, 297, 310, 656, 699
Davis, R. W., 961, 1012, 1039
Dawson, A. B., 1125
Day, F. H., 107, 153, 180, 186
Day, M. J., 147, 152, 186, 292, 311
Day, P. L., 937, 949
Deal, L. J., 147, 186
Debreuil, G., 554, 620
DeBruyn, P. P. H., 931, 943, 949, 1102,

1103
Dechaume, 1157, 1195
Decker, A. B., 985, 986, 1009, 1021
DeCoursey, E., 999, 1005, 1020
Dehler, H., 1171, 1193
Delbet, P., 1125
Delbriick, M., 130, 144, 412, 413, 464,

476, 526, 582, 597, 606, 624, 625,

630, 699
DeLong, C. W., 922, 949
Demerec, M., 345, 379, 380, 417, 422,

463, 464, 476, 478, 482, 494, 495, 505,

518, 521, 532, 533, 535, 540, 556, 604,

606, 608, 611, 620, 630, 632, 635,

644-648, 652, 656, 658, 659, 661-

663, 666-670, 673, 677, 678, 682,

683, 697, 699-701, 703, 1190, 1193
Demidova, Z. A., 476, 480, 607, 610
Dempster, E. R., 333, 334, 339, 344, 350,

483, 492, 518, 521, 607, 655, 669, 671,

683, 688, 700, 786, 824
Denniston, R. H., 994, 1012
Denstad, T., 962, 1012
Depenthal, 1171, 1193
Deringer, M. K., 847, 856, 858, 963, 1020,

1153, 1154, 1166-1168, 1180, 1193,

1198
Desjardins, A. U., 983, 990, 1012, 1030,

1094, 1125, 1127, 1128, 1131
Dessauer, F., 142, 248
Deufel, J., 774, 776, 779, 782, 799, 800,

818
Deupree, N. G., 1003, 1004, 1011
Devi, P., 557, 607

DeVries, H., 353

Dickey, F. H., 417, 464, 550, 607, 752,

757
Dickman, S., 551, 580, 603, 922, 937, 941,

948
Dickmann, S. R., 262, 278, 298, 299,

309
Dickson, H., 531, 607
Di Giovanni, H., 176, 185
Dillard, G. C. L., 966, 968, 1012
Dinning, J. S., 937, 949
Dirac, P. A., 40
Dische, Z., 943, 957
Dixon, F. J., 686, 710, 968, 983, 1012,

1027, 1057, 1058
Doan, C. A., 1155, 1156, 1200
Dobrovolskaia-Zavadskaia, N., 1155,

1193
Dobson, E. L., 978, 1012
Dobvns, B. M., 172, 186
Dobzhansky, T., 446, 455, 464, 607, 635,

640-642, 644, 648, 700
Dodds, E. C., 986, 1013
Dognon, A., 344, 558, 607
Dohnert, H. R., 1179, 1193
Dole, N., 961, 1012
Doll, E. A., 908, 916
Donaldson, D. D., 998, 1011
Donaldson, L. R., 448, 465, 710
Donaldson, M., 788, 817
Doniach, I., 1172, 1193
Doniger, J., 963, 1020
Dooley, R., 1065

Doring, H., 414, 472, 541, 586, 607, 623
Dotterweich, H., 576, 607
Doub, H. P., 1000, 1013
Dougherty, T. F., 990, 991, 1004, 1013
Douglas, D. M., 985, 1013
Doull, J., 937, 949, 987, 988, 1013
Dowdy, A. H., 923, 930, 935, 940, 941,

944-946, 949, 951, 960, 963, 993,

1016, 1019, 1037, 1039, 1044, 1063,

1066, 1067
Downes, Helen R., 348
Driesch, H., 353
Driessen, L. F., 865, 866, 869, 901, 908,

910, 914
Duane, W., 286, 311
Dubinin, N. P., 381, 465, 480, 481, 607,

640, 700
Dublin, L. I., 1162, 1193
Dubois, K. P., 937, 949, 987-989, 1001,

1013
Dubovsky, N. V., 410, 465
DuBow, R., 695, 706
Dubreuil, G., 831, 858
Dudgeon, E., 561, 562, 566, 567, 610
Dudley, H. C, 1061

NAME INDEX

1209

Dudley, R. A., 172, 186

Duggar, B. M., 630, 641, 700, 713, 767,

763, 818
Dulbecco, R., 543, 544, 607
Dunlap, C. E., 960, 966, 1013, 1039, 1061,

1094, 1099, 1155, 1157, 1161, 1193,

1201
Dupuis, P., 1158, 1198
Duryee, W. R., 628, 656, 700, 801-804,

806, 807, 818, 928, 949, 1094
Dutcher, R., 969, 1026
Dyke, C. G., 995, 1012
Dyroff, R., 869, 894, 914

E

Easley, M. A., 23, 142

Eberhardt, K, 334, 344, 480, 487, 607,

641, 700
Eby, F. S., 148, 187
Eddy, C. E., 153, 186
Edelmann, A., 989, 991, 1005, 1013
Eden, M., 993, 1001, 1022
Edington, C. W., 561, 563
Edison, T. A., 1148
Edmonds, H. W., 751, 760
Edmondson, M., 546, 569, 607
Edsall, D. L., 1000, 1013
Efroimson, W. P., 478, 582, 607
Ehrenberg, L., 584, 608
Ehrlich, G., 943, 957
Ehrlich, M., 147, 186
Eisen, H. N., 1004, 1013
Eldredge, J. H., 926, 952, 963, 965, 1018,

1070, 1073, 1077, 1078
Elias, C, 1190, 1197
Ellinger, F., 864n., 914, 925, 926, 932,

949, 950, 956, 977, 987, 990, 992,

1000, 1013, 1094, 1121, 1123
Ellis, C. D., 42, 66, 143
Ellis, F., 788, 819, 975, 1019
Ellis, M. E., 926, 949, 963, 965, 1011
Ellis, R. H., Jr., 168, 169, 171, 188
Elsberg, C. A., 995, 1012
Elson, L. A., 934, 950
Eltzholtz, D. C, 264, 278, 308, 310, 932,

935, 942, 943, 945, 949
Elward, J. F., 994, 1019
Ely, J. O., 692, 693, 700, 934, 950, 961,

964, 974, 981, 983-988, 1001, 1013,

1014, 1024
Emerson, L. C, 1167, 1193
Emile-Weil, P., 1162, 1165, 1193
Emmons, C. W., 531, 535, 536, 540, 573,

608, 611
Endicott, K. M., 1001, 1010
Enerson, D. M., 966-968, 1003, 1008,

1045

Engelstad, R. B., 983, 990, 1014, 1125,

1129-1131
Entenman, C, 975, 1021
Enzmann, E. V., 335, 344, 363, 386, 471,

566, 621, 741, 760
Ephrati, E., 941, 950
Ephrussi, B., 302, 312, 461, 596, 608, 683,

704, 929, 953
Erf, L. A., 339, 348
Erickson, R., 707
Ermala, P., 975, 1025
Ernst, H., 553, 616
Errera, M., 289, 311, 696, 700
Eschenbrenner, A. B., 829-831, 837, 857,

858, 963, 999, 1001, 1014, 1020, 1048,

1049, 1100, 1116, 1118, 1153, 1154,

1165-1168, 1173, 1180, 1188, 1198
Euler, H. von, 276, 278, 279, 305, 306,

308, 692, 696, 700
Evans, A. T., 416, 466
Evans, B. H., 628, 642, 699
Evans, E. I., 935
Evans, M. G., 256, 279, 309
Evans, R. D., 101, 142, 476, 608, 1155,

1157, 1158, 1176, 1178, 1179, 1183,

1193, 1194
Evans, T. C, 274, 279, 301, 303, 311, 336,

337, 345, 785, 818, 923, 925, 936, 940,

945, 950, 993, 998, 1014, 1044, 1048,

1061, 1131
Eversole, W. J., 989, 1013
Ewing, J., 1182, 1194
Exner, F. M., 262, 280, 527, 616, 643, 708
Eyring, H., 236, 252

Faberge, A. C, 480, 482, 488, 532, 533,
566-568, 573, 601, 608, 666, 700, 727,
729, 741, 744, 757

Fabian, G., 577, 608

Faerber, E., 911, 914

Faes, M., 163, 185

Failla, G., 152-154, 157, 162, 164, 167,
176, 179, 186, 188, 189, 274, 279, 301,
311, 319, 320«,., 343n., 345, 348, 349,
785, 818, 938, 939, 950, 972, 1014,
1096

Fainstat, T. D., 907-908, 914

Fairchild, G. C, 788, 819

Fairchild, L. M., 417, 464, 562, 563, 606,
745, 757

Falconer, D. S., 825, 857

Falls, H. F., 430, 470

Fankhauser, G., 643, 700

Fano, L., 14, 26, 59, 116, 144

Fano, U., 92, 109, 121, 131, 135, 139, 141,
142, 144, 157, 186, 237, 252, 344,

1210

RADIATION BIOLOGY

Fano, U. (cont.), 345, 482, 492, 494, 502,
521, 606, 608, 630, 646, 647, 653, 656,
658, 659, 663, 666-668, 671, 672, 676,
677, 680, 682, 683, 699, 700, 701
Farago, A., 345
Farrant, J. L., 153, 186
Farris, E. J., 994, 1015
Farrow, J. G., 644, 699
Farrow, J. H., 182, 183, 188
Fehr, A., 1132
Feigenbaum, J., 299, 311
Feinstein, R. M., 987, 1014
Feldsted, E. T., 994, 1018
Feldweg, P., 909, 914
Fell, H. B., 805, 806, 810, 823
Feller, D. D., 993, 1014
Fellner, O. O., 869, 894, 900, 914
Fenn, W. O., 1002, 1014
Ferkel, R. L., 692, 707
Fermi, E., 166, 186
Fernau, A., 295, 311
Fershing, J. L., 1001, 1015
Feygin, S., 1150, 1194
FIAT Review of German Science, 630,

701
Field, J. B., 966-969, 979, 1014
Fielding, U., 1168, 1192
Fifth International Congress of Radi-
ology, 151, 186
Figge, F. H. J., 1161, 1194

Findlay, D., 993, 1014
Fink, K., 973-975, 1014

Finkel, M. P., 864, 900, 914, 1159, 1160,
1166, 1177, 1180, 1184, 1192, 1197

Finkelstein, P., 295, 309

Fischel, E. E., 1005, 1014

Fischer, P., 941, 945, 948, 1004, 1019

Fisher, N. F., 990, 1014, 1020

Fishier, M. C, 926, 935, 948, 949, 953,
961, 963, 965, 982, 985, 1005, 1010,
1011, 1016, 1018

Fitch, S. H., 147, 186

Fitzmaurice, H. A., 879, 882, 915

Flaskamp, W., 908, 911, 914, 1123

Fleeman, J., 93

Fleischmann, W., 305, 312

Flint, J., 417, 464

Flood, V., 287, 288, 301, 303, 304, 309,
551, 580, 603

Focht, E. F., 86, 143, 176, 180, 182, 188

Fogg, L. C, 829, 837, 857, 999, 1014, 1095

Folley, J. H., 1163, 1194

Folsom, F. B., 943, 951

Ford, J. M., 538, 541, 593, 608, 615

Forfota, E., 1000, 1015

Forkner, C. E., 1155, 1156, 1200

Forro, F., Jr., 139, 143, 330, 348

Forssberg, A. G., 262, 279, 299, 311, 922,

924, 941, 943, 950, 956
Forstat, H., 165, 187
Forster, T., 230, 233, 252
Forsterling, K., 864, 914
Forsythe, W. E., 23, 142
Foster, R. F., 448, 465, 710
Foulds, L., 1157, 1194
Fowler, W. A., 59, 143
France, H. O., 939, 950, 969, 971, 1011,

1015
Francis, D. S., 274, 280, 335, 346, 769,
770, 784, 788-790, 820, 929, 935, 939,
951
Franck, J., 225, 227, 232, 233, 249, 250,

252, 253, 270, 279
Frankenthal, L., 305, 311, 961, 1002, 1015
Franks, W. R., 1161, 1194
Frantz, F., 93
Fraser, F. C, 907, 914, 915
Freed, J. H., 994, 1015
Freeman, P. J., 304, 310
Freund, L., 919, 950
Frey, H., 990, 1015

Fricke, H., 122, 142, 157, 186, 257, 258,

262, 274, 279, 284, 287, 288, 300, 307,

311, 328, 345, 478, 556, 557, 599, 608

Frieben, 919, 950, 1148, 1194

Friedell, H. L., 924, 926, 950, 955, 988,

1015, 1160, 1182, 1196
Frieden, J., 973, 1015
Friedenwald, J. S., 123, 142, 811, 812, 816,

817, 818
Friedman, M., 1182, 1192
Friedman, N. B., 982, 983, 986, 1015,

1027, 1108, 1125, 1161, 1201
Friedrick, W., 153, 186
Friesen, H., 359, 360, 465, 582, 609, 653,

701
Frillev, M., 274, 278, 299, 311, 331, 345,
864, 869, 876-877, 879, 888, 891, 901,
916, 917, 939, 950
Frings, H., 589, 609
Fritz-Niggli, H., 589, 609
Fry, E. G., 991, 1024
Fry, H. J., 771, 818
Fuerst, R., 417, 473, 548-551, 625, 751,

761
Furst, L. J., 910, 918
Furth, F. W., 926, 957, 1002, 1003, 1015,

1017
Furth, J., 933, 950, 961, 966, 968-971,
978, 979, 988, 1009, 1015, 1026,
1028, 1038, 1039, 1047, 1062, 1075,
1082, 1146, 1164, 1166-1169, 1171-
1174, 1180, 1187, 1190, 1192, 1194,
1200, 1201

NAME INDEX

1211

Furth, O. B., 926, 933, 948, 950, 960,
1000, 1164, 1166, 1168, 1174, 1180,
1194

G

Gadsden, E. L., 1171, 1172, 1194

Gager, C. S., 390, 391, 465, 554, 609, 627,
701

Gaither, N. T., 545, 564, 572, 580, 613

Gall, E. A., 345, 347

Gallico, E., 289, 311

Gardner, W. IL, 935, 957, 1066, 1079,
1167, 1168, 1170, 1194, 1197

Garrod, A., 461

Gastaldi, G., 996, 1015

Gaston, E. O., 926, 930, 935, 952, 963,
965, 967, 983, 1002, 1017, 1018, 1046,
1064, 1070-1074, 1076-1079

Gasvoda, B., 293, 301-303, 309

Gates, A. A., 1178, 1196

Gates, O., 1112, 1130

Gaulden, M. E., 561, 569, 609, 748, 757,
796, 799, 808, 814, 816, 817, 818, 904

Gauss, C. J., 908, 914

Gavrilova, A. A., 480, 613

Gay, D. M., 987, 988, 1014

Gay, E. H., 520, 610

Gay, H., 268, 280, 503, 569, 570, 613, 652,
657, 661, 664, 675, 676, 685, 691, 695,
696, 703, 739, 740, 758, 759

Geiger, H., 4

Geigy, R., 531, 609

Gelin, O. E. V., 274, 280, 555, 609

Gentner, C. F., 555, 609

Gentner, W., 5, 40, 72, 82, 83, 1 13, 142

Gerard, P., 1152, 1153, 1195

Gerritsen, A. N., 19-in., 252

Gershenson, S. M., 378, 448, 469, 6(54, 706

Gershon-Cohen, J., 926, 950, 1072

Gerstner, H. B., 1075

Gest, H., 587, 610

Ghent, W. R., 985, 1013

Giese, A. C, 630, 701, 763, 811, 818, 819,
823, 1094, 1097

Gilbert, C, 907, 914

Gilbert, C. W., 265, 268, 271, 275, 279,
296, 297, 310

Gilbert, L. A., 277, 278

Giles, N. H., Jr., 73, 139, 142, 341, 345,
399, 465, 484, 517-519, 521, 558, 559,
561, 568, 584, 588, 594, 596, 597, 606,
609, 616, 620, 656, 658, 668, 701, 721,
723-728, 733, 734, 742-749, 751-753,
757, 758, 759, 904, 940, 941, 950,
1094
Gillman, J., 907, 914

Gillman, T., 907, 914

Gjessing, E. C, 973, 1015

Glass, H. B., 385, 465, 503, 609, 079, 687,
701, 846, 857

Glasser, O., 23, 24, 86, 109, 142, 157, 180,
182, 186, 1146, 1194

Glasser, S. M., 992, 1013

Glenn, J. C, Jr., 1003, 1015

Glocker, R., 331, 340, 345

Glover, J. K, 943, 951

Glticksmann, A., 687, 701, 774, 776, 781,
782, 787, 797, 804-806, 810, 819, 823,
824, 828, 831, 857, 998, 999, 1015

Godwin, I. D., 966, 968, 1022

Goebel, O., 1152, 1153, 1195

Goldberg, R. C, 993, 1015, 1171, 1172,
1195

Goldfeder, A., 966, 1001, 1015

Goldhaber, G., 274, 280

Goldie, H., 1184, 1195

Goldman, D., 1000, 1016

Goldschmidt, L., 943, 955, 961, 962, 964,
1016, 1024

Goldstein, L., 908-911, 914

Gomez, E. T., 1123

Good, D. J., 784, 822

Goodgal, S. H., 545-547, 579, 600, 610,
623

Goodman, C., 1178, 1179, 1194

Goodrich, J. P., 936, 940, 945, 950

Goodspeed, T. H., 362, 389, 465, 610

Gorbman, A., 993, 1016, 1 167, 1 1 7 1 , 1 1 80,
1195

Gorin, M. H., 307, 311

Goudsmit, S., 198, 252

Gowen, J. W., 329, 345, 520, 610

Gower, J. S., 827, 828, 833, 838

Grachus, T., 986, 1009

Graffeo, A. J., 935, 950, 992, 1016

Graham, J. B., 935, 950, 992, 1016

Graham, R. M., 992, 1016

Grasnick, W., 627, 701

Gray, L. H., 66, 139, 142, 154, 157, 160,
162, 165, 187, 260, 271, 275, 279, 296,
299, 300, 307, 310, 317, 319, 320, 323,
328, 329, 335-337, 339, 340, 344-346,
348, 490, 517, 614, 628, 630, 658, 709,
710, 725, 732, 736, 750, 758, 759, 764,
775-777, 779, 786-788, 805, 810, 819,
823, 923, 928, 951, 954, 957, 976,
1002, 1021, 1146, 1195

Green, C. D., 1172, 1199

Greenberg, D. M., 1112

Grecnstein, J. P., 277, 281, 289, 313, 694,
709, 754, 761, 801, 823

Greisen, K., 97, 143 |

Gricouroff, G., 1094, 1099, 1117, 1118,
1121, 1122, 1124, 1132, 1157, 1195

1212

RADIATION BIOLOGY

Griffen, A. C, 424, 465

Griffith, H. D., 319, 333, 349, 478, 626

Griffith, J. Q., Jr., 926, 950, 968, 970, 976,

1016, 1017, 1022, 1072
Grimmett, L. G., 781, 790, 823
Grohman, A., 894, 896, 898
Grodner, R., 561
Grumbrecht, P., 1132
Grundhauser, W., 930, 955, 1159, 1160,

1177, 1184, 1197
Griissner, G., 980, 1009
Gudernatsch, J. F., 864, 879, 915
Guerin, M., 1157, 1200
Guilleminot, M. H., 554, 610
Gulbekian, C, 508, 516, 610
Gump, H., 1001, 1010
Gunz, F. W., 305, 312, 962, 1020
Gurney, R. W., 165, 187
Gurtner, P., 291, 312
Gustafson, G. E., 1002, 1003, 1016
Gustafsson, A., 275, 279, 436, 442, 465,

555, 556, 563, 575, 576, 578, 584, 590,

591, 606, 608, 610, 754, 758
Guyenot, E., 390, 465

H

Haas, E., 249, 252

Haas, F., 303, 313, 314, 417, 473, 537,

544, 548-551, 561, 562, 566-568,

610, 623, 625, 751, 761
Haba, G. de la, 573, 615
Haddox, C. H., 417, 473, 548-551, 625,

751, 761
Haerting, F. H., 1177, 1195
Hagen, C. W., Jr., 336, 346, 925, 930,

932-934, 951, 955, 964, 1017, 1031,

1044
Hahn, L., 696, 700
Hahn, P. F., 1060, 1184, 1195
Haines, R. B., 139, 143, 299, 301, 312,

319, 320, 330, 331, 347
Haissinsky, M., 256, 280
Halberstaedter, L., 274, 280, 682, 701,

961, 991, 1006, 1097, 1117, 1136,

1137
Haldane, J. B. S., 411, 419, 465, 590, 687,

701, 854, 857
Haley, T. J., 935, 946, 951, 993, 1016,

1031, 1046
Hall, B. V., 943, 951
Hall, C. C, 982, 985, 988, 1000, 1016
Hall, H., 166, 187
Hall, J. J., 979, 1024
Hall, M. E., 164, 189
Hall, T., 1003, 1004, 1024
Hall-Edwards, J., 1148, 1195
Halliday, D., 6, 13, 14, 142

Halpern, O., 166, 187

Hamburgh, M., 907, 915

Hamermesh, B., 117, 142

Hamilton, J., 1112

Hamilton, K., 1187, 1195

Hammond, C. W., 983, 1002, 1003, 1016,

1021
Hance, R. T., 1123
Hannah, A., 498
Hanschuldt, J. D., 972, 1016
Hansen, R. A., 1037, 1067
Hanson, A. O., 89, 112, 141, 142
Hanson, F. B., 346, 389, 465, 478, 479,

582, 610, 869, 876, 879, 894, 896-898,

915
Hardenbergh, E., 972, 974, 1010
Harrar, J. A., 908, 916
Harrington, N. G., 779, 780, 789, 800,

801, 817, 819
Harris, B. B., 393, 465
Harris, D. H., 1031, 1046
Harris, I. D., 967, 1010
Harris, P. S., 943, 948, 1001, 1011
Harris, R. S., 1155, 1183, 1193, 1194
Harris, W., 910, 916
Harrison, B., 287, 288, 308
Harriss, E. B., 172, 187
Hart, E. J., 122, 142, 257, 262, 274, 279,

287, 288, 311, 556, 609
Hartman, F. W., 1000, 1013
Harvey, E. B., 811, 822
Haskins, C. P., 335, 344
Haskins, D., 907, 915
Hatcher, C. H., 1180, 1181, 1195
Hawkins, J. W., 1162, 1195
Hawn, C. van Z., 1003, 1004, 1024
Hayden, B., 559, 610, 940, 951
Hayer, E., 1032
Hays, F. A., 532
Hedin, R. F., 985, 1016
Heiberg, T., 290, 310
Heidenthal, G., 404, 472, 667, 710
Heilbrunn, L. V., 785, 819
Heineke, H., 960, 1002, 1016, 1029, 1039,

1061, 1099
Heitler, W., 98, 143
Hektoen, L., 935, 951, 1002, 1003, 1016,

1080
Helber, E., 864, 900, 916, 1030, 1032
Heifer, R. G., 662, 677, 678, 701
Heller, M., 1109-1116
Hellner, H., 1155, 1195
Helwig, E. R., 628, 665, 701
Hempelmann, L. H., 776, 777, 782, 797,

820, 929, 935-937, 946, 951, 953, 993,

998, 1005, 1016, 1120
Hendlev, D. D., 987, 1014

NAME INDEX

1213

Hennessy, T. G., 943, 951, 962, 1016,
1037

Henry, J. A., 937, 946, 951

Henshaw, C. T., 788, 789, 820

Henshaw, P. S., 274, 280, 301, 311, 335,
337, 346, 647, 709, 769-773, 784, 785,
788-790, 796, 803, 819, 820, 922-925,
928, 929, 935, 939, 951, 996, 1016,
1030, 1031, 1042, 1044, 1049, 1058,
1061, 1063, 1094, 1146, 1160, 1162,
1167, 1177, 1189, 1195, 1199

Henson, M., 686, 701, 833-835, 837, 857

Heptner, M. A., 476, 610

Hereck, 1179

Hereford, F. L., 166, 187

Hermel, M. B., 926, 950, 1072

Hersh, A. H., 589, 610

Hershey, A. D., 587, 610

Herskowitz, I. H., 351, 416, 417, 465, 475,
499, 506, 611, 652, 653, 702

Hertel, E., 811, 820

Hertwig, G., 456, 466, 554, 611, 643, 702

Hertwig, O., 554, 611, 643, 702

Hertwig, P., 423, 456, 466, 554, 611, 627,
643, 686, 687, 702, 825-830, 832-834,
836-849, 851, 852, 857, 858

Herve, A., 275, 280, 941, 945, 948, 951,
990, 1021

Herz, R. H., 147, 185

Herzberg, G., 211-214, 219, 252

Hesse, O., 1150, 1151, 1177, 1195

Hesse, W., 1177, 1195

Heston, W. E., 424, 466, 856, 858, 963,
1020, 1048, 1049, 1054, 1061, 1100,
1107, 1153, 1168, 1173, 1180, 1188,
1190, 1193, 1195, 1198

Heusghem, C, 990, 1021

Hevesy, G. von, 276, 278-280, 305, 306,
308, 311, 692-694, 700, 702, 808, 820,
926, 928, 947, 951, 952, 958, 1017

Hewitt, H. B., 940, 952

Heys, F., 346, 479, 582, 610

Hicks, S. P., 879, 891, 894, 896-898, 901,
904, 915, 933, 952, 996, 1016

Higginbottom, C., 557, 607

Highman, B. J., 935, 956

Higinbotham, N. L., 1153, 1181, 1182,
1193, 1200

Hilcken, J. A., 345, 347

Hill, G. S., 794

Hill, R. F., 544, 611

Hillstrom, H. T., 1112

Himes, M. H., 695, 702

Hine, G. J., 167, 173, 188

Hinkel, C. L., 933, 952

Hinton, C. W., 653, 666, 702

Hinton, O. T., 700

Hinton, T., 416, 466

Hippel, von, 869, 879, 891, 900, 915

Hippie, J. A., 238»., 252

Hirsch, H., 990, 1017

Hirschf elder, J. O., 236, 252

Hobbs, A. A., Jr., 909, 915

Hoberson, J. H., 147, 186

Hochwald, L. B., 935, 936, 947

Hodes, P. J., 970, 976, 1017, 1022

Hodges, F. J., 923, 953

Hodges, G. R. V., 264, 280, 945, 946, 954

Hodges, P. C., 346

Hoecker, F. E., 172, 188

Hoffman, B. G., 87, 141

Hoffman, J. G., 776, 777, 782, 820, 998,
1005, 1016

Holden, C., 707

Holden, W. D., 966, 968, 1017

Hollaender, A., 127, 131, 143, 277, 281,
289, 301, 311, 313, 333, 349, 444, 466,
471, 476, 503, 505, 506, 509, 521, 531,
532-536, 540, 543, 546, 556-558, 561,
563, 564, 569-571, 573, 578-580, 601,
605, 606, 608, 611, 613, 620, 622, 623,
675-677, 694, 703, 709, 738-740, 744,
754, 758, 759, 760, 761, 774-776, 779,
782, 791, 792, 794, 800, 811-815, 817,
818, 822, 823, 856, 858, 940, 941, 944,
952

Hollander, W., 849, 859

Hollcroft, Joanne, 336, 346, 1146, 1195

Hollingsworth, J. W., 1004, 1017

Holmes, B. E., 262, 275, 276, 280, 300,
305, 306, 308, 310, 312, 692, 693, 702,
928, 944, 952, 975, 1017

Holmes, G. W., 995, 1017

Holt, M. W., 805, 823

Holthausen, C. F., 414, 470

Holthusen, H., 180, 187, 554, 558, 611,
627, 702, 927, 935, 939, 952

Holweck, F., 301, 312, 330, 331, 346, 349

Holzknecht, G., 147, 187

Honerjager, R., 25, 143

Hoover, M. E., 645, 647, 650, 683, 699,
700, 702

Hopkins, D. E., 441, 464

Hopwood, F. L., 291, 312, 773, 776, 823,
1097

Horlacher, W. R., 588, 611

Horn, E. C., 335, 346

Horning, E. S., 424, 464, 1190, 1192

Hornyak, W. F., 59, 143

Horowitz, N. H., 461

Hoth, I., 574, 605

Houlahan, M. B., 532, 540, 606

Howard, A., 544, 546, 549, 567, 611, 830,
858

Howland, J. W., 971, 974, 1002, 1003,
1005, 1009, 1017

1214

RADIATION BIOLOGY

Hubbard, H. S., 12, 141

Hubert, R., 1002, 1017

Hudson, J. C, 347

Hueck, W., 1195

Hueper, W. C, 986, 1017, 1164, 1166,

1179, 1188, 1195
Huff, R. L., 926, 943, 951, 957, 962, 1016,

1037, 1136
Hughes, A. F., 776, 782, 797, 824
Hughes-Schrader, S., 646, 647, 702
Huguenin, R., 1184, 1195
Hummel, R., 975, 1017
Humphreys, E. M., 986, 1023
Humphries, R. E., 1158, 1198
Hungate, F. P., 585, 612
Hunt, H. B., 1112
Hursh, J. B., 992, 1017, 1178, 1196
Hurwitz, H., Jr., 116, 143
Hussey, R. G., 299, 312
Husted, L., 714, 758
Hutchinson, W., 148, 187
Hutner, S. H., 595, 620

Ickowicz, M., 991, 1006, 1136, 1137

Impiombato, G., 988, 1017

Inda, F. A., 1164, 1166, 1174, 1198

Inglis, K., 1128

Ingraham, L. P., 999, 1009

Ingram, M., 965, 1017, 1031, 1048

Insch, G. M., 165, 166, 185

International Commission on Radio-
logical Units, 150, 151, 187

International Critical Tables, 118, 143

International Recommendations on Ra-
diological Protection, 1148, 1196

Isaacs, R., 1034

Isotopes Catalogue, 14, 26, 143

Ives, P. T., 414, 416, 466, 588

Ivy, A. C, 985, 986, 1017, 1127, 1132

Izzo, M. J., 961, 1012

Jackson, E. M., 942, 943, 945, 954, 966,
993, 1001, 1022, 1025, 1060, 1067

Jackson, M. A., 1184, 1195

Jacobson, L. O., 337, 346, 926, 930, 932,
935, 952, 955, 960-975, 980, 981, 983,
984, 989, 995, 1002, 1004, 1008, 1010,
1017, 1018, 1020, 1021, 1022, 1024,
1025, 1026, 1029-1031, 1034-1038,
1040-1042, 1044-1046, 1055-1065,
1069-1080, 1100, 1103, 1107, 1137

Jacques, P., 1180, 1196

Jacquez, J. A., 925, 932, 933, 952

Jaffe, G., 194n.

Jaffe, H. L., 1181, 1196

Jagie, N. von, 1162, 1196

James, H. M., 216, 252

Janeway, C. A., 1003, 1004, 1024

Janossy, L., 143

Janzen, A., 996, 1018

Jasny, G. J., 827, 828, 838

Jelatis, D. G., 985, 1016

Jenkinson, E. L., 985, 1018

Jennings, F. L., 934, 952, 986, 1005, 1018

Jensen, K. A., 416, 466, 547, 550, 577,

612, 751, 758
Jentschke, W. K., 148, 187
Jentzer, A., 1155, 1185, 1196
Jesse, W. P., 66, 143, 165, 187
Job, T. T., 864, 865, 869, 870, 873-874,

877-883, 886, 891-894, 896, 901,

915
Joel, C. A., 686, 710
Johannsen, W., 391, 466
John, E. S., 971, 990, 991, 994, 1001, 1022
Johnson, C. G., 934, 952, 1005, 1018
Johnson, P., 287, 288, 309
Jolles, B., 926, 952, 976, 977, 1018, 1136
Jollos, V., 582, 612
Jolly, J., 927, 939, 940, 945, 952
Jones, D. C, 935, 953, 1005, 1018
Jones, H. B., 928, 953, 978, 988, 993,

1012, 1014, 1018
Jones, H. W., Jr., 908-909, 915
Jonkhoff, A. R., 1152, 1196
Jordan, H. C, 888, 918, 1185, 1201
Jordan, M. L., 969, 1011
Jovin, J., 776, 822
Judge, J. T., 268, 280, 703
Jiingling, O., 776, 777, 782, 820
Just, E. E., 357, 466
Juul, J., 776, 782. 799. 811. 820, 1097

K

Rabat, E. A., 1005, 1014

Kahlau, G., 1178, 1180, 1199

Kahn, J. B., Jr., 937, 952, 1039

Kaliss, N., 1190, 1194

Kalnitsky, G., 307, 309

Kalter, H., 907, 915

Kamen, M. D., 587, 610

Kanellis, A., 481, 484, 565, 576, 605, 606,

612, 670, 671, 674, 698, 699, 702
Kanoky, J. P., 994, 1018
Kaplan, H. S., 926, 933, 947, 952, 991,

1005, 1018, 1020, 1164-1166, 1168,

1170, 1174, 1188, 1196
Kaplan, I. I., 999, 1000, 1018
Kaplan, R. W., 485, 537, 538, 541, 542,

555, 556, 568, 569, 575, 576, 591, 612,

614

NAME INDEX

1215

Kaplan, W. D., 417, 466

Karadv, S., 1000, 1016

Karnofsky, D. A., 925, 932, 933, 952, 953

Karr, J. W., 864, 869, 872-873, 876, 879,
882, 887-888, 891-892, 901-904, 907,
918, 932, 933, 957, 996, 1028

Karrer, E., 589, 610

Kasha, M., 227, 252

Kassabian, M. K., 1149, 1196

Katz, E. J., 961, 1024

Kaufmann, B. N., 570, 624

Kaufmann, Berwind P., 135, 143, 268,
280, 378-380, 463, 466, 483, 495, 503,
518, 521, 532-534, 569-571, 578, 604,
606, 613, 627, 631, 632, 635, 645-648,
650-657, 659-666, 668-671, 673,
675-681, 683, 685, 695-697, 699, 700,
702, 703, 738-740, 758, 759

Kausche, G. A., 510, 613

Kaven, A., 864, 869, 872, 874, 875, 879,
881-886 890-894, 896, 898, 901-902,
915

Kaye, G. W. C, 153, 187

Keating, R. P., 939, 956, 969-972, 974,
1026

Keith, C. K., 937, 949

Keller, M., 291, 312

Keller, M. E., 982, 985, 1023

Kelly, E. M., 388, 464, 683, 699, 838, 929,
949

Kelly, K. H., 994, 1018

Kelly, L. S., 928, 953, 988, 1018

Kelner, A., 543, 544, 613

Kelsall, M. A., 967, 1018

Kemp, T., 776, 782, 799, 811, 820, 1097

Kennard, E. H., 6, 7, 11, 13, 143

Kennedy, J. W., 587, 610

Kennedy, R. J., 108

Kenney, R. W., 97, 141

Kerkis, J. J., 396, 414, 466, 508, 588, 613

Kerr, H. D., 998, 1019

Kerst, D. W., 112, 141

Kersten, H. P., 299, 314

Khvostova, V. V., 378, 471, 480, 481, 607,
613

Kihlman, B., 578, 613

Kikkawa, H., 647, 703

Kile, J. C, Jr., 561, 620, 827, 828, 858

Kimball, R. F., 545, 564, 572, 580, 613

Kimeldorf, D. J., 935, 953, 1005, 1018

Kimura, S. J., 998, 1011

Kindler, K, 1172, 1196

King, E. D., 562, 565, 576, 614, 674, 704

King, R. C, 584, 585, 614

Kingsbury, A. N., 1002, 1021

Kinst, M., 403, 427, 472

Kirby-Smith, J. S., 137, 488, 614

Kirk, I., 416, 466, 547, 550, 577, 612, 751,

758
Kirk, M., 1003, 1004, 1024
Kirschbaum, A., 999, 1020, 1165, 1170,

1188, 1196, 1199
Kirschner, L. B., 993, 1001, 1018
Kirschon, A., 966-969, 1008
Kirsner, J. B., 986, 1023
Kirssanow, B. A., 480, 614
Kirwan, D. P., 538, 593, 608
Kisieleski, W. E., 923, 956, 1160, 1197
Klot, B. von, 864, 898, 908, 910, 916
Knapp, E., 476, 531, 540, 553, 555, 556,

591, 614
Knef, J. P., 1182, 1198
Knipling, E. F., 441
Knowlton, N. P., Jr., 681, 704, 776, 777,

779, 780, 782, 797, 820, 929, 935-937,

946, 961, 963, 993, 1016, 1120
Knuchel, H., 291, 312
Koch, H. W., 112, 141
Koch, J., 910, 918
Koerner, L., 480

Koernicke, M., 554, 614, 627, 704
Kohn, H. I., 935, 963, 966-969, 972-976,

979, 1004, 1018, 1019, 1027
Koletsky, S., 1002, 1003, 1016, 1160,

1182, 1196
Koller, P. C, 415, 466, 577, 614, 629, 660,

662, 678, 681, 686, 693, 699, 704, 722,

749, 769, 774, 799, 804, 820, 843,

848, 858
Kolmark, G., 416, 466, 547, 550, 577,

612, 751, 758
Komuro, H., 274, 280
Kornblum, K., 970, 980, 982, 1000, 1021
Kosaka, S., 864-866, 869, 872, 874, 876,

879-880, 888, 891, 893, 894, 896-898,

901, 904, 915
Kossikov, K. V., 409, 410, 466, 481, 522,

614
Kostoff, D., 573, 589, 614
Kotval, J. P., 488, 490, 517, 614, 724, 732,

736, 759
Kotz, J., 994, 1019
Koza, R. W., 801, 819
Kraemer, 908-909, 915
Kraemer, E. A., 293, 313
Krause, P., 554, 614, 627, 688, 704, 982,

1019, 1151
Krebs, C., 1164, 1178, 1196
Krejci, L. E., 293, 313
Krenz, F. H., 269, 280
Kretzschmar, C. H., 975, 1019
Krivshenko, J. D., 664, 704
Krizek, H., 988, 1019
Kroemeke, F., 1061
Krotkina, N., 1155, 1156, 1199

1216

RADIATION BIOLOGY

Kruger, P. G., 148, 187, 998, 1008
Krumpel, O., 292, 313
Kubowitz, F., 249, 252
Kuck, Kathryn D., 333, 350
Kiihn, A., 461
Kuntz, H., 629, 707
Kurnick, N. B., 695, 704
Kustner, H., 180, 187

LaBarre, J., 991, 1028

Lacassagne, A., 301, 312, 331, 346, 776,
822, 869, 864, 893-895, 898, 915, 916,
917, 936, 940, 945, 953, 982, 1023,
1029, 1094, 1099, 1117, 1118, 1121,
1122, 1124, 1132, 1145, 1152, 1153,
1155, 1161, 1162, 1165, 1177, 1182,
1183, 1185-1187, 1190, 1191, 1193,
1196, 1197

Lacomme, M., 909, 916

La Cour, L. F., 717, 721, 723, 741, 749,
757

Laidler, K. J., 193, 205n., 206, 223, 252

Laird, A. K., 929, 955

Lamerton, L. F., 172, 187, 934, 950

Lampe, L, 334, 339, 350

Lampe, J., 923, 953

Lams, H., 865, 866, 869, 879, 893, 894,
898, 916

Lamy, R., 388, 466, 683, 704

Landolt-Bornstein, 229, 252

Lane, G. R., 386, 467, 483, 485, 486, 516,
571, 572, 615, 727, 729, 730, 759

Langendorff, H., 331, 332, 345, 346, 657,
704, 776, 777, 782, 820

Langendorff, M., 332, 346

Langohr, J. L., 998, 1024

Lanzl, E. F., 112, 141

Lanzl, E. R., 923-925, 955, 982, 985, 1023

Lanzl, L. H., 89, 112, 141, 142

Laqueur, G., 811, 816, 822

Larkin, J. C, 338, 349, 976, 977, 996, 1019

Larsh, A. E., 148, 186

Larson, E., 990, 1014

Lartigue, O., 969, 974, 1020

Lasnitski, L, 319n., 335, 346, 629, 682,
704, 773, 776, 782, 787, 788, 791,
797, 804-806, 809, 810, 820, 821,
923, 924, 928, 953
Laszlo, D., 305, 312

Latarjet, A., 274, 280, 302, 312, 331, 345
Latarjet, R., 549, 683, 704, 929, 953, 1 190,

1197
Latchford, W. B., 1002, 1014
Langhlin, J. S., 112, 141, 146, 187
Laurence, G. C., 154, 155, 157, 166, 183,
187, 188

Lauritsen, T., 59, 143

Lavedan, J., 630, 704, 895, 916, 1029

Lawen, A., 1003, 1019

Lawrence, A. H., 1037, 1039, 1044, 1063

Lawrence, E. G., 650, 705

Lawrence, E. O., 336, 339, 347, 923, 953,

961, 1019, 1031, 1041, 1042
Lawrence, G. H., 990, 1019
Lawrence, J. H., 320, 330, 336, 339, 343n.,

344, 347, 923, 935, 947, 957, 961, 983,

1002, 1019, 1031, 1035, 1041, 1042,

1066, 1079, 1100, 1132
Lawrence, J. S., 926, 953, 960, 963, 966,

970, 978, 1008, 1019, 1027, 1035
Lazarew, N. W., 977, 1019
Lazarewa, A., 977, 1019
Lazarus-Barlow, W. S., 1127, 1151, 1153-

1156, 1197
Lea, D. E., 81, 130, 135, 137-141, 143,

180, 187, 194, 197, 252, 262, 263, 271,

280, 283, 299, 301, 312, 318-320, 323,

327, 329-331, 335, 342, 344, 346, 347,
479, 482, 487-491, 499, 500, 517, 521,
526-530, 542, 566, 567, 605, 615, 630,
632, 646, 647, 652, 653, 658, 660, 663,
666, 667, 675, 679, 685, 687, 698, 701,
704, 713, 714, 716, 717, 720, 724,
726-728, 730-732, 735, 741, 750, 755,
756, 757, 759, 763, 770, 788, 807, 821,
825, 835, 836, 858, 921, 924, 935, 953,
1147, 1189, 1197

Leach, J. E., 981, 1019

Le Bail, H., 256, 280

Leblond, C. P., 960, 991, 993, 1014, 1018,

1136, 1137, 1162, 1197
LeCalvez, J., 628, 646, 674, 697
Lecomte, J., 941, 945, 948, 1004, 1019
Leddy, E. T., 1175, 1197
Lederberg, E. Z., 420, 454, 467, 584, 609
Lederberg, J., 420, 454, 467
Lefevre, George, Jr., 654, 704
Lefort, M., 256, 274, 278, 280, 286, 309,

328, 344, 567, 604, 744, 750, 751, 757,
938, 948

Leibold, G. J., Jr., 879, 882, 915

Leighton, P. A., 193, 253

Leinfelder, P. S., 998, 1019

Leitch, J. L., 1161, 1174, 1197

LeMay, M., 937, 953, 989, 1005, 1014,

1020
Lengfellner, K, 894, 916
Lennihan, M. R., 268, 280, 703
L6obardy, J. de, 895, 916
Leppin, O., 919, 953
Lerche, W., 667, 687, 697
Leuchtenberger, C., 695, 707
Levan, A., 584, 608
LeVine, H. D., 176, 185

NAME INDEX

1217

Levine, M., 864, 899, 916

Levinson, L. J., 910, 918

Levit, S. G., 450, 467

Levitsky, G. A., 362, 363, 467, 480, 615

Levy, 0., 553, 554, 615

Lewis, D., 553, 566, 568, 615

Lewis, E. B., 650, 704

Lewis, G. N., 581, 619

Lewis, M., 53, 54, 56, 64, 87

Lewis, M. R., 775, 821

Lewis, W. H., 775, 821

Lewitsky, G. A., 799, 821

Li, M. H., 1168, 1197

Lick, L., 999, 1020, 1170, 1197

Liebmann, H., 328, 346

Liebow, A. A., 999, 1005, 1020, 1101

Liebowitz, J., 961, 1016

Liechti, A., 335, 347, 961, 1020

Light, A. E., 977, 1020

Limperos, G., 694, 705, 755, 759, 945, 953

Lindegren, C. C., 476, 533, 615

Lindegren, G., 476, 533, 615

Lindenfeld, B., 908, 916

Lingley, J. R., 345, 347

Linser, P., 864, 900, 916, 1030, 1032

Lisco, H., 932, 955, 961, 964, 967, 969-
975, 984, 989, 995, 998, 1000, 1001,
1005, 1016, 1023, 1159, 1160, 1166,
1177, 1180, 1184, 1192, 1197

Lison, L., 695, 705, 707

Little, C. C., 355, 390, 467, 469

Little, E. P., 274, 279, 301, 311, 785, 818

Lively, E., 420, 454, 467

Livingston, M. S., 89, 143

Livingston, R., 227, 233, 249, 252, 270,
279

Lloyd, 1148, 1197

Locher, G. L., 389, 470, 520, 618

Lockwood, L. B., 444, 467

Loeb, J., 390, 467

Loebl, H., 291, 312

Loevinger, R., 168, 169n., 170, 182, 187

Lohmann, A., 296, 313

Loiseleur, J., 274, 280, 289, 296, 312, 942,
946, 947, 953

London Conference, 630, 705

Long, C. N. H., 991, 1024

Loomis, A. L., 589, 610

Loos, G. M., 811, 817

Lorenz, E., 336, 346, 829, 830, 847, 856-
858, 926, 953, 960, 961, 963-965, 967,
991, 993, 1001, 1002, 1017, 1020,
1022, 1031, 1035-1037, 1040, 1045,
1048-1050, 1054-1056, 1061-1064,
1074-1076, 1146, 1153, 1154, 1165-
1168, 1170, 1173, 1174, 1177, 1180,
1184, 1188, 1195, 1198

Lotz, C., 417, 464, 550, 607, 752, 757

Lourau, M., 968, 974, 1020

Louviere, L. J., 1061

Love, W. H., 773, 821

Low-Beer, B. V. A., 975, 994, 1018, 1020

Ludewig, S., 300, 312, 975, 987, 1020

Ludin, M., 1152, 1198

Liiers, H., 542, 615

Luippold, H., 486, 621, 721, 730, 760

Lukens, R. M., 993, 1020

Lumb, G., 1158, 1198

Liming, K. G., 477, 483, 504, 509, 510,

513, 514, 520, 552, 570, 604, 615, 644,

697
Lnria, S. E., 262, 280, 330, 349, 527, 615
Luther, W., 776, 821
Lutwak-Mann, C., 305, 312, 962, 1020
Lyman, C. M., 305, 312
Lyman, E. M., 89, 142

M

McAulay, A. L., 538, 541, 593, 615
McCarthy, J. E., 985, 986, 1017
McClintock, B., 362, 363, 365, 366, 451,

467, 646, 675, 705
MacComb, W. S., 338, 347, 925, 953, 955
MacDonald, E., 1013, 1014, 1024
McDonald, M. R., 268, 269, 280, 656,

695, 696, 703
MacDowell, E. C, 906, 914
MacDuffee, R. C, 935, 955, 965, 1024
MacDuffee, R. S., 1065
McElroy, W. D., 547, 573, 574, 597, 615,

624
McGregor, J. H., 554, 616
Macht, S. H., 967, 1020
MacKee, G. M., 925, 953
Mackenzie, K, 532, 540, 616, 618
MacKey, J., 591, 610
MacMahon, H. E., 1157, 1184, 1198
McMaster, R., 765, 814, 816, 818
MacMillan, J. C, 686, 710, 983, 1027
MacNeal, W. J., 1151, 1198
McQuarrie, L, 988, 989, 1021
McQuate, J. T., 532, 533, 616, 625
Maddox, M. N., 973, 1024
Magee, J. L., 194, 252
Mahoney, J. F., 997, 1022
Maier-Leibnitz, H., 5, 40, 72, 82, 83, 113,

142
Mainx, F., 668, 705
Maisin, J., 1158, 1198
Major, M. H., 833, 838, 869, 877, 879,

888, 903-905, 917
Makhijani, J. K., 480, 483, 565, 616, 655,

671, 673, 705
Makino, S., 642, 705

1218

RADIATION BIOLOGY

Makki, A. I., 363, 380, 469, 480, 481, 494,

501, 618, 670
Maldawer, M., 555, 622, 690, 709, 722,

760
Mallet, L., 347, 782, 821
Mann, S., 935, 946, 951, 993, 1016
Mannell, T., 585, 612
Manollov, S. E., 299, 312
March, H. C, 1163, 1198
Marchbank, Dorothy F., 333, 360
Marcuse, W., 1148, 1198
Marder, S. N., 991, 1005, 1018, 1020,

1164, 1165, 1196
Marie, P., 1150, 1152, 1153, 1198
Marinelli, L. D., 135, 142, 153, 154, 157,

167-170n., 173, 182, 183, 185-189,

343n., 349, 484, 485, 616, 618, 727,

759
Mar-Kham, R., 312
Markham, R., 262, 280
Marks, E. K, 337, 346, 926, 930, 935,

952, 960, 961, 963-965, 967, 983,

1002, 1004, 1017, 1018, 1030, 1035,

1036, 1038, 1040-1042, 1044-1046,

1056, 1061-1064, 1069-1074, 1076-

1080
Marquardt, H., 484, 553, 616, 776, 799,

801, 804, 821
Marshak, A, 305, 312, 338, 341, 347, 575,

616, 661, 669, 684, 705, 775, 776, 798,

801, 811, 813, 821
Marshak, R. E., 116, 143
Marti, N. F., 926, 955
Martin, C. L., 985, 986, 990, 999, 1020,

1127
Martin, F. L., 131, 143, 301, 311, 333,

349, 506, 521, 558, 579, 611, 622, 940,

941, 944, 952
Martin, G. J., 979, 1009
Martin, S. F., 998, 1011
Martland, H. S., 964, 1000, 1020, 1099,

1158, 1159, 1178, 1181-1183, 1198
Mason, W. B., 965, 1017, 1031, 1048
Massey, H. S. W., 193, 202n., 205-207,

211, 236, 237, 239, 240, 243, 252, 253
Mather, K, 716, 721, 759
Matoltsy, G., 577, 608
Matthews, J. J., 937, 946, 954
Mavor, J. W., 358, 359, 467, 508, 616,

644, 705
Maximow, A., 1101, 1121
Maxwell, R. D., 939, 956, 969-972, 974,

1026
Mayer, E., 811, 821
Mayer, M. D., 910, 916
Mayer, M. M., 1004, 1013
Mayer, S. H., 937, 942-946, 954, 966,
1022

Maynard, R. M., 998, 1028

Mayneord, W. V., 146, 174, 185, 188, 318,

347, 1061, 1187, 1190, 1192, 1193,

1198
Mazia, D., 601, 616, 928, 953
Mead, J. F., 985, 986, 1009, 1021
Medvedev, N. N., 565, 574, 616
Meek, G. E., 1161, 1194
Meredith, W. J., 259, 260, 262, 263, 271,

272, 275, 279, 296, 299, 300, 310, 329,

344, 656, 699
Merrill, O. E., 998, 1024
Meschan, I., 937, 949
Metcalf, R. G., 1164, 1166, 1174, 1198
Metz, C. W., 650, 662, 665, 687-690, 705
Meyer, H. U., 535, 536, 540, 545, 546,

569, 602, 607, 616
Mickey, G. H., 565, 616, 642, 673, 705
Miescher, G., 976, 985, 1021, 1120
Miletzky, O., 926, 948, 963, 1010
Milham, W., 966-989, 1008
Miller, B. L., 172, 188
Miller, C. P., 983, 1002, 1003, 1016, 1021
Miller, E., 829, 837, 838, 857, 999, 1001,

1014
Miller, G. L., 973, 1024
Miller, H., 547, 574, 624
Miller, J. H., 926, 955
Miller, J. R., 908, 916
Miller, N., 270, 279, 280
Miller, W. R., 985, 1016
Miller, Z. B., 307, 309
Minder, W., 291, 312, 335, 347, 577, 606
Minot, G. R., 1029
Minowitz, W., 182, 187
Mitchell, J. R., 935, 956
Mitchell, J. S., 163, 188, 276, 280, 336,

338, 347, 692, 693, 705, 706, 785, 809,

821, 924, 928, 954
Mitchell, R. G., 976, 1018
Miwa, M., 771, 772, 785, 787. 789, 821,

822 824
Mixer, H. W., 999, 1020, 1165, 1170,

1188, 1196, 1197, 1199
Miyamoto, S., 1157, 1199
Miyazaki, K., 988, 1021
Mobius, W., 910, 916
Moelwyn-Hughes, E. A., 272, 280
Mohler, F. L., 148, 188
Mohler, J. D., 532, 608
Mohney, J. B., 992, 1017
Mohr, O. L., 358, 368, 467, 627, 644, 706
Mole, R. H., 264, 280, 945, 946, 954
Moll, F. C., 1003, 1004, 1024
Mollendorf, W. V., 811, 816, 822
Momigliano, E., 865, 869, 894, 916
Montgomery, H., 1175, 1176, 1200
Montgomery, P. O., 980, 981, 1021

NAME INDEX

1219

Moon, V. H., 970, 980, 982, 1000, 1021

Moore, C. E., 198, 253

Moore, D. E., 1003, 1004, 1011

Moore, D. H., 1004, 1013

Moore, M. C, 970, 971, 973-976, 978,
980, 981, 996, 1022

Moore, W. G., 513, 616

Mora, J. M., 1199

Morgan, D. R., 970, 980, 982, 1000, 1021

Morgan, L. V., 361, 467

Morgan, T. H., 390, 467, 638, 643, 698,
706

Mori, K., 771, 772, 785, 787, 789, 821,
822, 824

Morris, H. P., 1172, 1199

Morrison, P., 59, 143

Morrow, N. M., 182, 188

Morse, M. L., 579, 605

Morse, S., 257, 258, 279, 345

Moser, H., 600, 603

Moses, J. J., 723, 760

Moses, M. J., 695, 706

Moses, S. G., 811, 812, 816, 817

Mosher, W. A., 694, 705, 755, 759, 945,
953

Moshman, J., 770, 796, 818

Mott-Smith, L. M., 581, 618

Mottram, J. C, 275, 281, 336, 337, 345,
348, 553, 558, 616, 773, 776-777, 779,
782, 786, 819, 822, 923, 927, 929, 936,
939, 940, 945, 954, 976, 1021, 1030,
1158, 1178, 1187, 1190, 1199

Moulder, P. E., 966-968, 1003, 1008

Moulder, P. V., 1045

Moyer, A. J., 444, 467

Moyer, B. J., 12, 142

Midler, J. H., 335, 347, 348

Muller, H. J., 135, 136, 143, 334, 348, 355,
359, 362, 363, 365, 366, 377, 378,
380-382, 385, 388, 389, 391-393,
395-397, 399-401, 403, 405, 409,
411-414, 418, 419, 422, 423, 427, 430,
431, 435, 438, 446, 448, 451-453, 455,
456, 458, 460, 462, 463, 466 471, 476,
478-484, 487, 488, 492-495, 497-503,
506, 509, 511, 513-515, 519-523, 526,
527, 529, 532, 535, 536, 538, 540, 546,
552, 563, 565, 569, 581-584, 588, 596,
598, 600, 601, 602, 604, 614, 616-618,
625, 629, 630, 632, 635, 644, 646, 647,
650, 653, 655, 658, 661, 663, 664, 666,
668, 670, 671, 673, 676, 683, 687, 704,
706-708, 854, 858, 1190, 1199

Munick, R., 168, I70n., 188

Munoz, C. M., 943, 957

Muntz, J. A., 262, 278, 298, 299, 309,
551, 580, 603, 922, 941, 948, 973, 984,
986, 1001, 1009, 1021

Miintzing, A., 683, 706
Murphy, A. S., 1157, 1184, 1198
Murphy, D. P., 864, 879, 891, 901, 908-

911, 914, 916, 994, 1015
Murphy, E. D., 1174, 1196
Murphy, J. B., 1003, 1021, 1064, 1123
Murray, J. M., 687, 706
Murray, R., 961, 1021, 1031
Murray, R. G., 1104, 1106-1108, 1118
Mutscheller, M., 925, 953

N

Nadson, G. A., 390, 470

Nagai, M. A., 389, 470, 520, 618

Narat, J. K., 990, 1021

National Bureau of Standards, 179, 188

National Research Council Preliminary

Report No. 8, 175n., 188
Naumenko, V. A., 417, 470
Neary, G. J., 164, 188
Nebel, B. R., 484, 485, 616, 618, 727, 759,

811, 814, 822
Neel, J. V., 414, 430, 470
Neill, W., Jr., 908-909, 915
Nelms, A. T., 143
Nelson, L., 304, 309
Nelson, W. O., 1132
Nemours, A., 1184, 1195
Neuberg, C., 297, 313
Neufeld, J., 166n., 188
Neuhaus, M. E., 455, 470, 514, 618
Neuhaus, M. J., 654, 706
Neumann, F., 869, 894, 900, 914
Neve, R. A., 975, 1021
Newcombe, H. B., 545, 572, 591, 592,

595, 618, 724, 759
Newcomer, E. N., 589, 625, 737, 761
Nichols, H. J., 182, 190
Nickson, J. J., 952, 1011
Nickson, M. J., 977, 1010
Nims, L. F., 987, 990, 1021
Nix, M., 561, 609, 796, 818, 904
Nizet, E., 990, 1021
Nobele, de, 865, 866, 869, 879, 893, 894,

898, 916
Noethling, W., 476, 531, 540, 618, 623
Nogier, T., 863, 864, 917, 982, 1023, 1124
Norby, B., 558, 622
N0rgaard, F., 1158, 1199
Norrish, R. G. W., 193n., 227, 253
North, N., 987, 990, 1021
Northrop, J. H., 299, 313
Novick, A., 418-420, 470, 537, 543, 545,

619
Novick, R., 995, 1011, 1021
Novitski, E., 403, 427, 472, 565, 619
Noyes, W. A., Jr., 193, 253

1220

RADIATION BIOLOGY

Nurnberger, C. E., 271, 281, 296, 313,

328, 348
Nurnberger, L., 869, 894, 897-898, 916
Nybom, N., 590, 610
Nyka, W., 1155, 1183, 1185, 1197

O

Oak Ridge Symposium, 630, 707

Oakberg, E. F., 829re.

Oakley, H. E. H., 554, 623, 776, 799, 805,

823
Oberling, C, 1157, 1200
O'Connor, M. H., 1179, 1191
O'Connor, R. J., 932, 954
Odclie, T. H., 168, 169, 188
Oehlkers, F., 416, 470
Offermann, C. A., 479, 497, 552, 619, 650,

707
Ogur, M., 695, 707
Oldenberg, O., 220, 253
Oliver, C. P., 476, 588, 619, 621, 652, 707
Olson, A. R., 389, 465, 581, 619
Onufrio, O., 1167, 1199
Oosterkamp, W. J., 180, 188
Oppermann, K., 554, 619
Ordway, T., 1150, 1199
Orndoff, B. H., 985, 986, 1017
Ortega, P., 994, 1018
Osborne, J. W., 982, 985, 1023
Osgood, E. E., 339, 348, 1029, 1039
Oster, R. H., 1000, 1021
Otis, E. M., 849, 858, 909, 916
Ovadia, J., 146, 187
Owen, R., 848, 859
Owen, S. E., 1174, 1199

Packard, C, 300, 313, 348, 763, 784, 804,
822, 929, 935, 954, 1002, 1022, 1094

Packham, Evelyn A., 339, 348

Pagenstecher, H. E., 869, 879, 891, 898,
900, 915, 916

Painter, E. E., 932, 955, 961, 964, 967,
969-981, 984, 989, 995, 996, 1000,
1001, 1005, 1022, 1023, 1027, 1037

Painter, T. S., 363, 378, 446, 469, 470,
635, 647, 707

Palmer, W. L., 986, 1023, 1125

Panofsky, W. K. H., 97, 141

Panschin, I. B., 381, 470, 480, 493, 522,
619, 668, 707

Panschina, A. N., 493, 619, 668, 707

Papalashwili, G., 565, 619, 673, 707

Pappenheim, A., 1099

Park, G. S., 309

Parker, D. R., 654, 707

Parker, E., 994, 1019

Parkes, A. S., 832, 834, 835, 837, 839,

840, 856, 858, 864-866, 869, 870, 894,

896-898, 916, 999, 1022, 1119, 1168,

1192
Parrish, Mary E., 331, 334, 343m., 348, 350
Parsons, L. D., 1061, 1187, 1198
Passonneau, J., 1187, 1195
Pasteels, J., 695, 705, 707
Patau, K., 527
Paterson, E., 348, 657, 707, 788, 796, 819,

822, 925, 937, 946, 954
Patt, H. M., 264, 281, 308, 313, 930, 935-

939, 942-946, 953, 954, 956, 957, 965,

966, 971, 977, 979, 984, 985, 990-994,

1001, 1005, 1022, 1025, 1026, 1067
Patterson, J. T., 359, 393, 399, 455, 458,

470, 478, 514, 516, 526, 619, 641, 644,

650, 653, 689, 707
Patterson, P. A., 932, 933, 953
Pauli, W., 295, 311
Pauling, L., 32, 143, 250, 295, 313
Payne, F., 627, 707
Pearce, M. L., 962, 963, 1027, 1035, 1037,

1065
Pearlman, N., 337, 348, 966, 1025, 1044
Pekarek, J., 776, 782, 822
Pelc, S. R., 172, 188, 830, 858
PeUer, S., 1177, 1199
Pemberton, R., 1000, 1013
Pendergrass, E. P., 968, 976, 994, 997,

1015, 1016, 1022
Penick, G. D., 966, 968, 1022
Pennoyer, J. M., 268, 280, 703
Penrose, L. S., 461
Perje, A. M., 477, 604, 644, 697
Perlberg, H., Jr., 967, 1020
Perrot, M., 782, 821
Perry, S. P., 346
Perryman, R., 968, 1016
Perthes, G., 627, 707, 1094, 1112
Peter, O., 519, 606, 669, 670, 698
Petersen, B. W., 345
Peterson, S. C, 995, 1011
Petrakis, N. L., 692, 693, 707
Petrov, N., 1155, 1156, 1199
Petry, E., 927, 939, 955
Peyrou, P. P., 493, 619, 668, 707
Pfuhl, W., 629, 707
Philip, U., 361, 470
Philippov, G. S., 390, 470
Phillips, J. T., 291, 312
Philpot, J. St. L., 264, 280, 945, 946, 954
Picken, D., 849, 859

Pickering, B. I., 943, 965, 962, 964, 1024
Pickhan, A., 478, 572, 588, 619
Pierce, M., 961, 983, 1021, 1022, 1096,

1120, 1124, 1125, 1127

NAME INDEX

1221

Pinson, E. A., 964, 1026, 1075

Pintner, I. J., 595, 620

Planck, 18, 30

Platzman, R. L., 64, 88, 143, 197, 232,

241n., 243, 249, 250, 263
Plesch, J., 1099

Plomley, N. J. B., 538, 593, 615
Plough, H. H., 414, 470
Plunkett, C. R., 400, 471
Podljaschuk, L. D., 1132
Pohle, E. A., 976, 986, 1020, 1022, 1128
Poisson, S. D., 80, 133
Politzer, G., 554, 602, 628, 629, 696,

776, 782, 799, 804, 811, 816, 817, 822,

1097
Pollard, E. C, 139, 143, 330, 348
Pollister, A. W., 695, 707
Pontecorvo, G., 365, 456, 469, 471, 481,

482, 557, 607, 618, 619, 646, 647, 650,

682, 685, 706-708
Poppe, E., 1131
Porter, C. A., 1149, 1199
Porter, G., 193m., 253
Porter, L. M., 961, 988, 1024
Portmann, A. F., 966, 968, 1017
Portmann, U. V., 180, 186
Powers, E. L., Jr., 586, 587, 619
Pratt, A. W., 993, 1001, 1022
Present, R. D., 216, 252
Price, J. M., 929, 955
Price, J. P., 811, 812, 817
Pringsheim, P., 20ln., 202n., 217, 230,

232, 233, 253
Prokofyeva, A. A., 380, 448, 451, 469,

492, 499, 618
Prokofyeva-Belgovskaya, A. A., 377, 378,

448, 496, 471, 493, 495, 618, 619, 664,

706
Promptov, A. N., 531, 620
Prosser, C. L., 930, 932, 955, 961, 964,

966, 967, 969-975, 979-981, 984, 989,

993, 995, 1000, 1001, 1018, 1021,

1022, 1023, 1025, 1031, 1037, 1039,

1044, 1045
Provasoli, L., 595, 620
Prussia, G., 1157, 1199
Pullinger, B. D., 977, 1023
Pullman, E. W., 972, 984, 1022

Q

Quastler, H., 112, 136, 141, 142, 923-925,

933, 934, 955, 982, 993, 995, 1001,

1018, 1023
Quimby, E. H., 86, 109, 142, 143, 167,

173, 176, 180, 182, 183, 186, 188, 338,

347, 348, 925, 953, 955

R

Raab, W., 991, 1023

Rabinowitch, E., 230, 232, 253

Radu, G., 48, 483, 484, 605, 606, 670,

671, 684, 698, 699
Raffel, D., 380, 448, 469, 492, 493, 618,

664, 706
Rajewski, E. V., 294, 313
Rajewsky, B., 1178, 1180, 1199
Rajewsky, B. N., 567, 582, 620, 763, 822
Randolph, L. F., 457, 462
Raper, J. R., 319, 333, 336, 348, 350, 924,

926, 931, 935, 955, 961, 997, 1023,

1030, 1058, 1160, 1177, 1199
Raper, K. B., 444, 466, 467, 471, 540, 611
Rapoport, J. A., 416, 417, 471, 552, 557,

575, 576, 620
Rask-Nielsen, H. C., 1164, 1178, 1196
Raulot-Lapointe, G., 1150, 1152, 1153,

1198
Ray-Chaudhuri, S. P., 385, 471, 478, 480,

487, 620, 642, 671, 708
Raynaud, A., 864, 869, 876-877, 879, 888,

891, 901, 916, 917
Read, J., 139, 142, 275, 281, 306, 314, 320,

328, 335, 336, 339, 340, 345, 346, 348,

349, 557, 558, 567, 599, 624, 656, 710,

736, 744, 751, 761, 764, 775-777, 786,

787, 819, 923, 940, 951, 955, 957,

1178, 1199
Recknagel, R. O., 796, 824
Redd, J. B., 969, 1026
Redfield, A. C., 936, 955
Redfield, B., 286, 309
Redfield, H., 654, 708
Regaud, C., 554, 620, 776, 822, 831, 858,

863, 864, 917, 982, 1023, 1094, 1108,

1114, 1124
Reid, T. R., 926, 953, 964, 965, 1020
Rekers, P. E., 926, 955, 963-969, 979,

1003, 1009, 1014, 1023, 1037, 1075
Remley, M. E., 148, 187
Renyi, M. de, 864, 879, 891, 901, 916
Reuss, A., 331, 332, 340, 345, 346, 531,

540, 614, 620
Reynolds, J. P., 484, 620, 689, 690, 708
Rhoades, R. P., 987, 990, 1023, 1122,

1128
Rice, F. O., 32, 143
Richards, A., 783, 784, 822
Richards, P. I., 168, 188
Richmond, J., 962, 1002, 1008
Richtmyer, F. K, 6, 7, 11, 13, 143
Rick, C. M., 493, 494, 566, 620, 668, 708,

720, 724, 726, 759
Ricketts, W. E., 986, 1023, 1125
Ridgway, L. P., 932, 933, 953

1222

RADIATION BIOLOGY

Riesen, D. E., 112, 141

Rietz, L., 929-931, 948, 965, 1010, 1031,

1128, 1135
Rigdon, R. H., 978, 1023
Rigos, F. J., 1175, 1197
Riley, E. F., Jr., 336, 337, 346, 350, 922-

924, 933, 951, 958, 1030, 1049, 1146,

1160, 1167, 1177, 1195
Riley, H. P., 558, 559, 561, 568, 580, 609,

620, 656, 701, 714, 721, 742-745,

747-749, 752, 758, 759, 940, 950
Ris, H., 646, 674, 702, 708, 723, 759
Risse, O., 257, 281, 284, 313, 328, 345,

348, 531, 540, 614
Robbie, W. A., 936, 950
Robbins, L. R., 998, 1024
Roberts, J. E., 1187, 1193
Robertson, J. E., 1000, 1024
Robinett, P. W., 966-969, 979, 1019
Robinson, C. S., 112, 141
Robinson, J. C, 811, 817
Robinson, J. M., 338, 349
Robinson, M. R., 909, 910, 917
Robinson, Robert, 461
Robson, J. M., 136, 141, 363, 415, 463,

466, 551, 603, 657, 697
Robson, M. J., 926, 952, 963, 965, 983,

1004, 1017, 1018, 1045, 1080
Rogers, F. T., 985, 986, 1020, 1127
Rokizky, 833
Rollefson, G. K., 193, 253
Rolleston, H., 1000, 1024
Roman, H., 411, 454, 472
Rosen, G. U., 707
Rosenblum, C, 292, 313
Rosenfeld, F. M., 277, 281, 289, 313,

694, 709, 754, 760, 801, 822
Rosenthal, R. L., 943, 955, 961, 962, 964,

966, 968, 974, 1016, 1024
Roser, F. X., 166, 185
Ross, J. M., 1155-1157, 1184, 1199, 1200
Ross, M. H., 692, 693, 934, 950, 961, 964,

974, 981, 983-988, 1001, 1011, 1013,

1014, 1024, 1039
Rossi, B., 97, 143
Rossi, H. H., 162, 167-169, 171, 176, 186,

188, 320n., 345, 348, 611. 644
Roswit, B., 992, 1013
Rothberg, H., 570, 613, 657, 661, 664,

676, 685, 703
Rother, J., 980, 1026
Rothstein, K., 348
Rovissy, G., 1157, 1200
Rowlands, S., 985, 1013
Rowntree, C. W., 1149, 1200
Rubin, B. A., 168, 188, 586, 587, 620, 622,

750, 760
Rudali, G., 1155, 1177, 1197

Rudolphi, H., 1158, 1200

Rugh, R., 628, 643, 696, 708, 783, 800,
804, 822

Rusch, H. P., 1190, 1200

Russ, S., 773, 822, 1029, 1030, 1032, 1145,
1150, 1151, 1191, 1193

Russell, D. S., 995, 1024

Russell, L. B., 561, 620, 858, 861, 862,
864-867, 869-871, 873-879, 882-886,
888, 891-894, 896, 898-900rc., 901-
908, 910, 917, 932, 933, 955

Russell, W. L., 411, 423, 429, 432, 471,
561, 583, 620, 825, 827, 828, 833-835,
838, 841, 845, 850-853, 855, 856, 858,
865-867, 870, 877, 879, 900n., 903-
906, 908, 910, 917, 932, 955

Rutherford, E., 41-44, 51, 66, 87, 89, 143

Sabin, F. R., 1155, 1156, 1200

Sacher, G. A., 337, 348, 925, 931-934,
951, 955, 971, 1005, 1011, 1044, 1166,
1192

Sackis, J., 925, 955

Sadauskis, J., 66, 143, 165, 187

Salaman, M. H., 329, 347

Salerno, P. R., 926, 955

Salis, H. von, 1127

Salisbury, P. F., 926, 955, 1081

Sallmann, L. von, 943, 957

Saloman, K., 962, 1002, 1008

Salter, W. T., 1000, 1021

Saltmarsh, O. D., 227, 253

Sanderson, M., 966-969, 1008

Sanigar, E. B., 293, 313, 973, 1024

Sansome, E. R., 476, 505, 533, 535, 611, 620

Saretzky, 865, 869, 893, 894, 900, 917

Sarkar, I., 385, 471

Sarvella, P. A., 838

Saunders, T. S., 1175, 1176, 1200

Sax, K., 135, 144, 362, 363, 379, 385, 386,
471, 480, 484, 486, 487, 516, 562, 566,
571, 573, 576, 614, 621, 660, 666, 682,
683, 707, 708, 714-716, 719, 720-730,
735, 737, 740, 741, 749, 759, 760, 799,
822

Sayers, G., 989, 991, 1024

Sayers, M. N., 991, 1024

Schack, J. A., 935, 955, 965, 1024, 1065

Schaefer, H., 828, 829, 858

Schaefer, H. J., 583, 584, 621

Schall, L., 908-911, 917

Schechtmann, J. L., 478, 621

Scheuer, O., 286, 311

Schinz, H. R., 828, 837, 858, 863, 869, 893,
894,917, 1114, 1121, 1179, 1188,1200

Schjeide, O. A., 682, 708, 935, 936. 947

Schmidt I. K, 479

NAME INDEX

1223

Schmidtke, L., 576, 607

Schmitt, O., 578, 621

Schneider, L., 988, 1024

Schneiderman, H., 979, 1024

Schneiderman, H. A., 562, 576, 614

Schoenberg, M. D., 979, 1024

Scholes, G., 283, 290, 313, 694, 708, 753,

754, 760
Scholes, M. E., 775, 779, 819
Schon, M., 763, 822
Schottelndreyer, H., 976, 1024
Schraffenberger, E., 864, 869, 874, 879,

882, 891-892, 901, 907, 918
Schraub, A., 1178, 1180, 1199
Schreiber, H., 531, 540, 614, 811, 821
Schrek, R., 275, 281, 935, 939, 955, 960,

1024
Schrodinger, E., 249, 253, 597, 621
Schugt, P., 348, 1118
Schultz, J., 455, 465, 621, 652, 654, 657,

684, 708
Schulz, M. D., 995, 1017
Schiirch, O., 1152, 1153, 1155, 1156, 1200,

1201
Schutze, R., 574, 621
Schwab, L., 1003, 1004, 1024
Schwartz, D., 560, 621
Schwartz, S., 961, 988, 1019, 1024, 1028,

1039
Schwarz, G., 927, 939, 940, 945, 956,

1162, 1196
Scott, C. M., 144, 262, 281, 297, 300, 313,

1189, 1200
Scott, G. M., 773, 822
Scott, M. B., 89, 142
Scott, M. R., 14, 26, 59, 116, 144
Scott-Moncrieff, R., 461
Searle, A. G., 854, 858
Sebileau, 869, 894, 898, 918
Sedginidse, G. A., 1152, 1200
Seemann, H. E., 180, 189
Segal, G., 960, 991, 1019, 1136, 1137
Segale, G. C, 1112
Segre, E., 66, 87, 141
Seide, J., 784, 812, 822
Seitz, F., 196, 253
Seki, L., 303, 309
Selbie, F. R., 1155, 1157, 1200
Seldin, M., 986, 987, 1024
Seliger, H. H., 92, 99, 144, 160, 189
Selin, G., 1181, 1196
Sell-Beleites, I., 535, 621
Selling, L., 1029, 1039
Selye, H., 989, 1025
Semonov, L. F., 299, 312
Senn, N., 919, 956
Serebrovskava, R. I., 409, 471
Setala, K., 975, 1025

Settles, F., 366, 419, 470, 632, 706
Seymour, A. H., 448, 465, 710
Sgourakis, E., 561, 563, 565-568, 603,

655, 697, 936, 940, 948
Shacter, B., 975, 1025
Shapiro, J. R., 1188, 1196
Shapiro, N. I., 409, 471
Shapiro, N. P., 653, 654, 708
Shaver, A., 986, 1009
Shaw, J. C., 1061

Shechmeister, I. L., 934, 956, 1005, 1025
Shefner, D., 587, 619
Shelton, E., 926, 953, 964, 965, 1020
Sheppard, C. W., 160, 176, 189, 978, 979,

988, 1028, 1167, 1193
Sheremetieva-Brunst, E. A., 924, 948
Shimkin, M. B., 963, 994, 1018, 1020,

1157, 1177, 1192
Shirlock, M. E., 908, 916
Shorvon, L. M., 966, 1025
Shouse, S. S., 966, 1025, 1044
Shuler, K. E., 193, 205n., 206, 223, 252
Sick, 1148, 1200
Sidky, A. R., 363, 380, 469, 480, 481, 501,

519, 618, 621
Sidorow, B. N., 381, 465, 640, 700
Siebenrock, L. von, 1162, 1196
Sieburth, L. R., 558, 622
Siegel, P. W., 1032
Sievert, R. M., 924, 956
SiM, H., 1177, 1178, 1200
Silberbach, I., 988, 1019
Silverman, S. B., 967, 1025
Simmons, E. L., 925, 932, 933, 951, 961),

963-966, 1005, 1017, 1018, 1025,

1026, 1038, 1045, 1058-1060, 1064,

1069-1073, 1076-1080
Simon, N., 985, 1025
Simon, S., 643, 699
Simon-Reuss, J., 776, 778, 779, 782, 806,

822
Sinclair, W. K, 584, 603
Singer, G., 153, 179, 189
Singer, T. P., 262, 278, 298, 299, 309, 551 ,

580, 603, 922, 941, 948
Singleton, W. R., 533, 621
Sipe, C. A., 264, 278
Sipe, C. R., 308, 310, 934, 935, 942, 943,

945, 949
Siri, W. E., 1158, 1200
Skaggs, L. S., 112, 136, 141, 142
Skanse, B. N., 993, 1025
Skirmont, E., 1057, 1065, 1079
Skoog, F., 276, 281
Slack, C. M., 134, 144
Slater, J. C., 32, 144
Slaughter, J. C., 274, 279, 301, 311, 735,

818, 936, 940, 945, 950

1224

RADIATION BIOLOGY

Slizynska, H., 652, 708

Slizynski, B. M., 528, 532, 622, 645, 652,

661, 662, 668, 708, 850, 858
Slotopolsky, B., 828, 837, 858, 1114, 1121
Sluka, E., 1002, 1009, 1080
Smeltzer, J. C, 148, 189
Smith, D. B., 289, 313, 694, 708
Smith, D. E., 264, 281, 308, 313, 930, 935,

937, 938, 942, 944-946, 954, 956, 957,

965, 966, 977, 979, 984, 985, 992, 993,

1001, 1022, 1025, 1026
Smith, F., 935, 937, 956, 991-993, 1001,

1005, 1025
Smith, H. P., 122, 142, 257, 262, 279, 556,

609
Smith, K. A., 277, 278, 290, 310, 754, 757
Smith, K. M., 262, 280, 312, 329, 347
Smith, L., 553, 558, 559, 568, 571, 572,

605, 610, 622, 940, 951
Smith, T. R., 967, 1025
Smith, W. W., 929, 935, 937, 946, 948,

956, 993, 1010, 1065, 1067
Smyth, F. S., 986, 988, 1025
Snell, G. D., 337, 348, 362, 686, 708, 825-

828, 830-838, 841-850, 858, 859, 999,

1026
Snider, R. S., 337, 346, 929, 951, 995,

1026, 1030, 1058, 1093, 1119, 1121,

1131, 1160, 1177, 1195, 1199
Snyder, L. H., 355, 469
Snyder, M. L., 776, 782, 791, 792, 794, 818
Snyder, R. H., 923, 956, 1058
Snyder, W. S., 163, 189
Sobel, E., 993, 1014
Sobel, H., 1169, 1200
Soberman, R. J., 939, 956, 969-972, 974,

1026
Sokoloff, B., 969, 1026
Sommermeyer, K., 346, 657, 704
Sommers, S. C, 805, 823
Sonneborn, T. M., 424, 471, 593, 595, 622
Sonnenblick, B. P., 632, 647, 667, 687,

689, 709, 910, 918
Spargo, B., 1100, 1104, 1105, 1116, 1118
Sparrow, A. H., 277, 281, 289, 313, 555,

622, 630, 688-691, 694, 695, 706, 709,

722, 723, 748, 750, 754, 760, 801, 802,

822, 928, 956
Spear, F. G., 335, 336, 338, 345, 348, 628,

630, 709, 763, 773-779, 781, 782, 786,

787, 790, 794, 797, 804-806, 809, 810,
817, 819, 822, 823, 824, 928, 956, 957
Speder, E., 1127
Spence, I., 907, 914
Spencer, L. V., 71, 109, 144
Spencer, W. P., 476, 622
Sperduto, A., 25, 141
Spiegel-Adolf, M., 292, 313

Spiegelman, M., 1162, 1193

Spiers, F. \V., 164, 185, 189, 934, 956

Spies, T. D., 934, 952, 985, 1005, 1009, 1018

Spinks, J. W. T., 584, 603

Spitz, S., 1153, 1200

Sponer, H., 222, 225, 227, 252, 253

Sprague, G. F., 531, 533, 540, 622

Spurring, R., 1029

Spurr, C. L., 967, 1025

Ssipowsky, P. W., 1136

Stadler, L. J., 362, 363, 366, 389, 411, 454,
472, 476, 480, 494, 531, 533, 534, 540,
552, 555, 565, 574, 622

Stanton, Elizabeth, 346

Stapleton, G. E., 131, 143, 301, 311, 333,
334, 336, 337, 346, 349, 350, 506, 521,
556-558, 564, 579, 580, 605, 611,
622, 922-924, 935, 940, 941, 944, 951,
952, 956, 1005, 1026, 1030, 1049,
1146, 1160, 1167, 1177, 1195

Steacie, E. W. R., 193, 208, 253

Steamer, S. P., 925, 932, 956, 960, 963,
964, 966, 989, 1026, 1031, 1064

Steele, R., 723, 760

Steggerda, F. R., 975, 980, 981, 1000, 1028

Stein, G., 147, 152, 186, 257, 267, 268,
270, 281, 290-292, 297, 311 313, 694,
708

Steinglass, P., 587, 620

Stenbeck, T., 919, 956

Stenstrom, W., 296, 313

Stern, C., 131, 141, 361, 368, 403, 404,
427, 472, 476, 478, 605, 622, 625, 635,
638, 653, 709 1

Stevens, L. G., 919, 956

Stewart, F. W., 1181, 1182, 1193

Stinson, F., 71

Stock, H. C., 1004, 1013

Stone, L. H. A., 774, 823

Stone, R. S., 338, 349, 1005, 1018

Stone, W. S., 303, 313, 314, 417, 455, 470,
473, 511, 514, 537, 544, 545, 548-552,
557, 561, 562, 566, 567, 598, 606, 610,
619, 623, 625, 626, 641, 650, 707, 751,
761

Stoneburner, C. F., 153, 179, 189

Storaasli, J. P., 966, 968, 1017

Storer, J. B., 914, 915, 957

Storey, R. H., 966-969, 971, 978, 979,
988, 1009, 1026, 1028, 1039

Storey, W. F., 1162, 1197

Stowell, R. E., 692, 709

Strandskov, H. H., 826, 828, 831-833,

836-839, 859
Strang, V., 966, 1011
Strangeways, T. S. P., 554, 623, 773-776,
778, 799, 805, 806, 810, 823, 1097

NAME INDEX

1225

Straube, R. L., 264, 281, 308, 313, 937,
938, 942-946, 954, 956, 957, 965, 966,
990-992, 994, 1005, 1022, 1026, 1067

Strauss, O., 980, 1026, 1132

Strelin, G. S., 926, 957

Str^mnaes, 0istein, 688, 709

Stroud, A., 1074

Stubbe, H., 414, 415, 472, 476, 478, 510,
531, 540, 541, 553, 576, 586, 589, 607,
613, 618, 623

Sturm, E., 1003, 1021

Sturtevant, A. H., 368, 373, 382, 438,
472, 635, 638, 643, 709

Suche, M. L., 641, 653, 707

Sugiura, K, 339, 349, 923, 957, 981, 1018,
1096

Supplee, H., 972, 1016

Suter, G., 960, 963, 1026, 1031, 1035,
1037, 1048, 1049

Sutton, E., 518, 521, 538, 606, 645, 647,
648, 661, 666, 669, 700, 709

Sutton, H., 1035, 1048, 1055

Svedberg, T., 296, 313

Svenson, H. K, 359, 467

Svihla, G., 930, 956, 977, 979, 1025

Swann, M. B. R., 985, 1026

Swanson, C. P., 366, 472, 485, 503, 504,
533, 534, 537, 546, 547, 569, 570-574,
578, 579, 597, 600, 611, 615, 623, 660,
661, 682, 683, 708, 709, 714, 721-723,
738-741, 760, 799, 814, 822, 823

Swenson, P. A., 304, 310

Swift, H. H., 695, 709

Swift, M. N., 926, 932, 935, 936, 940, 945,
946, 948, 954, 955, 961, 964, 965, 967,
969-975, 979-982, 984, 985, 989, 993,
995, 1000, 1001, 1005, 1010, 1022,
1023, 1025, 1031, 1037

Swisher, S. N., 926, 957, 1082

Sydow, G., 574, 605

Szilard, B., 148, 189

Szilard, L., 418-420, 470, 537, 543, 545,
619

Tahmisian, T., 301, 313
Tait, J. H., 163, 189
Takamine, N., 811, 815, 823
Talbot, J. M., 964, 1026, 1075
Taliaferro, L. G., 934, 957, 1003, 1005,

1026
Taliaferro, W. H., 934, 957, 1003, 1005,

1026
Tansley, K., 338, 348, 628, 709, 774, 776,

781, 782, 787, 805, 806, 808-810, 819,

823, 928, 956, 957, 995, 1024
Tarlov, I. M., 995, 1012

Tarr, R., 1004, 1013

Tatum, E. L., 424, 461, 465, 593, 594,
596, 624

Taurog, A., 993, 1014

Tavernier, G., 163, 164, 185

Taylor, B., 277, 281, 289, 313, 314, 694,
709, 754, 761, 801, 823

Taylor, C. S., 148, 187

Taylor, C. V., 303, 314

Taylor, Grant, 1201

Taylor, H. D., 1032

Taylor, H. S., 236, 252

Taylor, L. S., 109, 121, 142, 148, 152, 153,
157, 179, 182, 186, 188, 189

Taylor, W. R., 632, 709

Tazima, Y., 457

Teleky, L., 1201

Telfer, J. D., 563

Teller, E., 32, 143, 222, 225, 227, 252, 253

Templeton, F., 1125

Tennant, R., 336, 347, 983, 1002, 1019, 1100

Tessmer, C, 966, 1012

Thaddea, S., 990, 1026

Thew, K., 14, 26, 59, 116, 144

Thilo, E. R., 134, 144

Thoday, J. M., 275, 281, 306, 314, 340,
341, 349, 484, 487, 491, 517, 557, 558,
566, 567, 599, 605, 624, 632, 656, 660,
668, 675, 698, 710, 716, 717, 720,
724-728, 731, 736, 741, 744, 748, 751,
756, 761, 940, 957

Thomas, H., 581, 624

Thomas, J. O., 303, 314

Thomas, M. M., 1032

Thompson, E. C, 994, 997, 1025

Thompson, E. O., 1065

Thompson, M. V., 657, 796, 822, 925, 954

Thompson, W. R., 299, 312

Thomson, J. F., 988

Thoraeus, R., 180, 182, 189

Timofeeff-Ressovsky, N. W., 127, 130,
139, 144, 319, 333, 349, 394, 396, 399,
410, 412, 414, 464, 472, 476, 478, 479,
481, 499, 508, 509, 512-514, 516, 521,
523, 526, 565, 572, 582, 588, 597, 606,
619, 620, 624 626, 630, 658, 670, 684,
710

Ting, T. P., 961, 1026

Tinsley, M., 988, 1024

Tipton, S. R., 748, 757

Titterton, E. W., 164, 189

Tittle, C. W., 164, 189

Tobias, C. A., 139, 302, 316, 317, 332,
344, 349, 350, 1189, 1201

Tompkins, M., 983, 1002, 1003, 1021

Tompkins, P. C, 1058

Toolan, H. W., 964, 1026

Torda, C, 985, 996, 1026

1226

RADIATION BIOLOGY

Torgersen, O., 990, 991, 1014, 1027, 1131

Toulis, W. J., 328, 349

Toussey, S., 863, 918

Toyoma, T., 980, 985, 1027

Troadwell, A. de G., 935, 957, 1066, 1079

Tribondeau, L., 553, 604, 627, 681, 697,

927, 948, 1114, 1133
Trillmich, F., 865, 866, 869, 893, 894, 918
Troland, L. T., 392, 473
Tnijillo, T. T., 935-937, 946, 951, 993,

1016
Trump, J. G., 93, 144
Tsuzuki, M., 983, 1027
Tullis, J. L., 930, 957, 972, 974, 1010, 1099
Turkowitz, H., 940, 947
Turner, C. W., 1123
Tweedie, M. C. K., 259, 272, 279
Twombley, G., 157, 186
Tvler, W. J., 848, 859
Tyree, E. B., 264, 281, 308, 313, 935, 939,

942-945, 954, 956, 957, 965, 984, 985,

990, 992, 993, 1001, 1022, 1025, 1026,

1067
Tytell, A. A., 299, 314

U

Uber, F. M., 531, 534, 540, 622
Uehlinger, E., 1155, 1156, 1179, 1188,

1200, 1201
Uggeri, B., 685, 699
Ullmann, H. J., 999, 1009
Ullmann, T., 987, 1027
Ullrich, F. W., 966, 1012
Ulrich, H., 1162, 1201
Uncapher, R. P., 932, 949, 969, 1011
U.S. Atomic Energy Commission, 442,

473
Unterberger, F., 390, 473
Upcott, N. B., 721, 757
Uphoff, D., 403, 427, 472, 476, 478, 625,

926, 953, 964, 965, 1020, 1035, 1048,

1075, 1165, 1173, 1188, 1198
Upton, A. C, 1146, 1163, 1164, 1201
Uri, N., 256, 279

Valencia, J. I., 385, 409, 411, 470, 479,
481, 488, 493, 495, 496, 498, 500, 502,
503, 514, 519, 521-523, 533, 552, 618,
622

Valencia, R. M., 385, 409, 411, 470, 479,
488, 503, 514, 552, 618

Valentine, W. N., 926, 953, 960, 962, 963,
970, 978, 1004, 1012, 1019, 1027,
1032, 1035, 1037, 1039, 1044, 1063,
1065

Van de Graaff, R. J., 11, 25, 93, 141, 144

Van Dyke, D. C, 926, 957, 1136

Van Valkenburg, P. A., 992, 1017

Velley, G., 942, 946, 953

Vennesland, B., 308, 310

Venters, K. D., 975, 1027

Villard, P., 148, 189

Villee, C. A., 503, 625

Vilter, C. F., 934, 952, 1005, 1018

Vilter, R. W., 985, 1009

Vintemberger, P., 935, 947

Vinzent, R., 1152, 1155, 1183, 1186, 1197

Vogt, E., 1171, 1201

Vogt, M., 416, 473, 480

Volkin, E., 973, 1027

Von Halle, E. S., 561, 603

W

Waddington, C. H., 907, 918, 1189, 1201

Wagner, A., 1164, 1178, 1196

Wagner, R. P., 417, 473, 548, 549-551,
625, 751, 761

Wakely, G. P. G., 1151, 1154, 1201

Waletzky, E., 848, 859

Walkhoff, 1150

Wallace, B., 422, 437, 464, 473

Wallace, J., 988, 1024

Wallace, R. H., 589, 625, 737, 761

Walsh, D., 919, 957, 982, 1027

Walters, O. M., 1132

Warburg, O., 304, 314

Ward, F. D., 389, 473

Warkany, J., 864, 869, 873-874, 879, 882-
886, 891-892, 901, 907, 918

Warren, S., 686, 710, 763, 773, 776, 805,
823, 929, 930, 950, 957, 966, 976, 982,
983, 988, 990-993, 996, 999, 1005,
1018, 1020, 1027, 1094-1097, 1117,
1123, 1130, 1131, 1101, 1168, 1201

Warren, S. L., 966, 982, 983, 1000, 1002,
1005, 1017, 1025, 1027, 1028, 1029,
1044, 1057, 1058, 1092, 1094, 1124,
1126-1128, 1131

Washburn, M., 307, 311

Watson, C. J., 961, 1024

Watson, M. D., 569, 572, 625

Wattenberg, L., 988, 1028

Wattenwyl, H. von, 686, 710

Way, K., 14, 26, 59, 116, 144

Weatherwax, J. L., 109, 142, 182, 186,
189

Weber, R. P., 975, 980, 981, 1000, 1028

W.-bster, J. H. D., 986, 1013

Weichert, U., 990, 992, 1028

Weinstein, A., 389, 473

Weiss, J., 257, 267, 268, 270, 281, 290,
291, 296, 297, 312, 313, 694, 708, 750,
753, 754, 761, 937, 957

Weiss, L., 1182, 1192

\\ elander, A. D., 448, 465, 628, 710

NAME INDEX

1227

Wells, P. H., 811, 823

Wels, P., 305, 314, 1001, 1028

Welt, P., 481, 519, 606, 669, 670, 671, 698

Wertheimer, J., 936, 955

Wertz, E., 555, 625, 939, 957

Westergaard, M., 416, 466, 547, 550, 577,

612, 751, 758
Westman, A., 994, 1009
Wettstein, W. von, 584, 608
Wheeler, B., 1060
Whipple, G. H., 966, 982, 983, 985, 988,

989, 1000, 1002, 1016, 1021, 1025,

1027, 1028, 1029, 1044, 1124, 1127,
1128

Whitaker, D. M., 543, 625

White, A., 990, 991, 1013, 1024

White, C. J., 1149, 1199

White, G. R., 24, 38, 40, 41, 70, 104, 105,

110, 111, 144
White, H. E., 30, 144
White, M. J. D., 628, 643, 646, 665, 710,

799, 823
White, T. N., 153, 154, 189, 343n., 349
Whiting, A. R., 384, 388, 456, 473, 510,

625, 643, 657, 667, 683, 688, 689, 710,

799, 808, 823
Whiting, P. W., 453, 473, 608, 711
Whittinghill, M., 653, 654, 702, 711
Wick, G. C, 166, 189
Widner, W. R., 681, 779, 780, 820
Wilbrandt, W., 961, 1020
Wilbur, K. M., 796, 824
Wilder, H. C., 998, 1028
Wilhelmy, E., 478, 508, 625
Willey, E. J. B., 193, 202n., 204, 205, 253
Williams, A. E., 1174, 1199
Williams, F. C., 87, 141
Willis, G. S., 1151, 1198
Willis, R. A., 1157, 1201
Wilsey, R. B., 147, 190
Wilson, C. T. R., 144
Wilson, C. W., 174, 190, 776, 782, 797,

810, 819, 824, 932, 957
Wilson, G. B., 485, 618
Wilson, H., 1132
Wilson, J. G., 864, 869, 872-874n., 876,

879, 882-883, 885-888, 891-892, 897,

901-904, 907, 918, 932, 933, 957, 996,

1028, 1185, 1201

Wilson, K., 675, 676, 703, 740, 759

Wilson, R. R., 4, 5, 81, 100, 144

Wimpfheimer, S., 910, 916

Winchester, A. M., 689, 707

Winter, K. A., 971, 1009

Wish, L., 969, 970, 978, 979, 988, 1026,

1028, 1039
Witkin, E. M., 125, 144, 422, 464, 553,

625

Wolbach, S. B., 1122, 1123, 1143, 1150,

1175, 1201
WToldrich, A., 1179, 1201
Wolf, B. S., 182, 187
WTolff, H. G., 985, 996, 1026
Wolfson, N. D., 769, 779
Wolkowitz, W., 984, 986, 1001, 1009
Wollman, E., 330, 349
Wood, F. C., 349
Woodard, H. Q., 1181, 1182, 1193
Woods, M. C., 966, 968, 971, 978, 979,

1009
WPA, Federal Works Agency, 171, 190
Wrenshall, G. A., 182, 190
Wright, G. P., 775, 824
Wright, K. A., 93, 144
Wright, S., 854, 859
Wuensche, H. W., 1034
Wyckoff, H. O., 108
Wyckoff, R. W. G., 330, 349
Wyman, R., 703
Wyss, O., 303, 313, 314, 417, 473, 511,

537, 544, 545, 548-552, 557, 562, 598,

606, 610, 623, 625, 751, 761

Yamashita, H., 771, 772, 785, 787, 789,

821, 822, 824
Yamawaki, T., 1163, 1194
Yost, H. T., Jr., 570-572, 624, 626, 739,

741, 760, 761
Young, L. E., 961, 1012
Young, R. A., 785, 819

Z

Zamenhof, S., 419, 420, 473

Zerahn, K, 306, 308, 926, 947

Zerwic, M. M., 988, 1013

Ziegler, K., 554, 614, 627, 688, 704, 982,
1019

Zimmer, E. M., 533, 535, 611

Zimmer, K. G., 127, 130, 139, 144, 319,
320, 333, 349, 478, 487, 508, 521, 523,
526, 565, 572, 574, 588, 597, 605,
607, 619, 624-626, 630, 710

Zinder, N., 420, 454, 467

Zirkle, R. E., 138, 139, 144, 278, 281, 302,
316, 319, 327, 330, 332-334, 336, 337,
343n., 344, 346, 348-350, 655, 711,
786, 796, 824, 921-925, 928, 933, 938,
939, 948, 951, 957, 958, 961, 963, 965,
972, 983, 1008, 1011, 1017, 1026,
1030, 1042, 1044, 1058, 1094, 1131,
1146, 1189, 1201

Zuitin, A. I., 414, 473

Zunz, E., 991, 1028

SUBJECT INDEX

Abdomen, shielding of, 1072
Aberrations, chromatid, 715

chromosome (see Chromosome aberra-
tions)
diagrams of types, 716, 718
distribution in nuclei, 720
effects of modifying factors on, 736jf.
efficiencies of radiations in producing,

732
induced by absorbed radioisotopes, 733
induced by slow neutrons, 733
one-hit, 723
radiation-induced, in chromosomes of

Tradescantia, 715-720
relation of yield, to dose, 723

to intensity, 727
spontaneous vs. induced, 720
two-hit, 723
Abnormal bleeding, 966-969
Abnormal mitosis, 628-630, 632, 646, 647,

664, 1092
Abnormalities, chromosome, induced,
627, 628
in tumors, 338

(See also Chromosome aberrations;
Morphological changes follow-
ing irradiation; Mutations)
following prenatal irradiation, 879-912
critical periods for induction of,
880-887, 891-893, 901, 902, 904,
906, 909, 912, 913
Abortion following irradiation of embryo,

866, 910-911
Absorption, of electromagnetic radiation,
17, 20, 45-48
energy (see Energy absorption)
fat, 986

glucose, 307, 986
from intestine, 986
linear energy, 316
X rays, 49

broad-beam, 108
narrow-beam, 107, 108
Absorption coefficient, /3-ray, 102
light, 118
linear, 104

Absorption coefficient, mass, 104

narrow-beam, 106

neutrons, 114

X rays, 103

true (adjusted), 110
Absorption spectrum, 47

albumin, 294
Accelerated atomic nuclei, 316-318
Accelerator, electrostatic, 11

induction (betatron), 12

linear, 12
Accidents, radiation, 998, 1005, 1006
Acenaphthene, 577
Acetic acid, 576

Acetylcholine synthesis by brain, 996
Acetylene polymerization, 291
Acid-base balance, 974
Acquired radioresistance, 1136

of intestine, 1127
Acridine orange, 542
Acrinavin, 596
Activated (intermediate) complex, 220,

222, 223»., 224
"Activated water," 284, 937
Activation energy, 35, 60, 210, 215-216

distribution of, 60
Activations, 60, 129

distribution of, 135-136

ionizing, 129

mechanism of, 62

and mutation, 490

by secondary electrons, distribution of,
80

spatial distribution of, 76
Acute radiation (see Radiation ; X radia-
tion)
Acute-radiation reaction, 980
Acute-radiation syndrome, 1005-1008
Adaptation syndrome, 989-990
Additivity, different radiations, 924

radiobiological, 326
Adenosine triphosphate, 289, 987
Adenosinetriphosphatase (myosin), 298,

922
Adenylic acid, 289
Adipose tissue, 1097

Adrenal cortex, atrophy of, following pre-
natal irradiation, 889

1229

1230

RADIATION BIOLOGY

Adrenal cortex, extracts of, 935, 992

lipids of, 990
Adrenal insufficiency, 934, 992, 1005
Adrenal medulla, 991
Adrenal radiation exposure, 982
Adrenalectomy, effect on leukemogencsis,

1165
Adrenals, 306, 929, 989-992

catechols, 991

hyperemia, 1131

immunity, 1004

latent period after irradiation, 1132

radioactive isotopes and, 1132

radium and, 1132

X rays and, 1131
Adrenocorticotropic hormone, 990, 992
Aerobic glycolysis, 306
Age, and radiation effects, 1093

and radiosensitivity, 933
Alanine, 305
Albumin, 293

absorption spectrum, 294

egg, 293

plasma, 295

serum, 294-296
Albumin-globulin ratio, plasma, 973
Alcohol, 291

ethyl, 580
Alcohol dehydrogenase, 299
Aldehydes, 417

aliphatic, 290
Algae, 322, 445
Alkalosis, 1000
Alkaptonuria, 461
All-or-none phenomenon, 121
Allantoin, 988
Alleles, 355

iso-, 403

positional, 404

pseudo-, 594
Allium, 589, 683, 737, 776, 782, 799, 801,

815
Alopecia, 919, 1148

Alpha irradiation, internal, 964, 970, 973,
1000

oxygen effect, 940
a particles, 318, 320

atomic weight, 7

charge, 7

effects of, on chromosomes, 724, 726
as modified by oxygen, 748, 751, 752

ejection of, 35
spontaneous, 36

relative efficiency in chromosome-
aberration production, 732

scattering, 42
Alpha radiation, effects of, on carboxy-
peptidase, 271

Alpha radiation, effects of, compared
with X rays, 271
cytosome, 785
on glycine deamination, 271
mechanism, 271
mitotic, 785-787
on tyrosine, 271
a rays, 3

(See also Alpha radiation)
Altitude, 935, 965
Ambystoma, 783
Ameba, Chaos, 304
Amenorrhea, 999

Amino acid oxidase, X-ray effect on, 261
D-amino acid oxidase, 299
Amino acids, 307, 324, 328
deamination, 296
oxidation, 307

(See also individual amino acids)
D-amino acids, 306
Amino groups, 292
Ammonia, 414, 575
Ammonia formation, 306, 390
Ammonium hydroxide, 575
Amorphous cementing substance and

bone, 1108
Amphibia, 554, 628, 643, 656, 801, 803,

804, 806
Anaerobic glycolysis, 306
Anaerobiosis, 940-941
Anamnestic reaction, 1004
Anaphase, 628, 632, 645, 647, 661, 662,

669, 689
Androgenesis, 510
Anemia, 922, 961

and bone marrow, 1100
induced by acute radiation, 1036-1041,
1047, 1079
effect on, of irradiation, 1064-1066
mechanism of recovery from,
1038-1040
of spleen-shielding and increments
to spleen, 1079
induced by chronic radiation, 1050-
1052, 1054-1056
by gamma rays, 1054-1056
macrocytic, 1056
by radioisotopes, 1058-1060
recovery from, 1056
induced by fast neutrons, 1043
induced by gamma radiation (see

Gamma radiation)
induced by X radiation (see X radia-
tion)
pathogenesis of, 964
phcnylhydrazine-induced, and X radia-
tion, 1064-1066
recovery from, 1067, 1068

SUBJECT INDEX

1231

Anemia, radium-induced, 1059

(See also Cobalt-induced polycyth-
enia; Erythrocytes)
Anesthesia, nembutal, 937, 946
Aneuploidy, 842, 849, 854

and radiation damage to embryo, 903,

913
of whole chromosomes, 358-359
Angiosperms, 554
Anlagen, optic, 458
Annihilation, 41, 69, 98
Anomalous viscosity, 289
Anorexia, 982, 984-985, 1001

neurogenic origin of, 985
Anoxia and radiosensitivity, 940, 1065-

1066
Antibiotic therapy, 1002, 1003
Antibiotics, 445
Antibody formation, 1002, 1005

effect on, of appendix shielding, 1080,
1081
of spleen shielding, 1080, 1081
.and splenectomy, 1080, 1081
Anticoagulants, circulating, 966
Antigens, 593, 595
Antioxidants, 308
Antirrhinum, 414, 415, 417, 476, 478, 531,

540, 541, 553, 556, 586, 589, 591
Anurans, 554

Appearance potential, 239, 240
Appendicular skeleton (limb, foot, girdle),
abnormalities following prenatal ir-
radiation, 879-882, 885, 887-888,
892, 902, 904-905, 907, 909
Appendix, 1101

effect of shielding, on antibody for-
mation, 1080
on lymphopoiesis, 1063
on survival, 1070, 1071
Arbacia, 300, 301, 304, 766, 769, 771, 772,

784, 785, 789, 796, 803, 812, 814
Arginase, 300, 987
Argon, 559
Arsenic, 574
Arsenoxide, 946
Arteriolar change in skin, 1122
Ascaris, 554, 558, 784, 812
A scar is eggs, 628, 674, 935
Ascomycete, 593
Ascorbic acid, 288, 942, 945, 967
of adrenals, 990
of serum, 975
Ash content of intestinal cells, 983
Aspergillus, 444, 503, 506, 521, 535, 537,
546, 547, 556, 564, 570, 578, 579
spores. 326
Asphyxia, tissue, 1001

Asters, supernumerary, X-ray-induced,

803
Astrocytes, radiosensitivity of, 995
Atmospheric oxygen, 307
Atomic-bomb effects, human, 966, 998,
999, 1005

leukemogenesis by, 1163

{See also Hiroshima)
Atomic nuclei, accelerated, 316-318

excitation of, 57

structure of, 27
Atomic number, 27

effective, 105n.
Atomic weight, 6, 27
Atoms, aggregates of, 31

excitation of, 197-208

hydrogen, 285

structure of, 27
Atrophy of tissues, causes, 927
Atropine, 980-981, 996
Atypical bone, 1101, 1111, 1112
Auger effect, 34, 47
Aureomycin, 1003
Autoradiographs of bone, 1111
Autotrophism, 462
Avian tissue, 629
Azide, 551, 562

B

Backscattering, electron, 91

neutron, 116
Bacteremia, 923, 983, 1000, 1006
Bacteria, 321, 324, 330-331, 415, 417,

444, 462, 553
endotoxins, 1003
Bacterial invasion, 965
Bacterial toxins, 1000, 1003
Bacteriophage, 543, 544, 569, 572, 587,

591, 592
Bacterium prodigiosum, 538, 541, 542
BAL, 579

protection against, 942, 945
Barium, 574
Ba140-La140, effect of, 1058

on intestine, 1127

on spleen, 1107

on testis, 1117
Barley, 442, 485, 552, 555, 556, 559, 565,

568, 569, 574-576, 578, 590
Barley seeds, 940
Basophilic outstretched cells in testis,

1114, 1116
Bats, radiation effect on, 930

wing circulation, 977
Beans (see Vicia faba)
Beets as dietary factor, 969
Bellevalia, 484, 801
Benadryl, 981

1232

RADIATION BIOLOGY

Benzene, 290, 291

hydroxylation of, 257
reaction with neutrons, 290
recoil protons, 290
Benzol, 922, 1094
Benzpyrene, 577
0 disintegration, 8, 35
Beta irradiation, external body, 931, 970,
984, 997
internal, 924-926, 963-964, 970, 973,
978, 980, 993, 1003
(See also specific isotopes)
/3 particles, negative (see Electrons)

positive (see Positrons)
/3- to X-ray ratio, 923
0 rays, 3, 317, 319

effect of, 1030, 1058, 1059
lethal, on cells, 809
mitotic, 771, 776, 787, 788
emission of, 3

energy release in tissues, 764, 785, 786
secondary, 117
Betatron, 12, 1030
Bile pigments, excretion of, 961
Bile salts, 988
Biliary passages, 1128
Biliary tract, carcinogenesis of, 1171
Binding energy, 28, 60
Biological effectiveness, relative, 321,

322, 328-342
Biological variability, 27
Birefringency, 289
Bleeding, abnormal, 966-969
Bleeding time, effect of irradiation on,

1044-1046
Blood, dyscrasias after irradiation, 963
electrolytes, 974

flow of, effect on radiosensitivity, 927
hepatic, 977-978
and radiation injury, 940
phosphorus content of, 1000
platelets, 966, 978

radiation effects, 922, 960-976, 1029-
1082
on capillary fragility, 1046
on clot retraction, 1046
on clotting time, 1044, 1046, 1047
red cells (see Erythrocytes)
urea of, 973, 989
uric acid of, 925, 973, 1000
white cells (see Leukocytes; Lympho-
cytes)
Blood calcium, 967
Blood cell mass, 960

Blood cells (see Erythrocytes; Leuko-
cytes; Lymphocytes)
Blood chloride, 974
Blood clotting mechanisms, 966-968

Blood coagulation, 1044-1047
effect on, of radiation, 1044
of toluidine blue, 1044
Blood counts, 325, 337

following irradiation during fetal pe-
riod, 895-896
Blood-forming tissues, recovery of, 926

regeneration of, 963-964
Blood plasma, constituents, 972-982
globulins, 973
histamines, 975, 980
lipids, 974-975
potassium, 974
proteins, 972-973
iodinated, 970
sodium, 974
sulfhydril, 969
electrophoretic pattern, 973
refractive index, 972
turbidity, 974-975
volume, 961, 969
Blood-pressure fall, 970, 980
Blood transfusions, 966, 967, 1006

effect on irradiated dogs, 1081, 1082
Blood vessels, fragility of, 968
obliterative changes in, 1176
Body fluids, 969-976
Body size following prenatal irradiation,
869, 876-877, 879, 896, 900, 905, 912
Body weight and radiosensitivity, 933
Bone, 1108#.

amorphous cementing substance in,

1108
autoradiographs of, 1111
calcium phosphate, 1108
carbonate, 1108
citrate, 1108

collagenous fibers of, 1108
cortex, 1111
devitalized, 1111
epiphysis, 1108
growth, endosteal, 1108
periosteal, 1108
stunted, 1110
hydroxyapatite, 1108
metaphysis, 1108, 1109, 1111
osteoblasts, 1108, 1110, 1111
osteoclasts, 1108, 1110
osteocytes, 1108
osteogenic sarcomas and radioisotopes,

1111
radiation effects on, 999-1000
carbon14, 1110
Plutonium, 1110, 1112
radium, 1110-1112
strontium89, 1110
Xrays, 1110
yttrium91, 1110, 1112

SUBJECT INDEX

1233

Bone, radiosensitivity of, 929
Bone marrow, 339, 1098, 1099
bone-seeking isotopes and, 1100
effect of irradiation, 926, 960-965
by fast neutrons, 1101
gamma radiation, 1056
nonspecificity of, 1101
by slow neutrons, 1101
by strontium89, 1059
by X radiation, 1066, 1099, 1101
and phenylhydrazine-induced ane-
mia, 1066
and spleen shielding, 1071
erythrocytes of, 1099
granular leukocytes of, 1099
Hiroshima studies of, 1101
injections of, modification of radiation
injury by, 926, 964-965, 107-1,
1075
heterologous, 964, 1075
homologous, 1074, 1075
macrophages, radioresistance of, 1100
mast cells and, 1100
megakaryocytes, 1099
metabolism, 962

of homogenates, 1002
myelocytes, 964, 1099
rabbit, glycolysis in, 305
regeneration of, 1101
uptake of iron by, 962
Bone-marrow changes and toxins, 1101
Bone matrix, 1108
Bone repair, 1111
Bone-seeking isotopes and bone marrow,

1100
Bone tumors, 1147, 1152, 1155, 1157,
1160, 1170, 1180-1184
induced by plutonium239, 1060
Bragg curve, 87
Brain carcinogenesis, 1185
"Breakage-first" theory, 480
Breathing retardation, 940
Bremsstrahlung, 23, 69
Broad bean (see Vicia faba)
Bromus, 798

Bronchial epithelium of lung, 1130
Bronchogenic carcinoma, 1153, 1157,

1160, 1177-1180
Brownian movement, 383
Brunner's glands, 1126
Budding of nuclei, 1096
Build-up, neutrons, 116

X rays, 108
Build-up factor, 108, 116

calculations, 109
Butterflies, 390

Cabbage as dietary factor, 968

Cadmium, 946

Caffeine, 576

Cage effect, 231, 246

Calcium, 295

in blood, 967, 974
Calcium phosphate, 1108
Callitroga americana, 441
Cancer (see Carcinogenesis; Carcinogenic
agents; Leukemia; Leukemogenesis;
Radiation; Radioactive elements;
Tumors; specific animals and sites)
Cancer Commission of Harvard Uni-
versity, 1150
Cancer theory, mutation, 1170, 1189

virus, 1170
Cancerogenic agent (see Carcinogenic

agents)
Cancerolytic agent, 283
Capillaries, dilatation of, 976
Capillary fragility, effect of irradiation

on, 1044
Carbamates, 417
Carbohydrates, 298

in cells, 1095
Carbon14, 305

and bone, 1110
Carbon dioxide, 575

combining power of, 974
production of, 962
Carbon monoxide, 562
Carbon tetrachloride, 291
Carbonate and bone, 1108
Carboxylic acids, salts of, 580
Carboxypeptidase, 297, 299, 307
radiation effect on, 258, 271
alpha-, 271
Carcinogenesis, endocrine imbalance and,
1167, 1170
ionizing radiation compared with other

carcinogens, 1185-1188
mechanism of, 1188-1191
physical aspects of, 1146-1148
radiation-induced, and treatment, 919
by radioactive elements (see Radio-
active elements)
relative effectiveness of different radia-
tions, 1146, 1147
threshold dose in, 1167
(See also specific animals and organs)
Carcinogenic agents, 283
aging, 1191

bone seekers (see Radioactive elements)
cosmic rays, 1161
endogenous, 1190

by radioactive substances, 1153-1160

1234

RADIATION BIOLOGY

Carcinogenic agents, estrogen, 1173
fission products, 1159
ionizing radiations, 1145-1201
neutrons, 1100, 1161, 1166, 1167

cataract, 1167
ultraviolet radiation, 1150, 1190
(See also Radioactive elements)
Carcinogenicity and mutagenicity, 421-

424
Carcinoma in rats, 684
Cardiovascular system, 976-982
Cartilage, degenerative changes of, 1112
ear, 1000

hypertrophic, 1112, 1113
provisional calcification, 1112
radiation effects, 999-1000
radioactive isotopes, 1113
radium, 1113
strontium89, 1113
X rays, 1113
trachea, 1000
Cascade showers, 63, 70, 95, 111
Castration cells, 999
Cat, 863
Catalase, 286, 288, 299, 300, 303, 544,

549, 551, 562, 581, 941, 987
Cataracts, 325, 337, 1131

following prenatal irradiation, 879, 896,

909, 912
radiation, 943, 998

induced by neutrons, 1167
Catechols of adrenal, 991
Cathepsin II, 300
Cathode rays, 3
Cell damage and radiation effects on

embryos, 903-904, 906, 913
Cell death, 1092, 1133

X rays, 1134
Cell division, 289, 682

interference with, 357-359
Cell membrane, 1094, 1095
Cell physiology, 462

Cell viability (see Viability effects, cell)
Cells, metabolism of, 922
swelling of, 938, 972
turnover of, in radiosensitivity, 929
Cellular functions, effects on actual and

potential, 1092
Cellular inclusions, 1095
Central nervous system (brain, spinal
cord), damage following prenatal
irradiation, 880, 881, 883, 888-891,
896-898, 902, 909, 912
Centrifugation, effect on radiation-in-
duced chromosome aberrations, 737
and mutation, 573, 589
Centriole, 525
Centromere, 354, 850

Centrosome, 1095
Cerebellum lesions, 995
Cerium-praseodymium and thymus, 1105
Cessation, of mitosis, 1096
of spermatogenesis, 1116
Chaetomium, 531, 538, 541, 593, 595
Chaetopterus, 804
Chain reaction, 264, 270
Characters, 395
Charge exchange, 237-238, 245

(See also Electron-transfer processes)
Charged particles, acceleration of, 1 1
collision of, 41

elastic, 42
heavy (positive ions), 7
sources of, 10
nuclear, 13
Cheese, 445
Chemical bond, 31

energy of, 28, 31, 60
Chemical changes, 60
Chemical forces, 34, 60
Chemiluminescence, 201
Chemostat, 418, 419
Chiasma, 357
Chick embryo, 335

Chickens, 774, 776-778, 781, 782, 787, 788,
790, 794, 796, 797, 805, 806, 809-811
carcinogenesis in, 1155, 1158
eggs, 935

radiation effects, 925
time of death, 932
Chloride, excretion of, 989
of plasma and blood cells, 974
in tissue, 972
Chlorine, 552
Chloropicrin, 551
Chloroplastids, 595
Cholera vibrio, 1003
Cholesterol, 291
of adrenals, 990
of liver, 987
of plasma, 975
Cholic acid, 291

acetylation, 291
Choline acetylase inhibitors, 996
Chortophaga (see Grasshopper)
Chromatid breaks, 717

relation of, to radiation dose, 724
to radiation intensity, 727
Chromatid exchanges, 717

relation of, to fast-neutron dose, 726
to radiation intensity, 727
to X-ray dose, 725
symmetrical and asymmetrical, 720
Chromatids, in chromosomes, 628, 632,
642, 647-650, 659-661, 664, 675
sister, 354

SUBJECT INDEX

1235

Chromatin, 1096

Chromatolysis of ganglian cells, 995
Chromonema effects, 801, 802, 814
Chromophil substance, 1095
Chromophoric groups, 74, 84, 289
Chromosome aberrations, 289, 324-326,
334, 338, 341-342, 627, 628, 717, 798,
825, 1135
in animal cells, induced, 627, 629, 630,
632-635, 642, 645, 648, 650, 652,
653, 655, 656, 659, 666, 669, 671,
673, 674, 676, 682-690
in mammals, 826, 831, 832, 835-837,
839, 842, 843, 854, 855
and radiation damage to embryo,
902-904, 913
in microspores, 715
oxygen tension, 940
radiation-induced, effect on, of centrif-
ugation, 737
of colchicine, 737
of sonic vibration, 737
in Tradescantia, 713-761
regenerating liver, 1128
tumors, 338
Vicia faba, 306
Chromosome breaks, 124, 132, 135, 136,
138, 140, 272, 364, 446, 598, 717,
801, 802, 836, 839, 846, 847
in animal cells, induced, 628-630, 632-
635, 637, 641, 642, 647, 649-671,
673-682, 687, 688, 690
distribution in chromosomes, 720
and dosage, 482, 724
frequency per cell, 125
healing of, 366, 491
and infrared, 571
intragenic, 497
and ionization, 480-492
isochromatid, 490
multiple, 366-377
point mutation, 496-507
restitution of, 363, 483, 499, 500, 722
reunion of, 722, 730, 749
single, 362-366
Chromosome changes, in form, 801, 814

nonrandom incidence of, 377-380
• and ultraviolet radiation, 531-535
Chromosome exchanges, 717

relation of, to fast-neutron dose, 726
to radiation intensity, 727
to X-ray dose, 725
Chromosome matrix, 555
Chromosome matrix effects, 814
Chromosome number, in Drosophila
melanogaster, 631, 642
in Tradescantia, 631

Chromosome rearrangements, 132
in animal cells (see Chromosomes)
gross, and ionization, 380, 480-488
minute, 380
Chromosome ring, 372, 497, 498, 500, 514
Chromosome stickiness, 777, 779, 798-
801, 814, 1097
dose and, 800
thymonucleic acid depolymerization

and, 800, 801
time after treatment and, 798-800
Chromosomes, 352, 355, 1096
acentric, 363, 364
adhesive ends, 446
aneuploidy of whole, 358-359
in animal cells, adhesion, induced, 694,
695
chromatids, 628, 632, 642, 647-650,

659-661, 664, 675
deficiencies, 640-642, 645-650, 661-

663, 667-670
division, 628, 677-680
duplications, 636, 637, 640-642, 645,

648-650, 660, 662, 689
fragmentation, induced, 628, 629,

631, 632, 659, 665, 668
inversions, 635-638, 647, 650, 651,

654, 661-663, 670, 677-680, 685,

687, 689

isochromatid breaks, 632, 660
neuroblast, 635, 638, 648, 651, 659,

660, 666, 667, 679
rearrangement, 627, 629, 630, 632,

635-637, 641, 642, 649-657, 660,

661, 663-666, 668-681, 684, 685,

688, 689

recombination, 630, 642, 649, 652-

655, 660, 661, 663, 665, 670, 671,
674-680, 682, 687, 690

reconstitution, induced, 628, 629
salivarv-gland, 447, 494-496, 507,
635-637, 646, 648-652, 660, 661,
663-665, 668-670, 673, 677-679,
689
translocation, induced, 635, 637-642,
651, 652, 659, 661-663, 669-671,
673, 674, 679, 680, 685, 687, 689
transpositions, 650, 651, 689
arrangement in sperm, 520
balancing, 457
breakage-fusion-bridge cycle, 365, 451,

456, 515, 560
bridge formation, 482
centric, 363, 364
and deficiency, 449
deletion in, 371, 372
dicentric, 364
distribution, 630

1236

RADIATION BIOLOGY

Chromosomes, euchromatic region in,
645, 663, 664, 677, 678, 681
extended, 447
fragmented, 1096
heterochromatic regions in, 635, 636,

645, 651, 652, 663-665, 677-681
homologous, 370

indirect radiation effects, 753, 754, 756
intranuclear disorientation, X-ray-in-
duced, 804
inversion in, 374
iso-, 363
metastable, 571
minute structural changes in, by

ionizing radiation, 492-496
monocentric, 370
radiation effects in Tradescantia, 713-

761
"shift" rearrangement in, 376
sticky (see Chromosome stickiness)
structural changes, 362
telomeres, 366, 446
terminal deficiency, 366
time of division, 721
translocation, 999
union of fragments, 446
variations in radiosensitivity, 721
X, 377, 378, 396, 407, 497, 498, 500,
514, 641, 644-648, 650, 654, 661,
663-668, 677, 678, 680, 682, 839,
842
Y, 455, 514, 641, 644, 645, 649, 651,

654, 663, 668, 674, 842
(See also Chromosome aberrations;
Chromosome breaks)
Chronic-radiation effects (see Anemia;

Erythrocytes)
Chylomicron count, 975
Ciliary motion, 1095
Circulation, radiation effects, 960, 976-
982
and radiosensitivity, 940
Citrate, and bone, 1108

in liver, 988
Cleavage, irradiation during, 868
multipolar, X-ray induced, 803
retarded, following irradiation of early
mouse embryo, 866
Cleavage furrow, ultraviolet-induced

change in position, 815, 816
Cleavage time, 304

Clinical literature on prenatal irradia-
tion, 908-911, 913
Clot retraction, effect of irradiation on,

1044
Clotting time, effect of irradiation on,

1047
Cloud chamber, 4

Cluster, 80

Coat color, mammalian, 461
Cobalt, 946

Cobalt-induced polycythemia, effect on,
of phosphorus32, 1065
and X radiation, 1067
of X radiation, 1067
erythropoiesis in, 1065
Cocarcinonogenesis factors, 1186-1188
Cohesive forces, 31
Colchicine, 577, 578

effect on radiation-induced chromo-
some aberrations, 737
Cold environment, 935
Collagen changes, 1175
Collisions, average energy dissipated, 55
cooperative action, 138
distance, 77

law of probability, 79-80
elastic, 42, 45, 49, 50
charged particles, 42
energy dissipation by, 67
neutrons, 74
fast, 50

of first kind, 200
glancing, 51, 56, 62

characteristics of, 63
inelastic, 49, 62

energy dissipation by, 51
mechanism of, 50
probability of, 55
interval, 136
knock-on, 51, 56, 63
neutralizing one another, 140
radiative, 42, 49, 50, 63, 95
relative frequency, 55-56
resonance rule in, 205, 207, 208, 220,

235, 237-238
of second kind, 200, 202-208, 219-220,

228-229, 237-238
slow, 50

spin-conservation in, 205, 208, 220
Colloid retention, 978
Coloboma following prenatal irradiation,

879, 883, 887, 902, 909
Colon, 982, 1124

carcinoma induced by Y91 in, 1159
Colony formation, inhibition of, 330-331,

333
Columnar epithelium of digestive system,

1124
Compound (heterozygote), 404
Compound nucleus, 58
Compton effect, 38, 69
Compton scattering (see Scattering)
Conjunctiva, 1131

radiation effects, 998
Con variant reproduction, 352

SUBJECT INDEX

1237

Conversion, internal, 34
Copper sulfate, 414
Coproantibodies, response, 1003
Coproporphyria 988
Cornea, epithelium, 926
radiation effects, 998
Corpora lutea, 1117, 1118
Corpuscular radiation (see Radiation)
Cortisone, effect of, on leukemogenesis,
1164, 1188
on mouse embryo, 907
Corynebacterium creatinovorans, 301
Cosmic rays, 3
Crepis, 363, 589

Critical periods for induction of abnor-
malities by prenatal irradiation,
880-887, 891-893, 901, 902, 904, 906,
909, 912, 913
Crocus, 774
Crop plants, 442
Cross circulation, 926
Cross section, definition of, 37n.
Crossing over, 356, 359-362, 508

in Drosophila, 635, 638, 642, 653, 654,

668
somatic, 458
Crypt epithelium of intestine, 982
Crystal diffraction, neutrons, 115

X rays, 105
Cumingia, 784
Cumulative dose, 766, 770
Cumulative effect, 1136
Curie (unit), 13
Current, oscillating, 20, 30, 33
frequency of, 30
resonating, 17, 20
Cyanide, 551, 562, 577, 580
of potassium, 576
and radiosensitivity, 941, 945
Cyclotron, 12

Cysteine, 295, 304, 551, 579, 586
liberation of H,S from, 267-268
modification of radiation injury by,
1067, 1068
and estradiol, 1068
and estrogens plus spleen shielding,

1079, 1080
and/or spleen shielding, 1080
protection by, 942-945, 965

mechanism of protective effect, 1067,
1068
Cytochemical techniques and analysis in

radiobiology, 692, 693, 695
Cytochrome c, 287
Cytochrome oxidase, 544, 550, 551, 562,

576
Cytochrome reductase, 941

Cytological techniques and analysis in
radiobiology, 632, 635, 638, 648, 673,
696
Cytoplasm, decrease in viscosity of, 1095
injections of, 928
and nucleus, 928-929

relative radiosensitivity of, 1094
Cytoplasmic inheritance, 595, 596
Cytoplasmic movements, 1095

D

Daily irradiation of ovary, 1118

Datura, 391

Deamination, of amino acids, 296

glycine, effect of alpha radiation on,

271
by X rays, 265-266

effect of solute concentration, 268-

270
pH dependence, 266-267
Death, genetic, 426, 430

(See also Mortality)
Decrease in viscosity of cytoplasm, 1095
Decticus, 358
Deficiency, 449, 854, 855
Degenerative phenomena, 1092
cartilage cells, 1112, 1113
epidermis, 1121
Dehydration, 969
11-Dehydrocorticosterone, 992
Delayed effects, 1097
Deletions, 480, 498, 842
8 rays, 81, 271

Dermatitis, radiation, 919, 1148
Desoxycorticosterone, 992
Desoxyribonucleic acid (DNA), 305
chromosome breakage, 723
radiation effects on, 276, 277, 753-755
depolymerization, 277
inhibition of formation, 276
oxygen and radiosensitivity of, 755
synthesis, 929, 962
Desoxyribonucleic acid depolymerase,

299
Desquamation, 338
Deuterons, 3
atomic weight, 7
charge, 7
Development, 458-462

mammalian prenatal, broad divisions
of, with respect to radiation
response, 862-863, 886, 911
radiation effects on, 861-918
Developmental anomalies, 861-918, 1185
Devitalized bone, 1111
Diagnostic irradiation and hazards to
embryo, 910, 913

1238

RADIATION BIOLOGY

Diarrhea, 969, 984, 1127
Diazomethane, 547
Dicumarol, 967
Diet and radiosensitivity, 933
Differentiation, 459

as related to radiosensitivity, 927
and type of radiation damage to
embryo, 902-903
Digestive system, columnar epithelium

of, 1124
Dilution effect, 259, 274
Dinitrophenol, 551, 935
Diphenyl, 290

Diphosphopyridine nucleotide, 288, 289
Diploid, 354

Diploid cells, 642-644, 647, 683
Direct action theory of radiation (see

Target theory)
Direct effects of radiation (biological),
245-252
on chromosomes, 735, 753
Disability, average, 432
Disintegration, 32, 35

different modes of, 35
Disordering of solids, 196
Dissipation of energy (see Energy dis-
sipation )
Dissociation, 33, 213-219, 224-225, 238-
239, 246, 248
following ionization, 225, 235, 238-239
Dithiophosphonate, 942
Divided doses, 1092
Dog, 863

LD60 for, 930
Dominance, 402-405

effective, 427
Dosage compensation, 401, 449
Dosage rate, effects of, 924

on chromosome aberrations, 727
lethal, on cells, 810
mitotic, 788-793, 813
Dose, 120

cumulative, 766, 770
divided, 1092

effect of varying, in prenatal irradia-
tion, 882-887, 904-907, 913
and mitotic effects of ionizing radia-
tion, 769, 772, 782, 783
permissible average or total, 433, 434
Dose action (see Dose effect)
Dose effect, exponential curves, 123-125,
130, 131, 135, 136
range of validity, 125-127
shape of, 326

theoretical significance, 126
sigmoid curves, 132, 135
Dose-effect relations, 123

Dosimetry, 145

comparison of units, 149, 150

of internal emitters, 166-174

of ionizing particles, 164-165

by means of ionization in gases,

148-150
of neutrons, 161-164
of photons, 151-154, 177
practical aspects of, 174-185
ionizing particle, 174-177
photons, 181-183
survey of dosimetric methods, 146-148
of very-low-energy X ray, 179-180
(See also Dosage rate; Dose; Radiation)
Drosophila, 324, 325, 333-335, 358-363,
373, 375, 377, 378, 380-383, 387, 389,
390, 392, 394-396, 399, 403, 405-
412, 414, 416-423, 427, 437, 438, 443,
447-450, 452-455, 457, 458, 460,
476-479, 481-486, 492-494, 497,
502-507, 509, 512, 513, 516, 518, 519,
521, 524-526, 529, 531, 532, 534, 535,
539, 542, 545-547, 551, 557, 561,
565-572, 575-578, 581-584, 589, 595,
600, 738-740, 746, 807, 924, 935, 936,
939, 1005
carcinogenesis in, 1190
crossing over, 635, 638, 642, 653, 654,

668
eggs, 682, 687, 689
embryos, 683
funebris, 410

genetic effects in, compared with
mammals, 825, 835, 836, 842, 844,
846, 847, 849, 854-856
melanogaster , 410, 456, 553

chromosome number in, 631, 642
simulans, 410, 456, 553
spermatogonia, 685, 686
spermatozoa, 631, 632, 635, 644, 669,

670, 673-676, 682, 686
stocks, 457
testes, 685, 686
Drugs, parasympatholytic, 985
Ducklings, radiation effects, 925
Duodenum, 982
Duplication, 353
Dye, 289, 292, 295
Dyspnea, 995

E

Ear cartilage, 1000
Echinoderms, 357, 811
Ectopic erythropoiesis, 963
Ectopic myelopoiesis, 1103, 1106
Edema, 971

intestinal, 983

lymphatic organs, 1103

SUBJECT INDEX

1239

Effective areas, 128
Effective volume, 128, 130, 137
Electric relaxation, 69
Electrocardiograph, 981
Electrolytes, metabolism, 969
Electromagnetic radiation (see Radia-
tion)
Electron attachment to neutral mole-
cules, 240-241
Electron tracks, 90
Electron-transfer processes, 232, 237,

242-243
Electron volt, 8

Electronic equilibrium (X rays), 111
Electrons, atomic weight, 6
average deflection, 92
backscattering, 91
capture, 69
charge of, 6

energy dissipation at various depths, 93
energy distribution, 90
excitation of atomic, 30
negative, 6
penetration, 90, 99
penetration data, 100
positive (see Positrons)
primary, 57
recoil, 38
secondary, 21, 57, 69, 138

energy dissipation by, 68
sources of, 10
Electrophoretic mobility, change in, 328
Electrophoretic pattern of plasma, 973
Electrostatic accelerator, 11
Elementary processes, 26, 45
chains of, 62
free atomic particles, 44
in mercury vapor, 199, 200, 202, 206-
208, 215
Embryo, human, effects of radiation on,
908-911, 913
mammalian, 861

irradiated during period of major
organogenesis, 862, 868-893, 909-
910, 912
irradiated in preimplantation stages,

862, 864-868, 911-912
irradiation, of individual, 864, 901
of selected parts of, 864, 877, 889-
890, 911
mechanism of radiation effect on,
900-908, 913
role of maternal body in, 861,
900-901, 911, 912
ra iation effects, 925
rat iosensitivity, 932

hypoxia and, 904-906, 913

Embryo mash, effect on radiation injury,

1074
Embryology, mammalian experimental,

radiation effects and, 861-862, 911
Embryonic precursor, radiation effect on,

902-904, 906, 908, 913
Emitters, internal (see specific elements)
Endocrine imbalance and carcinogenesis,

1167, 1170
Endocrine organs, carcinogenesis in,
1167-1172

radiation effects, 989-994
Endosteal growth and bone, 1108
Endothelium, damage to, 978
Endotoxins, bacterial, 1003
Energy, activation, 210, 215-216

average, per ion pair, 65, 66, 158, 165,
166, 237, 241, 251

binding, 28, 60

of chemical bond, 28, 31, 60

migration of, 202, 233-234, 246n.,
248n., 249
Energy absorption, linear, 316

and neutron flux, 163

per roentgen, 150

in tissues, ultraviolet radiations, 764
Energy dissipation, average, in collisions,
55

by elastic collisions, 67

by electrons at various depths, 93

by inelastic collisions, 51

rate of, 87

by secondary electrons, 68
Energy distribution, electrons, 90
Energy flux per roentgen, 159
Energy loss, rate of, 316
Energy scales, 9
Energy transfer, linear, 316
Enterase, 987

Enterogenous infection, 982-983, 1002
Entropy, 291
Environment of cells and radiosensitivity,

1135
Enzyme systems, 297
Enzymes, 937, 941

amino groups, 297

double bonds, 297

flavone, 562

and genes, 354, 462

hydroxyl groups, 297

inactivation of, 324, 329

prosthetic group, 297

sulfhydryl, 937

sulfhydryl groups, 297
Eosin, 541

Eosinophils, effect of irradiation on, 1063
Epidermis, 1097, 1119

cellular polymorphism, 1121

1240

RADIATION BIOLOGY

Epidermis, degeneration of, 1121
friable, 1121
latent period in, 1119
loss of elastic fibers in, 1122
regeneration of, 1121
Epidermititis, moist, 997
Epididymis, 1116

Epilation, 337, 919, 997, 1120, 1123
Epilobium, 553
Epinephrine, 977, 980, 991
Epiphysis of bone, 1108
Epithelioma radiosensitivity, 929
Epithelium, intestinal, 982, 983
radiation damage, 960
stratified squamous, 1124
Epoxides, 417
Erythema, 338, 1120, 1121, 1150

of skin, 926, 976-997
Erythroblasts, 1009

sensitivity to radiation, 930, 1036-1037
Erythrocytes, 960-901, 970, 974, 979,
1002
and bone marrow, 1099
chloride, 974

chronic-radiation effects, 1048, 1049,
1136
and cutting of distal vessel of spleen,

1069
on fragility, 961, 1039

of red-cell mass, 1039, 1040
and hemolysis, 1039
of gamma radiation, 1050-1052,

1054-1056
of radioisotopes, 1056, 1061

strontium89 and splenectomy, 1060
and spleen shielding, 1038
of X radiation, 1037-1041, 1067
in phenylhydrazine-induced ane-
mia, 1065
disappearance of, 979
fowl, metabolism, 1002
iron59 uptake in, 1037
mass, 960

morphological changes in, 1061-1062
nucleated, 305
tagged, 970
Erythropoiesis, ectopic, 963

induced by phenylhydrazine and/or
bleeding, 1064-1066
inhibition of, 1037-1039, 1047, 1065-
1066
and spleen, 1107, 1108
Erythrosine, 542

Escherichia coli, 417, 418, 420, 454, 537,
542-546, 549, 564, 572, 579, 587, 591,
592, 935, 940, 944
Esophagus, 1124
Ester linkages, 290

Esterase, 300

Estradiol, effect of, on peripheral blood,
1067
and modification of radiation injury,
1066-1067, 1077-1080
by cysteine, 1080

and/or spleen shielding, 1079,
1080
Estrogens, treatment with, 935, 965

(See also Estradiol)
Estrus suppression, 337
Ethanol, 942, 946
Ethyl alcohol, 580
Euchromatic region in chromosome, 645,

663, 664, 677, 678, 681
Euchromatin, 361, 450
Evans blue, 971, 978
Evaporation, 36

Exchange reactions, 205-206, 220
Excitation, 60, 193-194, 197-234, 920
of atoms, 30, 197-208
in condensed systems, 229-234
of diatomic molecules, 209-220
effects of, 32
energy sharing, 32
interatomic motion, 32
of polyatomic molecules, 220-229
Exercise, 934, 937, 946, 1005
External beta rays and thymus, 1105
Extracts, cell-free, effect of, 1082
Extremities, protection of, 926
Eye, latent period, 1131
in lens, 1136
radiation effects, 943, 998

damage following prenatal irradia-
tion, 879-881, 883, 887-889,
891-892, 896-898, 909, 912
(See also specific abnormalities)
gamma rays, 1131
neutrons, 1131
X rays, 1131
Eye pigments, insect, 461
Eyelid carcinoma caused by Th02, 1158

Fallopian tubes, 1117
Fast neutrons, 1033, 1042-1044, 1048
anemia induced by, 1043
biological effectiveness of, and X rays,

1048
comparative effect of, and X rays, 1044
effect of intensity, 723Jf.
effects of, on abortive rise, 1043-1044
on chromosomes, 723^.
modified by oxygen, 747, 752
on morphological cell changes, 1044
on peripheral blood, 1033, 1042, 1043

SUBJECT INDEX

Fast neutrons, relation of aberration
yield to dose, 726
relative efficiency in chromosome-
aberration production, 732
toxicity of, 923, 961, 998
Fat, absorption of, 986

in cells, 1095
Fat cells, bone marrow, 1100
Fat infiltration of liver, 987
Fatty acids, 298
Fenton reagent, 290
Fern, 576, 786, 796
Fern spores, 322, 324, 333, 936, 939
Ferric sulfate, 575
Ferricytochrome c, 289
Ferrocytochrome, 287
Ferrocytochrome c, 307
Fertility, effects of radiation, in mam-
mals, 826-831
effects of prenatal irradiation on, 889-

890, 896-898, 909, 912
(See also Mutations, partial sterility,
sterility; Sterility)
Fetal period, irradiation during, 862, 863,
893-900, 912
and time of parturition, 898-899
Fetus, human, effects of radiation on,
908-911
mechanism of radiation effect on,

900-908
(See also Fetal period)
Fibrinogen, 965, 967, 973
Fibroblasts, 321, 335-336
Fish, LD6o for, 930
Fission, 14, 36

Fission products and intestine, 1127
Flavone enzymes, 562
Flavonoids, 968-969, 979
Flavoproteins, 551
Flower pigments, 461
Fluids, body, 969-976

extracellular, of intestine, 984
Fluorescein, 978

Fluorescence and phosphorescence, 22,
46, 201-202, 216-217, 224, 227, 230,
231, 233, 246
quenching of, 202, 203, 206, 207, 231
sensitized, 203, 207, 228, 237re., 248n.
X-ray, 47, 69
yield, 107
Fluoride, 941, 946
Fluoroacetate, 946, 988
Fodder, 445

Follicular epithelium of ovary, 1117
Food consumption, 984
Food production, 445
Formaldehyde, 417, 422, 547
Formate, 581

1241

Formic acid, 288
Fractionated doses, 925

effects on chromosome-aberration vield
729
Fragility, capillary, 1044

vascular, 979
Franck-Condon principle, 204-206, 208
214-218, 220, 223-224, 227, 235, 238
Frog eggs, 935
Frog radiosensitivity, 936
Fructose phosphorylation, 986
Fucus, 543

Fungi, 324, 333, 415, 444, 462, 505, 531,
535, 543, 557

G

Galactose, 304
Galactozymase, 304
Gall bladder, 1128

carcinogenesis in, 1154, 1156, 1184
Gallium72, effect on peripheral blood,

1061
Galls, 444

Gamma radiation, 408, 518, 521-523
anemia induced by, 1050-1052, 1054-
1056
effect of, on bone marrow, 1056
on peripheral blood, 1049-1056
on survival, 1052
and eye, 1131
genetic effects of, in mammals, 825,

830, 847
genetic hazards of, 854-856
and lymph nodes, 1104
and mutation, 479, 487
pancytopenia induced by, 1055, 1056
and spleen, 1107
and testis, 1114, 1116
7 rays, 3

anemia induced by (see Gamma

radiation)
definition of, 20n., 21
energy release in tissues, 764, 785, 786
lethal effect on cells, 805, 808-810
measurement of, 318-319
mitotic effects, 771, 776-779, 782, 784,

786, 787, 789-794, 797
and mutations, 479, 487
ratio to X rays, 923, 976
secondary, 117
sources of, 26

(See also Gamma radiation; X rays)
Ganglion cells, 995
Gastric acidity, 985-986
Gastric atrophy, 985-986
Gastric secretion, 1125

1242

RADIATION BIOLOGY

Gastrointestinal tract, 982-986
carcinogenesis in, 1184
fluid content, 972
Geiger counters, 4, 204
Gelatin, viscosity of solutions, 295
Gels, 292
Genes, 352
accessory, 402
as catalyst, 353
conjugator, 447

dosage, effects of changing, 396-402
and enzymes, 354, 462
equilibrium frequency of, 430
linkage of, 638, 642
marker, 406
modifying, 401
mutations (see Mutations)
mutator, 414, 496
noncompoundness of, 601
and nucleic acid, 353
number of, estimates, 407
primary, 402

sensitive volume of, 526, 527, 529
shape of, 527
size of, 527, 528
Genetic death, 426

overlapping in, 430
Genetic effects, 135, 136

of radiation, in mammals, 825-859
proportionality to dose, 126
Genetic hazards of radiation, 854-856

(See also Radiation)
Genetic "load" or disability, 427
Genetic material, fundamental proper-
ties of, 352-354
transmission of, 354-357
Genetic-strain differences in response to
prenatal irradiation, 872, 906-907,
913
Genetic technique and analysis in radio-
biology, 632, 635, 638, 639, 641, 668,
670, 673, 677
Genetics, developmental, 459
Genomeres, 452

Germ-cell age and mutation, 504, 552
Germinal centers, 1103, 1106
Germinal selection, 408, 502, 685, 687,

689, 837-839, 846
Germinal tissues, radiosensitivity of, 929
Germination inhibition, 331, 333
Gesonia, 385

Gestation period, length of, and prenatal
irradiation, 898-899, 912
survey of radiation effects on entire,
862, 880
Globulins of plasma, 973
Gluconeogenesis, 974

Glucose, 303, 942, 946

absorption of, 307, 986

of plasma and lymph, 974
Glucosides, flavonol, 968-969
L-Glutamate, 306
Glutathione, 287, 298, 299, 304, 308

liberation of H2S from, 267

modification of radiation injury by,
941-945, 1067
Glutathione reductase, 308
Glycerine, 580
Glycerol, 946
Glycine, 296, 297
Glycogen, 1095

metabolism, 987
Glycols, 580
Glycolysis, 305

aerobic, 306, 941

anaerobic, 306

bone-marrow, 305

inhibition of, 339
Glycosidic linkages, 290
Goat, LD60 for, 930
Gold, colloidal, 979, 988
Gold198 irradiation, 926

effect of, on peripheral blood, 1060,
1061
on sedimentation rate, 1061

toxicity of, 988
Goldfish, LD5o for, 930
Golgi net, 1095
Gonadotrophic response, 999
Gonads, 325

carcinogenesis in, 1168-1171

radiation effects, 998-999
Gradient of injury, bone marrow, 1101
Granular leukocytes and bone marrow,

1099
Granulocytes (heterophils), 965
Granulocytopenia, 935
Granulocytosis, 960, 964
Graphite hydrosol, 324, 328
Grasshopper (Chortophaga), 484, 486,
569, 628, 629, 632, 659, 660, 666, 667,
682, 685, 766, 768, 772, 774, 776, 777,
779-781, 783, 791-796, 801, 803, 806,
808, 811-815
Grasshopper eggs, 301
Graying of hair, 997-998
Growing follicles of ovary, 1117, 1118
Growth, 459

and oxygen tension, 940

retardation of, 339, 340, 869, 876-877,
879, 896, 900, 905, 912, 922
Guanidine of blood, 1000
Guinea pigs, 825, 826, 828, 831, 833, 836,
838, 839, 863

SUBJECT INDEX

1243

Guinea pigs, anemia induced by gamma
radiation in, 1052, 1054-1056
survival of, 1052
broad developmental divisions of, 863
carcinogenesis in, 1152, 1153, 1155,

1164, 1174
effect of bone-marrow injections from,

on irradiated mice, 1075
irradiation of, during fetal period,
894, 897-898
during period of major organo-
genesis, 869, 874, 879, 880
during preimplantation period, 864-
866
LD6o for, 930-931, 969
Gynandromorph, 359, 514, 515

H

Habrobracon, 384, 388, 453, 454, 510, 643,

657, 667, 683, 688, 689, 799, 808
Hair, 1119, 1122
epilation, 1123
permanent, 1123
temporary, 1123
graying of, 998
radiation effects, 997
Xrays, 1123
Hair follicles, carcinogenesis in, 1176

destruction of, 1122
Half deaths, 426
Half-value layer, 104
Hamster, LD5o for, 930
Haploid cells, 335, 642-644, 668, 683
Hassall's bodies of thymus, 1105
Hatching, inhibition of, 325, 334-335
Hazards of radiation, human, 854-856,
910, 913
(See also Radiation illness; Radiation
injury)
Head protection, 926

by shielding, 1070, 1072, 1073
Heart-muscle damage, 981
Heart rate, 981
Heavy particles, deflection of, 89

penetration of, 87
Helianthus, 589
Helium, 559
Hematocrit, 964

effect of radiation on, 1037, 1038, 1055
and phosphorus32, 1057

in cobalt-induced polycythemia,
1067
Hematologic effects of ionizing radia-
tions, 1029-1090
Hematopoiesis, radiation effects, 960-969

in testis, 1116
Hematopoietic cells, regeneration of, 1 100

Hematopoietic tissues, radiosensitivity

of, 929
Hemin synthesis, 962
Hemoconcentration, 969-970
Hemocyanin, 295
Hemocytoblasts, 1099, 1107
Hemoglobin, 295

effect of radiation on, 1037-1041

(See also Erythrocytes)
Hemoglobin concentration, 964
Hemolysis, 961
Hemophilia, 460
Hemorrhage, 922, 923

induced by radiation, 1039-1040, 1045-
1047, 1056, 1072

intestinal, 983
Heparin injections, 967
Heparin-like material, circulating, 967
Hepatic blood flow, 977-978
Hertzian waves (radio waves), 3, 14, 20
Heterochromatic regions in chromosomes,
635, 636, 645, 651. 652, 663-665,
677-681
Heterochromatin, 361, 448, 450, 493
Heterochromatin blocks, 378, 447
Heterophil leukocytes, 1103, 1106

abortive rise in, 1034-1036

effect on, of fast neutrons, 1042-1044
of gamma radiation, 1050-1052,
1055, 1056
Heterosis, 429
Heterozygote, 402
Hibernation, 930

radioresistance during, 977
Hiroshima, bone-marrow studies, 1101

leukemia incidence, 1163
Histamine of plasma, 975, 980
Histidine, 292
Histology of damage following prenatal

irradiation, 880, 897, 904, 912
Hit theory (see Target theory)
Homozygote, 402
Hormone imbalance, 1167, 1170
Horse serum, 935
Human cells, 629
Human fetus (see Fetus)
Human radiation effects (see Man)
Human subjects, 338, 339

(See also Man)
Humoral theory of regeneration, 1077-

1081
Hyaline membrane in lung, 1130
Hyalinization of collagen in skin, 1122
Hyaluronidase, 968, 979
Hybrids, introgressive, 456

species, 553
Hydration of ions, 232, 242-243, 247,
250-251

1244

RADIATION BIOLOGY

Hydrocarbons, carcinogenic, 422
Hydrocephalus following prenatal irra-
diation, 879-881, 883, 909
Hydrogen atoms, 285
Hydrogen bonding in protein, breaking

of, by radiation, 250-252
Hydrogen chloride, 291
Hydrogen effect on radiation-induced

chromosome aberrations, 746, 753
Hydrogen ion concentration and radio-
sensitivity, 939
Hydrogen peroxide, 256, 271, 274, 275-
277, 303, 306, 939
formation of, 285, 286, 328
and mutation, 547-552, 581
origin in irradiated water, 750
in relation to radiation-induced chro-
mosome aberrations, 751
Hydroperoxyl radicals, 294
origin in irradiated water, 751
in relation to radiation-induced chro-
mosome aberrations, 752
Hydrosulfite, 942, 945
Hydroxyapatite and bone, 1108
/3-Hydroxyglutamic acid, 292
Hydroxyl groups, 292
Hydroxyl radicals, 285

origin in irradiated water, 750
in relation to radiation-induced chro-
mosome aberrations, 752jf.
Hydroxylation of benzene, 257
Hydroxyproline, 292
Hymenoptera, 454
Hyperesthesia, 996
Hypertension and adrenal irradiation,

991
Hyperthermia, 996

Hypopharynx, carcinogenesis in, 1172
Hypophysectomy, 1005
Hypophysis, 1132
anterior lobe, 1132
growth, 1132
Hypotension, 980

Hypoxia, effect on radiation-induced
genetic damage, 856
and radiation sensitivity of embryos,
904-906, 913

Immunity to radiation effects, 1002-1004

Immunization, 935

Implantation process, radiation effect on,

868
Inactivation of viruses, 324, 329-330
Indirect action of radiations, 960
Indirect action theory of radiation, 257-

260, 272

Indirect effects of radiation, biological,
244-245, 1136
on chromosomes, 753, 754, 756
as related to radiodecomposition of
water, 750
Individual radiosensitivity, 932
Infections, 934, 960, 1002

enterogenous, 982-983, 1002
Inflammation facilitating carcinogenesis,

1186-1187
Inflammatory reaction in skin, 1121, 1122
Infrared effects in combination with X

rays, 739
Inheritance, cytoplasmic, 595, 596
Intensity, duration factor, 925

(See also Dosage rate)
Interference, 356
Internal conversion, 34, 225, 226-228,

246-249
Interphase, 628, 682, 684
Interphase nucleus, 1096
Interstitial cells, 1116-1118
of ovary, 1117, 1118
of testis, 999, 1116, 1117
Interstitial deletions, 717

in relation to X-ray dose, 726
Intestinal absorption, 986
Intestinal edema, 983
Intestines, 929, 982-983, 1006, 1124
acquired radioresistance of, 1127
duodenum, 1125
effect on, of, Ba140-La140, 1127
of fast neutrons, 1126
of fission products, 1127
of phosphorus32, 1127
of plutonium, 1127
of radium, 1127
of shielding, 1070, 1072, 1073
of slow neutrons, 1126
of sodium24, 1127
of strontium89, 1127
of yttrium91, 1127
of Zr93-Cb93, (Zr95-Nb95), 1127
epithelium of, 929
hemorrhages, 1124
motility of, 985
small, 306, 307, 1124
ulceration, 1124
Inversion, paracentric and pericentric,

374
Iodine, 414, 552

Iodine131, effect of, on lymphopoiesis,
1057
on thyroid, 1132
toxicity of, 993
uptake, 992-993
Iodoacetate, 941, 946
Ion beams, 3

SUBJECT INDEX

1245

Ion density, 315, 920, 923
Ion pairs, definition of, 5

per cluster, 81
Ionic yield, 259n.

of dry substances, 272
of small and macromolecules, 272
and solute concentration, 268
Ionization, 32, 234-244
biological effect, 920
and chromosome breakage, 480-488

number required, 488-492
in condensed systems, 241-243
definition of, 5
density of, 197, 243-244, 248
dissociation following, 225, 235, 238-

239
and gross chromosomal rearrange-
ments, 480-488
mean energy per ion pair, 65, 66, 158,

165, 166, 237, 241, 251
measurement of, 5
multiple, 196, 243
and point mutation, 475-479
recombination of, 194n., 239-240
spatial distribution of, 194, 197
specific (see Specific ionization)
by X-ray beam, 113
yield, 65
Ionization chambers, 6

air wall or thimble, 154, 157-161
calibration in roentgens, 177-179
correction factors, 155, 156
extrapolation, 176, 182
standard air, 151-154, 179
Ionization potential, 28, 29
Ionizing radiations (see Radiation)
Ions, charge of, 67-68

columnar distribution of, 83
hydration of, 232, 242-243, 247, 250-

251
negative, 80

formation of, 69
positive, 7

sources of, 10
recombination, 68
spatial distribution of, 194, 197
Iron, 574

uptake by bone marrow, 962
Iron59 uptake by rat erythrocytes, 1037
Iron pigment, 1096
Iron porphyrin, 287

Irradiation (see Alpha irradiation; Beta
irradiation ; Gamma radiation ; Radi-
tion; X radiation)
Ischemia, 939, 945
Isochromatid breaks, 632, 660, 717
relation of, to radiation dose, 724
to radiation intensity, 727

Isotopes, 27

radioactive (see Radioactive isotopes)

Japanese bomb casualties, 996, 998, 999,
1005
(See also Hiroshima)
Jaw abnormalities following prenatal
irradiation, 880-881, 892

Karyorrhexis, 1097
Ketones, 417

17-Ketosteroids, excretion, 990
Kidney, 306, 1128

carcinogenesis in, 1154, 1155, 1161

chick, damage to, 989

failure of, 925

radiation effects, 982, 988-989
and shielding, 1070, 1072, 1073
Knock-on collisions, 51, 56, 63
Kynurenic acid excretion, 988

Lactate, 305

Lactation suppression in mammary gland,

1123
Lactic acid formation, 305
Larynx, carcinogenesis in, 1151, 1180
Latent period, 922, 1136

in adrenal, 1132

in epidermis, 1119

in eye, 1131
lens, 1136

in lung, 1129

in lymphatic tissue, 1104

of protein denaturation, 294

in skin, 1121
Lead, 574

Lens, cysteine protection, 943
Lethal action, acute, 336
Lethal dose (LD5o), at birth, following
prenatal irradiation, 875, 912

for mammals, 930

(See also specific animals)
Lethal effects, cell (see Viability effects,

cell)
Lethal mutations (see Mutations)
Leukemia, 1153, 1154, 1162-1167, 1170,
1191

induction of (see Leukemogenesis)

lymphoid, 1164, 1165

myeloid, 1162, 1163, 1165, 1166, 1184

among physicians, 1162, 1163, 1165

in radium workers, 1162

1246

RADIATION BIOLOGY

Leukemia, in relation to myeloid hyper-
plasia, 1164
reticulum cell, 1167
thymic, 1164
Leukemogenesis, by atomic bomb, 1163
effect on, of adrenalectomy, 1165

of cortisone, 1188
in dogs, 1164
dose rate, 1166
factors of, 1165
indirect mechanism, 1165
Leukocytes, 922-923, 930, 935, 960, 964-
965, 977
abortive rise in, 1035, 1036, 1043, 1044
after exposure to fast neutrons, 1043,
1044
acute-radiation effects, 1031-1036
and cutting of distal vessel of spleen,

1070
of fast neutrons, 1033, 1042-1044
of gamma radiation, 1049-1053,

1055-1056
of radioisotopes, 1056-1061
of X radiation and fast neutrons,
1033
adherence to blood vessels, 977
morphological changes in, 1062, 1063
polymorphonuclear, 930
Leukocytosis, 964
Leukopenia, 922, 923
Lewisite, 551

(<See also Mustard gas)
Liberation of H2S, from cysteine, 267-268
pH dependence of, 267
from glutathione, 267
Life span, decrease in, 337
of mammals, 922
shortening of, 1005n.
Light, 3, 14

absorption coefficient of, 118
infrared, definition of, 20
penetration of, 118
primary action of, 74
production of, 21
ultraviolet, definition of, 20
visible, definition of, 20
Light effects, characteristics of, 84
Linear accelerator, 12
Linear energy absorption, 316
Linear energy transfer, definition, 316
Linkages, 356
ester, 290
gene, 638, 642
glycosidic, 290
Lipids, of adrenal cortex, 990

of plasma, 974-975
Liquid solutions, 229-233

Litter size, 826, 830-840, 843, 848, 849,
855

from females irradiated during preg-
nancy, 866, 870-874, 890-891
Liver, 306, 1128

blood flow, 977-978

carcinogenesis in, 1157, 1160, 1184

irradiation of, 978, 982, 986-988

effects of shielding, 1070, 1072, 1073

plutonium and, 1128

regeneration of, 1128, 1135
and radiosensitivity, 929

retention of colloids, 979, 988

sensitivity of, to prenatal irradiation,
880, 886-889, 896-898, 902, 904

thorotrast, 1128

uric acid of, 988

yttrium and, 1128
Liverwort, 531
Local irradiation, 1136

vs. total-body irradiation, 1091
Loci, 356

duplicate, 455

groups of, 454
Locust, 358, 385

Loss of elastic fibers in skin, 1122
Lotus, 583
Luminescence (see Fluorescence and

phosphorescence)
Lung, 1129

bronchial epithelium, 1130

carcinogenesis in, 1153, 1157, 1160,
1177-1180

latent period in, 1129

relative radioresistance of, 1129
Lupin, 567
Lyeopersicum, 776
Lymph, constituents of, 972-976

extravasation of red cells, 966, 968, 971,
979

flow of, 970
Lymph nodes, 960-966, 1101, 1103, 1104

effect on, of fast neutrons, 1104
of repeated X irradiation, 1104
of slow neutrons, 1104
Lymphatic organs, 1101

edema of, 1103

involution, 991

latent period, 1104

Peyer's patches, 1101

radioactive isotopes and, 1104

regeneration of, 1103

solitary lymphatic nodules, 1101

spleen (see Spleen)

tissue, 1101

tonsils, 1101
Lymphatic vessels, dilatation of, 976

SUBJECT INDEX

1247

Lymphocytes, 1101, 1103, 1104, 1133
effect on, of acute radiation, 1031-1036
fast neutrons, 1042, 1043
gamma radiation, 1049-1052, 1055
morphological changes, 1062, 1063
radioisotopes, 1056-1060
Lymphoid tissue, indirect action, 960

nucleic acids, 929
Lymphoma after irradiation, 933
Lymphopenia, 922, 935
Lymphopoiesis, effect on, of appendix
shielding, 1063
of spleen shielding, 1063
and spleen, 1106

M

Macrophages, 1100, 1103

Maize, 363, 410, 450, 454, 457, 531, 533-

536, 540, 560
Malformations (see Morphological
changes following irradiation)
Malonate, 946
Mammalian coat color, 461
Mammals, 415

radiation effects, on embryo, 861-918
genetic, 825-859
on reproduction, 826-831
(See also Radiation)
Mammary gland, 1117, 1123
during pregnancy, 1123
suppression of lactation, 1123
Man, 410
LD50 for, 930

mutations in, 419, 583, 854-856
radiation effects, 338, 339, 919, 966,
998, 999, 1005
on development, 862, 908-911
radiation hazards, 854-856, 910, 913
Manganese, 946
Maps, linkage, 356
Mass number, 27
Mast cells, 1100, 1105, 1107
Maternal body, role of, in radiation effect
on mammalian embryo, 861, 900-
901, 911, 912
Mature follicles of ovary, 1117
Mature sex cells, 1133
Mean square deviation, 80
Mechanism, of radiation effect on em-
bryo and fetus, 900-908, 913
of radiobiological action, 327-328
Megakaryocytes, 1099, 1106, 1108
Meiosis, 354
Meiosis-meiotic cycle, 640, 642-644, 653,

654, 689, 690
Melanoblasts, 998

Menadione, 551, 552

Meninges, carcinogenesis in, 1156

Mercaptocarboxylic acids, 479

Mercaptosuccinate, 942, 945

Mercuribenzoate, 946

Mercury, 946

Mercury vapor, elementary processes in,

199, 200, 202, 206-208, 215
Mesons, 3, 59
Mesothelium, 1130
Metabolic activity, 922
Metabolic processes and gene mutation,

418-421
Metabolic rate and radiosensitivity, 931,

935-937
Metabolism, radiation effects on, 1000-

1002
Metals, heavy, and mutation, 574
Metaphase, 628, 632, 638, 643, 659, 662,

689, 690, 928
Metaphysis of bone, 1108, 1109, 1111
Metastasis facilitated by irradiation,

1174-1175
Methemoglobinemia, 941
Methionine, 942, 945
Methyl-3: 12-diacetoxy-7-keto cholanate,

291
Methylacrylate, 291
Methylcholanthrene, 422
Methylene blue, 289, 945
Methylmethacrylate, 291
Microcephaly following prenatal irradia-
tion, 880, 889, 909
Micrococcus (Staphylococcus) aureus, 537,

545, 548-552, 562
Microglia radioresistance, 995
Microphthalmia following prenatal irra-
diation, 879, 883, 887, 888, 902, 907,

909
Microspores, 341

chromosome aberrations in, 715
Microwaves, 3, 20
Midbrain radiation exposure, 980
Migration of energy, 202, 233-234, 246n.,

248n., 249
Milk agent in mammary tumor induction,

1173
Miners, carcinogenesis in, 1177-1179
Mitochondria, 1095
Mitosis, 289, 354, 1096, 1133, 1135
abnormal, 628-630, 632, 646, 647, 664,

1092
cessation of, 1096
inhibition of, 322, 325, 331-333, 335-

336, 340, 561, 1092
multipolar, 1135
potential, 1133

1248

RADIATION BIOLOGY

Mitosis, rate (frequency) of, 922

correlated with type of radiation dam-
age to embryo, 902, 906, 908, 913
sensitivity of cell, 928
Mitosis-mitotic cycle, 627, 628, 642, 644,
655, 659, 677, 681, 682, 684, 685, 688,
691, 695, 696
Mitotic activity and radiosensitivity, 1 162
Mitotic effects, 766-798, 811-814
of alpha radiation, 785-787
of ionizing radiations, 766-798
acceleration of, 783, 784
calcium and, 796
cell structures involved, 784, 785
chromosome number and, 798
critical period, 768, 769
dosage rate and, 788-793
dose and, 769, 772, 782, 783

for different species, 781, 782
duration of mitotic decrease and,

775-777
on egg, 769

fractionated treatment and, 793, 794
kind of radiation and, 785-788

a rays, 786, 787

|8 rays, 785, 787, 788

y rays, 785-787

neutrons, fast, 786, 787

X rays, 786-788
kind of tissue and, 797, 798
length of mitotic cycle and, 775
mechanism of action, 785
normal mitotic rate and, 778-781
oxygen concentration and, 796
pH and, 796
recovery from, 770, 782

in relation to dose, 782
retardation of, 768, 769, 771, 772,

782, 783
reversion of, 768
sensitivity of different stages, 768,

769, 771-774
on sperm, 769
temperature and, 794-796

during treatment, 794

following treatment, 794-796
type of preparation and, 797
urethane and, 796
methods of study of, 766-768

counting of mitotic stages, 767, 768
timing of cells, 766, 767
of ultraviolet radiations, 811-814
acceleration of, 811, 812
chemical treatment and, 813, 814
intensity and, 813
regression of, 813
retardation of, 811
sensitivity of different stages, 812, 813

Mitotic effects, of ultraviolet radiations,

wave length and, 811, 812
Mitotic mechanism, injury of, and radia-
tion damage to embryo, 902-904
Mitotic spindle (see Spindle)
Modification of radiation effects (see

Radiation injury)
Modifying factors, effects of, on radia-
tion-induced chromosome aberra-
tions, 736ff.
Molds, 391, 399, 417, 444, 461
Molecular chaos, 392
Molecular rays, 3
Mongolism following prenatal irradiation,

909
Monkey, LD60 for, 930
Monochromatic radiation, 16
Monocytes, effect of radiation on, 1033

morphological changes in, 1063
Monosomies, 644

Monsters (see Morphological changes fol-
lowing irradiation)
Morphine, 937

Morphological changes following irradia-
tion, compared with effects of other
agents applied to embryos, 907-908,
913
during fetal period, 863, 895-898
of human embryos, 908-911, 913
intermediate effects of maternal body

on, 901, 911, 912
nature of primary damage leading to,

902-904, 906, 913
during period of major organogenesis,
862, 868-869, 878-893, 909-910,
912-913
during preimplantation period, 862,

864, 866-867, 886
(See also names of specific abnor-
malities)
Mortality, dose curves, 932
neonatal, and irradiation, during fetal
period, 893-894
during period of major organo-
genesis, 869, 874-875, 890, 904-
905, 912
during preimplantation period,
875
postnatal, and irradiation, during fetal
period, 893-895
of human embryo or fetus, 911
during period of major organo-
genesis, 869, 904
prenatal, influence of radiation effects
on mother, 900-901
and irradiation, during fetal period,
862, 893-894

SUBJECT INDEX

1249

Mortality, prenatal, and irradiation, dur-
ing period of major organogenesis,
862, 868-874, 888, 890, 904, 912
during preimplantation period,
862, 864-868, 872, 911-912
Mosaics, 481, 507, 513, 514, 560
in Drosophila, 648, 649, 653, 661
tobacco, 510
Motility of intestine, 985
Motor excitability, 995
Mouse, 336, 337, 390, 432, 459, 561, 583,
776, 777, 797, 806, 809
broad developmental divisions of, 863,

909
carcinogenesis in (see specific site)
effects of radiation, during fetal period
of, 871, 894, 896-899
genetic, 825-859
during period of major organogenesis

of, 869-892, 905-908
during preimplantation period of,

864-868, 871
on reproduction, 826-831
LD6o for, 924, 930-931
mutation rate in, 850-856
Mouth, 1124
Mucous tissue, 1097
Multilobular nuclei, 1096
Multiple-hit, 133

determination of number of hits, 134
Muscle, 1097

carcinogenesis in, 1177

effect of implants of, on radiation

injury, 1073
radiosensitivity of, 929
striated, contraction of, 922
Muscle cell, radiosensitivity of, 1161
Mushrooms, 442
Mustard, nitrogen (see Nitrogen raus-

Mustard gas, 415, 512, 514, 547, 548, 551,
579, 594, 600, 1094
derivatives, 424
Mutagenesis, and biological conditions,
552-555
and chemicals, 574-581
and infrared, 565-574
and physical factors, 565-574
selective, and radiations, 590-597
and temperature, 565-574
and ultraviolet radiation (see Ultra-
violet radiation)
and water and oxygen, 555-564
Mutants, achaete, 507
albino, 590, 591
alboviridis, 590
alboxantha, 590
Bar M2, 493

Mutants, Beaded wings, 390
cubitus interruptus, 481, 487
bithorax, 454
dumpy, 454
histidineless, 537, 542
inositol, 594
kappa, 595
lozenge, 454, 507
Minute bristles, 449
mottled eye, 595
Notch, 494, 495
phage, 595
purine, 564

R and Q type, 556, 591
scute, 452, 493, 507
streptomycin, 564, 587, 591, 592, 595
Stubble, 454
tigrina, 590
Truncate wings, 390
tryptophane, 592
viridis, 590, 591

white, 454, 504, 507, 515, 516, 526
yellow, 504, 507
xantha, 590
Mutational spectrum, 590, 594, 596
Mutations, 121, 138, 249, 289, 303, 324,
333, 334, 352
accumulation of, 424-432, 855, 856
and activations, 490
and active radicals, 551
amorphic, 398, 597
antimorphic, 398
biochemical, 462
and carcinogens, 421-424
and centrifuging, 573
clustered, 523, 853
and cosmic radiation, 581-584
cryptic, 404

deficiency, 449, 854, 855
delayed, 512, 514, 600
detrimental class of, 395
in different species, 410-412, 835, 836,

844, 854-856
direct or abnormad, 398
dominant, 381, 402
elimination of. 424-432, 837, 839, 842,

843, 846, 855, 856
and energy levels, 597
expression of, 424-432
and fluorescent substances, 541
fractional, 512
and gamma rays, 479, 487

(See also Gamma radiation)
gene, 389, 394, 598, 630, 638, 675, 825,
843, 854
and abnormalities from irradiation

of embryos, 903, 908, 913
and metabolic processes, 418-421

1250

RADIATION BIOLOGY

Mutations, gene, radiation-induced, 405-

409

in mammals, 825-859

spontaneous, 405-409
and germ-cell age, 504, 552
and heavy metals, 574
and hydrogen peroxide, 547-552
hypermorphic, 398, 597
hypomorphic, 397, 449, 597
lethal, 394, 652, 653, 666, 1133

cell, 838

dominant, 359, 504, 632, 644, 646,
655, 657, 663, 666-668, 670, 675,
676, 682-685, 687-689, 825,
831-840, 844, 847, 854, 856
and abnormalities from irradia-
tion of embryos, 903, 913

recessive, 851-855

sex-linked, 140, 405, 476, 851

somatic, 1135
in mammals, 825-859
in man, 419, 583, 854-856
and natural radiation, 581-584
neomorphic, 381, 399, 597
not radiation-induced, 412-418
and nuclear transmutation, 584-590
and nucleic acid, 540
and oxygen, 561, 856
and ozone, 557, 563
partial sterility, 825, 840, 842-850, 854
point, 389, 394, 411, 825, 826, 836, 837,
839, 855

and ionization, 475-479

relation to chromosomal breakage,
496-507

and temperature, 565

and ultraviolet radiation, 531-542
pre-, 511

and pressure, 573
proportionality of, to dose, 477
radiation, 389-396

spacially limited, 507-511, 517-525

time-limited, 511-517
and radiation duration and intensity,

478
and radioactive phosphorus, 585, 587
and radioactive sulfur, 585
rate of, critical, 433

of immature female germ cells, 552

of individual genes, 407, 853

mammalian, 903

in man, 583, 854-856

and meiotic stage, 566

in mouse, 850-856

of spermatogonia, 552, 850-853, 855,
856

of spermatozoa, 552, 850, 851, 855
recessive, 381, 402, 450

Mutations, reverse or normad, 398, 596
597

semilethal, dominant, 831-840
recessive, 851-855

semisterility, 825, 840, 842-850, 854

sex-linked, 140, 405, 476, 851

somatic, in mouse, 903

and sonic vibrations, 572

specific loci, 850-856

spontaneous, 389-396

in statosphere, 582

sterility, 840-843, 854, 855

sublethal class of, 395

subvital, 831-840, 851, 852, 855

and supersonics, 589

"suppressor," 399, 441

and target hypothesis, 525-531

and temperature, 412

and thermal neutrons, 588

translocations, 842-844, 846-850, 854

transverse induction of, 509

and undernourishment, 576

"unit loads" of, 434

and viability, 395, 853-856

viable, recessive, 851-854

visible, 395, 477, 502, 506
dominant, 850-851, 855
recessive, 851, 852

and X-ray wave length, 478
Myaesthenia gravis, 460
Myelocytes, 1099, 1106
Myeloid cells, maturation of, 964
Myocardial damage, 967, 981
Myofibrils, 1095

Myosin (adenosinetriphosphatase), 298
Myotis lucifugus, 977

N
Nails, 1119
Narcissus, 489
Nasopharynx, 1124
Nausea, 982

(See also Radiation illness)
Necrosis following prenatal irradiation,

897
Nembutal anesthesia, 937, 946
Nephritis, 305

Nerve cells, radiosensitivity of, 929, 930
Nerves, 1097, 1131

peripheral, 996
Nervous system, radiation effects, 994-

996, 1006
Nervous tissue, developing, 996

of embryos, 1131

of newborn, 1131
Neuroblasts, radiosensitivity of, 996
Neurofibrils, 1095
Neurogenic origin of anorexia, 985

SUBJECT INDEX

1251

Neurological damage, 967
Neurospora, 382, 399, 454, 461, 476, 505,
533, 535, 541, 545-550, 562, 576, 585,
586, 594, 596, 751
Neutrino, 3

Neutrons, 3, 27, 318, 320, 479, 492, 517-
524, 526, 528, 588
atomic weight, 7
backscattering, 116
capture, 58, 114, 116
collisions, 113
crystal diffraction, 115
dosimetry of, 161-164
ejection of, 36
elastic collisions, 74
energy distribution in space, 116
energy release in tissues, 786
and eye, 1131
fast (see Fast neutrons)
genetic effects of, in mammals, 825,
827, 830, 833, 838, 841, 845, 846
in n units, 320
no electric charge, 7
penetration of, 113, 114
scattered, 116
secondary, 116

slow, chromosome-aberration produc-
tion by, 733
effect of, on blood-forming tissue,
1030
on bone marrow, 1101
on intestinal mucosa, 1126
on lymph nodes, 1104
on spleen, 1107
on thymus, 1105
relation of, to biological efficiency,
734
sources of, 14

and testis, 1114, 1116, 1117
Neutropenia, 922
Neutrophils (see Heterophils)
Nicotine, 980
Nicotiana, 589
Nitrite, 941, 945
Nitrobenzene, 291
Nitrogen, 559-561, 576
excretion by urine, 1000
nonprotein of blood, 973, 989, 1000
Nitrogen mustards, 285, 424, 507, 577,
922, 967
effect of, on morphological changes in
peripheral blood, 1061
on rat embryo, 907
Nitrophenols, 291
Nondisjunction, 358, 508
Nonspecificity of radiation effects, 1093

on bone marrow, 1101
Nor-di-guairelic acid, 550

Nuclear changes, 60

production of, 62
Nuclear forces, 27

effect of, 44
Nuclear reactions, injuries, 1005
Nuclear reactor, 14
Nuclear resonance effect, 58
Nuclear transformations, types of, 59
Nuclear transmutation and mutation,

584-590
Nucleated red cells, 305
Nucleic acids, 290, 303, 527, 540, 550,
587, 692, 696, 929

intestinal, 983

of liver, 988

synthesis of, 305, 962
Nucleolus, 1096

ultraviolet-radiation effects, 816

X-ray effects, 803
Nucleoproteins, 290, 580
Nucleotides, 498
Nucleus, 1094

budding, 1096

compound, 58

and cytoplasm, 928-929

relative radiosensitivity of, 1094

membrane, 1096

structure of, 27

swelling of, 1127
Nutrition, disturbances of, and blood
effects, 960, 965

and radiosensitivity, 985
Nutritional deficiency, 1005

O

Oenothera, 382, 390, 553, 566
Oligodendroglia, radioresistance, 995
One-hit aberrations, 723
1/r2 law, 85
Onion, 578, 580
Oocvtes, 387, 837, 844, 847
Ootid, 374
Optic anlagen, 458
Organic chemistry, 462
Organic matrix of bone, 1112
Organogenesis, irradiation during period
of major, 862, 868-893, 905, 909-
910, 912
Organs, reduplication of, 325, 335

susceptibility of, to tumor induction,
1161-1162

weights of, 1001
Oscillating current, 20, 30, 33
Osmotic pressure, cellular, 938
Osteoblasts and bone, 1108, 1110, 1111
Osteoclasts and bone, 1108, 1110
Osteocytes, bone, 1108, 1110, 1111

1252

RADIATION BIOLOGY

Ova, 1117, 1118
developing, 930
primitive, 1117
Ovarian tumors, 999, 1151, 1153, 1168-

1171
Ovaries, 1117, 1136

daily irradiation of, 1118
follicular cells of, 1117
interstitial cells of, 1117, 1118
plutonium and, 1118
radiation effects, 994, 998-999
radium and, 1118
sterility, 1117
"Overlapping" in genetic death, 430
Oviduct, 1118
Ovocytes, 1117
Ovulation, 1118

Oxidants in physiological effects of radia-
tion, 921
Oxidation and mutations, 417
Oxidation-reduction, 256
Oxidation-reduction potentials, 307
Oxidation-reduction systems, 286, 289
Oxygen, 417

atmospheric, 307
dissolved, 284, 286

effect of, on blood-forming tissue, 1065,
1067
on radiation-induced chromosome

aberrations, 736, 744jf.
on radiosensitivity, 927, 939-941
influence of, on high-energy muta-
gensis, 552, 555-564, 577
(See also Hypoxia)
as related, to radiation intensity, 749

to temperature effects, 742
tissue utilization of, 1001
Oxygen concentration, influence on radia-
tion action, 131
Oxygen consumption, 962
Oxygen tension in mammals, 936
Ozone and mutation, 557, 563

Pair production, 39, 44, 69

cross section, 40
Palate, cleft, following injections of
cortisone, 907
following prenatal irradiation, 883,
892, 907, 909
Pancreas, 1128
Pancreatic fibrosis, 460
Pancytopenia, effect of spleen shielding
on, 1069
induced by gamma radiation, 1055-
1056 "
Papain, 299

Para-aminopropiophenone, 941, 945

Parabiosis, 926

Paramecium, 424, 545, 564, 572, 580, 581,

586, 587, 593, 595, 939
Parasites, insect, 444
Parasympatholytic drugs, 985
Parathyroid, carcinogenesis in, 1172
Partial body irradiation, 925
Pathology, 458-462
Penetration, 84-119

/3 rays in thick foils, 100
electrons, 90
data on, 100
thick foils, 99
thin foils, 99
exponential law, 103-104
heavy charged particles, 87
light, 118
neutrons, 113, 114

narrow-beam, 114
X rays, 102

broad-beam, 108
narrow-beam, 103
Penicillin, 1003
Penicillin resistance, 303
Penicillium, 444, 535, 594
Pentaploid cells, 643
Pentaploidy, 357
Pepsin, 299

Periosteal growth and bone, 1108
Permeability, vascular, 978-979
Peroxides, 417, 937, 939, 941

organic, and mutation, 303, 550, 752
(See also Hydrogen peroxide)
Persistence, 426
Petechiae, 966
Peyer's patch, effect of shielding, on

lvmphocytes, 1063
pH, 575
Phenol, 290
Phenotype, 396

and gene dosage, 401
Phenylhydrazine, 935, 965

stimulation of erythropoiesis by, 1064-
1066
Phenylhydrazine-induced anemia, effect
of X radiation on, 1064-1066
mechanism of recovery from, 1067,
1068
Phenylpyruvic amentia, 461
Phlebotomy-induced anemia (see Phenyl-
hydrazine-induced anemia)
Phosphate, inorganic, 290
Phosphatase, alkaline, 987
of intestine, 983
of plasma, 975
Phosphoglyceraldehyde dehydrogenase,
298, 299

SUBJECT INDEX

1253

Phospholipids, liver turnover, 988

of plasma, 975
Phosphorus (P32), 305, 476
effect of, 1057, 1058
on bone marrow, 926
on erythropoiesis, 1065
external irradiation by, 997
on intestine, 1127
on polycythemic rats, 1057
on testis, 1116, 1117
inorganic, of plasma, 974
radioactive, and mutation, 585, 587
toxicity of, 988, 1003
Phosphorus content, of blood, 1000

of liver, 987
Phosphorus32-tagged erythrocytes, 970
Phosphorylation of fructose, 986
Photochemical reactions, 46, 193, 194
primary processes in, 195-196
sensitized, 195, 208
Photoelectric effect, 18, 20, 46, 69
Photoelectric threshold, 18
Photoelectrons, 20

Photon-electron equilibrium, 150, 183-185
Photon flux per roentgen, 159
Photons, definition of, 19

dosimetry of, 151-154, 177
Photoreactivation, 543, 569
Phycomyces, 922
Pigment, loss of, 997-998
Pigment granules, 1095
Pigment tissue, 1097
Pigmentation in skin, 1120, 1121
Pile, 14

Pilocarpin, 576
Pisum, 776

Pituitary-adrenal system, 1004-1005
Pituitary damage following prenatal

irradiation, 877, 889
Pituitary gland, 991-994
Pituitary tumor, 1167-1169, 1171
(See also Radioactive elements)
Placenta, degenerative changes in, fol-
lowing irradiation, 901
Planck constant, 18, 30
Plankton, 445
Planorbis, 783
Plasma (see Blood plasma)
Plasma cells, 1103, 1116, 1133
effect of radiation on, 1033
Plasma proteins, iodinated, 970
Plasma volume, 961, 969
Platelet transfusion, 968
Platelets, effect on, of fast neutrons, 1043,
1048
of gamma radiation, 1050, 1051,

1054-1056
of radioisotopes, 1056-1061

Platelets, morphological changes in, 1062

thrombi, 977
Pleura, 1130
Ploidy, 683

Plutonium239, effect of, on bone, 1110, 1112
on intestine, 1127
on liver, 987, 1128
on ovary, 1118
on peripheral blood, 1060
on skin, 1122
on spleen, 1107
on testis, 1117
on thymus, 1105
toxicity of, 964, 970, 973
Pneumococci, 353
Point heat, 62
"Point sensible," 283, 284
"Poison" produced by radiation (see

Toxins)
Poisons, effects of, on embryonic devel-
opment, 862, 907-908, 913
Poisson distribution law, 80, 133
Polar cap, 531
Polar force, 31
Polarization effects, 75
Pole cells, 534, 536, 538, 539, 545, 546
Pollen, 341-342, 936
Polonium sources, 320
Polycythemia, cobalt-induced, effect of

phosphorus32 on, 1065
Polydipsia, 971
Polymerization, 290, 291, 292
acetylene, 291
acrylonitrile, 257
Polymorphonuclear neutrophils (see

Heterophils)
Polyphenol, 290
Polyploidy, 357-358, 1097
and radiosensitivity, 929
Poly sondes, 644
Position effects, 376, 380-383, 450, 487,

502, 503, 505, 507, 516, 849
Positrons, 7

atomic weight, 6
charge of, 6
penetration of, 98
sources of, 10
Posture abnormalities, 995
Potassium, 581
cyanide of, 576
excretion of, 989
of intestine, 984
permanganate of, 417, 552
. of plasma, 974

in tissue, 972
Potency, 8, 61, 135-136
corpuscular radiations, 8
electromagnetic radiation, 18

1254

RADIATION BIOLOGY

Potential, appearance, 239, 240
Potential barrier, 35
Potential curves, 211-215, 226-227
Potential surfaces, 220-226
Poultry, 459

(See also Chickens)
Precursor, embryonic, radiation effect on,

902-904, 906, 908, 913
Predissociation, 218-219, 224-225, 227,

246
Pregnancy, irradiation during, 932
and mammary gland, 1123
termination of, following irradiation,

867, 870, 910, 912
unsuspected, and radiation-induced

abnormalities, 910, 913
(See also Abnormalities ; Embryo ; Fetus)
Preimplantation period (of embryo),
irradiation during, 862, 864-868,
875, 886, 911-912
Pressure and mutation, 573
Primitive ova, 1117
Primitiveness and radiosensitivity, 930
Primordium damage as result of prenatal

irradiation, 887n., 902-904, 913
Progynon, 576
Propane-l,3-dithiol, 287
Prophase, 628, 635, 638, 643, 648, 653,

660, 664, 682, 688-690, 696, 928
Propylene glycol, 305
Protamine, 967-968
Protecting agent, 307
Protection, by hypoxia against radiation
damage to embryo, 904-905, 913
from radiation effects, 245, 934, 1006
ionizing radiations, 656
Protection effect, 260-262, 274

and reactivity of solutes, 262-264
Protein depletion, 934
Protein molecule, 292
Proteins, 692, 695, 696
breakdown of, 306
concentration of, 293
degrees of reactivity, 292
denaturation of, 292, 293. 294, 308
latent period of, 294
by radiation, 250-252
hydrogen bond rupture, 292
of intestinal cells, 983
of lymph, 972-973
molecular weight, 292
of plasma, 972-973
prevention of enzyme inhibition, 307
shape, 292

side groups, 292, 295
solutions, 292, 294, 295
spiral structure, 295
subunits, 195

Proteins, synthesis of, 305
Prothrombin-concentration studies, 1046
Prothrombin time, 967
Protons, 3, 27

atomic weight, 7

charges of, 7

ejection of, 36

energy release in tissues, 764, 786

range of, 87

sources of, 10
Protozoa, 303, 444
Primus, 553

Pseudocentrotus, 771, 772, 785, 787
Pulse rate, 981
Purine, 290
Purine bases, 290
Purine derivatives, 578
Purine ring, 290
Purpura, 966
Pyknosis, 1096
Pyrimidine ring, 290
Pyruvate, 305, 306

Q

Quality of radiation in physiological

effects, 922
Quantity of radiation in physiological

effects, 922

R

Rabbit, 336, 554, 629, 825, 831, 833-835,
837. 863, 900-901
bone-marrow glycolysis, 305
broad developmental divisions of, 863
carcinogenesis in, 1152, 1153, 1155,

1164, 1174, 1183
irradiation of, during fetal period,
893-896, 898
during period of major organogenesis,

868-869, 876, 879, 880, 891
during preimplantation period, 865-
866
LD6o for, 930-931
time of death, 932
Radiation, action of, 85

influence of oxygen concentration on,

131
working models, 119, 130
acute, effects of, 1136

on peripheral blood, 1031-1041
by fast neutrons, 1041-1044
by X radiation, 1031-1041
(See also Anemia)
and age, 1093
alpha, 526, 540, 964, 970, 973, 1000

effects of, 271, 940
breaking of hvdrogen bonding in pro-
tein by, 250-252

SUBJECT INDEX

1255

Radiation, classification of, 2

comparative effectiveness of, 135-136
corpuscular, analysis of, 4
classification of, 2
definition of, 2
electric charges, 6
energy transport by, 4
masses, 6

methods of detection, 4
potency of, 8
secondary, 21, 57
types of, 6
cosmic, and mutation, 412, 581-584
damage by (see Radiation injury)
definition of, 2
delta-ray, and mutation, 479
denaturation of protein by, 250-252
direct action theory of, 258, 272
dose (see Dosage rate; Dose; Dose

effect; Dosimetry)
earth, 412
and effect of cell stage on chromosomal

breakage, 383-389
effects of, biological, direct, 245-252
indirect, 244-245

on chromosomes, 753, 754, 756
protection from, 245, 656, 934,
1006
on fertility in mammals, 826-831
(See also Mutations, partial steril-
ity, sterility)
immunity to, 1002-1004
modification of (see Radiation injury)
and temperature, 1093
in water, 244n., 273, 274
electromagnetic, 2

absorption of, 17, 20, 45-48
action of, 16
classification of, 2, 20
detection of, 21
mechanism of, 15
potency of, 18
scattering of, 37, 45-48
gamma (see Gamma radiation)
grenz-rays, 478
hemorrhage induced by, 1039-1040,

1045-1047, 1056, 1072
indirect action theory of, 257-260, 272
infrared, 503

and chromosome breakage, 571
influencing high-energy mutagenesis,
565-574
ionizing, 61, 1029-1082

comparative effectiveness of, 139
degradation of, 61

differences in cell sensitivity to,
681-691

Radiation, ionizing, effects of, on blood
coagulation, 1044-1047

on blood-forming tissue, 1063,
1064
comparative, 1030-1031
direct vs. indirect, 1030
and minute structural chromosome

changes, 492-496
mitotic effects (see Mitotic effects)
and mutations, limited by space,
507-511, 517-525
limited in time, 511-517
protection from, 656
secondary, 63
similar effects, 84
massive doses, 922, 923, 995, 996
and metastasis, 1174-1175
monochromatic, 16
mutation and fractionated doses, 485
natural, and mutation, 581-584
nuclear particle, effect on atomic

nuclei, 58
poison produced by, 245
practical genetic applications of, 440-

445
quality and quantity of, in physio-
logical effects, 922
resonance, 199, 202, 206, 216
secondary, 72

and selective mutagenesis, 490-497
and speed of evolution, 435-440
stimulation of blood-forming tissue by,

1064
as tool for study, of biochemistry,
458-462
of chromosome behavior, 445-448
of chromosome properties, 445-448
of development, 458-462
of gene evolution, 448-458
of gene properties, 448-458
of pathology, 458-462
of physiology, 458-462
ultraviolet (see Ultraviolet radiation)
visible, and mutation, 588
X (see X radiation)
Radiation accidents, 998, 1005, 1006
Radiation action (see Radiation, action

of)
Radiation chemistry, 194

primary processes in, 196-197
Radiation illness, acute, 919
clinical, 966

following irradiation during fetal pe-
riod, 893
of mother and effect on embryo, 190
nausea, 982

1256

RADIATION BIOLOGY

Radiation injury, 553, 934, 1005

mechanism of recovery from, 1069,

1074-1075, 1079-1082
modification of, 274-276, 1064-1077,
1079, 1080
by absence or presence of oxygen,

275
by combined prophylactic and thera-
peutic measures, 1079, 1080
by prophylactic measures, 579, 1064-
1068
cysteine, 1067
estradiol, 1066, 1067
glutathione, 1067
H02, 274, 275
H202, 274, 275
H2S and ammonia, 276
induced hyperplasia, 1064-1066
oxygen deprivation, 275, 1067
sulfhydral compounds, 1067
by therapeutic measures, 1068-1077
blood transfusions, 1081, 1082
embryo mash, 1074
heterologous bone-marrow injec-
tions, 1075
heterologous transplants, 1075
homologous bone-marrow injec-
tions, 1074, 1075
parabiosis, 1075, 1076
spleen implants, 1073-1076
spleen shielding, 1068-1073
splenectomy, and spleen implants,
1073, 1074
and spleen shielding, 1077-1081
tissue shielding, 1069-1073
neurological, 967

in subsequent generations, 432-435
(See also Chromosome aberrations;
Morphological changes following
irradiation; Mutations)
Radiation length, 96
Radiation quantum (photon), 19
Radiation syndrome, acute, 960, 1005-
1008
treatment, 1006
Radiationless transition, 34-35
Radiative collisions, 42, 49, 50, 63, 95
Radicals from water, by irradiation,
256-257, 262, 270, 750
number of, 270

specificity of reaction of, 265-268
Radio waves, 3, 14, 20

and mutation, 588
Radioactive elements, 13

artificial isotopes (P32, I131, Au198, Mn62,

Ga67, Sr89"90), 1158
bone seekers, rare earth, 1160
transuranic elements, 1160

Radioactive elements, carcinogenesis by,
Au198, 1158, 1165, 1167, 1184
I"1, 1167, 1171
ionium, 1158
mesothorium, 1156, 1157
P32, 1148, 1158, 1160, 1166, 1177
Po210, 1148

Pu239, 1148, 1159, 1160, 1177
Ra226, 1147-1148, 1151, 1153-1158,

1160, 1177
radon, 1155, 1158, 1176-1177, 1178,

1180, 1183
Sr89"90, 1160, 1185
thorium dioxide (thorotrast), 1155,
1157, 1158, 1167, 1177, 1179, 1184
uranium, 1177
Y91, 1159, 1177, 1184
maximum permissible amount of [Ra226,
Pu239, Sr89"90 (+Y90), Po210, H3,
C14, Na24, P32, Co60, I131], 1148
strontium, 586
sulfur, 585

(See also Radioactive isotopes)
Radioactive istopes, 1093

artificial (see Radioactive elements)
chromosome aberrations produced by,

733
effect of, on adrenal gland, 1132
on bone, 1110
on bone marrow, 1100
on cartilage, 1113

on gastrointestinal epithelium, 1127
injected into pregnant females, 864,

899-900
on kidney, 1128
on lymphatic tissue, 1104
on osteogenic sarcomas, 1111
on ovary, 1118
on skin, 1122
on spleen, 1107
on testis, 1116
on thymus, 1105
on thyroid, 1132
(See also specific elements)
Radioactive transformations, 13
Radiobiological action, mechanism of,

327-328
Radiobiological additivity, 326
Radiobiology, primary processes in, 194-

195, 197
Radiochemical events in biological sys-
tems, 937
Radiodermatitis, 1151, 1175-1176
Radiomimetic agents, 285, 922, 937
Radioresistance, 1092, 1133, 1135
acquired, 1127, 1136
induced, 934-944
of lung, 1129

SUBJECT INDEX

1257

Radioresistance, of macrophages in bone
marrow, 1100
of reticular cells, 1133, 1135
Radiosensitivity, 1133
alteration of, by combined measures,
1079, 1080
by prophylactic measures, 1064-1068
by therapeutic measures, 1068-1077
by tissue shielding, 1068-1073
biological factors, 927-934
and cellular environment, 1135
of chromosomes, 721, 741, 748
of cold-blooded vs. warm-blooded

vertebrates, 1093
dependent on developmental stage,
861-862, 867-868, 874-877, 879-
882, 906-907, 909, 911
differential, of parts of embryo, 862,
889, 892, 899-900
(See also Critical periods for induc-
tion of abnormalities by prenatal
irradiation)
effect of induced hyperplasia on, 1064-

1066
experimental modification of, 927-934
induced, 934-944
of organs, 1135
physical factors, 927-934
recovery in, 928
in relation to water, 754
relative, 1092, 1094, 1132
Radiothyroidectomy by I131, 1171-1172
Radium, 390, 391

anemia induced by, 1059
calcification of vessels by, 1122
effect of, on adrenal gland, 1132
on bone, 1110-1112
on cartilage, 1113
on intestine, 1127
on peripheral blood, 1059
on ovary, 1118
on skin, 1122
on spleen, 1107
on testis, 1117
and mutations, 478, 581
toxicity of, 1000
Radium workers, leukemia among, 1162

tumor incidence in, 1158, 1182-1183
Radon sources, 321
Raman effect, 46
Rana, 802

Rana spermatozoa, 643
Randomness of atomic events, 26
Range, 87

extrapolated, 100

maximum, 100, 102

relative, of electrons and photons, 111

true, 92, 100

Rate of energy loss, 316
Rats, 774, 776, 781, 805, 806, 808, 809,
811, 812, 816, 825, 827, 831-835, 837,
848, 863, 907
baby, LD50 for, 931
broad developmental divisions of, 863
carcinogenesis in, 1152, 1157, 1159,

1164, 1172, 1174, 1180, 1184
carcinoma in, 684

irradiation of, during fetal period, 893-
894, 896-898
during period of major organogenesis,
869, 872-874, 876-888, 891-892,
901
during preimplantation period, 864-
866
LD60 for, 930-931
sarcoma in, 693
Reactor, nuclear, 14
Reciprocity, 924
Recombination, 355

of ions, 194n., 239-240
Recovery in radiosensitivity, 928
Red cells, 960-961, 970, 974, 979, 1002
nucleated, 305
(See also Erythrocytes)
Red pulp of spleen, 1106, 1108
Reduplication of organs, 325, 335
Refractive index of plasma, 972
Regeneration, 459, 1135

of blood-forming tissues, 963-964
of bone marrow, 1101

hematopoietic cells, 1100
capacity for, and type of radiation

damage to embryo, 902
of epidermis, 1121

following injury from prenatal irradia-
tion, 880, 888
inhibition of, 335
of liver, 1128, 1135
of lymphatic tissue, 1103
of spleen, 1106
Regression of tumors, 325, 339
Regulatory powers of mammalian blasto-

meres, 868, 904
Relative biological effectiveness, 321,

322, 328-342
Relative radiosensitivity, 1092, 1132

nucleus and cytoplasm, 1094
Relaxation, electric, 69
Relaxation length, 104
Relaxation time, 76
Rem (roentgen equivalent mammal),

1147
Remodeling of bone, 1112
Renal failure, 925

Rep (roentgen equivalent physical), 1147
Repair of bone, 1111

1258

RADIATION BIOLOGY

Repeated X irradiation of lymph nodes,

1104
Reproduction, convariant, 352

in mammals, effect of radiation on,
826-831
Reproductive system, effects of prenatal
irradiation on, 889-890, 896-898,
909, 912
Resonance, 16, 46
Resonance effect, neutron collisions, 114

nuclear, 58
Resonance radiation, 199, 202, 206, 216
Resonance rule in collisions, 205, 207,

208, 220, 235, 237-238
Resonating current, 17, 20
Resorption, of bone, 1110

of embryos after irradiation, 864, 866,
873
(See also Mortality, prenatal)
Respiration, endogenous, 302
of intestinal tissue, 984
single cells, 300
Restitution, average time for, 728

of chromosome breaks, 363, 483, 499,

500, 722
oxygen effect on, 749
proportion of breaks undergoing, 731
space factor in, 730
Retardation, of breathing, 940

of growth, 339, 340, 869, 876-877, 879,
896, 900, 905, 912, 922
Reticular cells, 930, 1135
bone marrow, 1099
lymphatic organs, 1101, 1103
radioresistance of, 1133, 1135

bone marrow, 1100
spleen, 1106
thymus, 1104
Reticuloendothelial system, 926, 1004-

1005
Reticulocytes, effect on, of fast neutrons,
1043, 1048
of gamma radiation, 1055
of spleen shielding, 1078
and splenectomy, 1078
of X radiation in phenylhydrazine-
induced anemia, 1036
Retina, 306, 339

damage to, following prenatal irradia-
tion, 880, 889, 891, 897-898, 909,
912
degeneration in, 338
Reunion of chromosome breaks, 722
oxygen effect on, 749
space factor in, 730
Reversible degenerative phenomena, 1092
Rhodanase, 987

Rili abnormalities following prenatal

irradiation, 882, 884, 887, 892, 904
Ribonuclease, 299

inactivation of, 944
Ribonucleic acids, 289, 305, 929

synthesis of, 962
Rodents, 686, 687
Roentgen rays, 3
Roentgen units, 120, 319

definition of, 149
Root-mean-square deviation, 80
Roots, pea, 341

sunflower, 340

tomato, 341

Vicia faba, 322, 325, 340, 341

wheat, 339
Rotifers, 583
Ruminants, 444

Rutherford scattering, 41, 43, 44, 51
Rutin, 968, 969, 979

S

Salamander, 335, 554, 776, 811, 816
Salivary-gland chromosomes in animal
cells, 635-637, 646, 648-652, 660,
661, 663-665, 668-670, 673, 677-679,
689
Salivary-gland secretion, 1127
Salivation, excessive, 995
Salmon, 448, 628, 629
Saltants, 593
Sarcoma, Jensen, 305

rat, 693
Saturation effect, 140
Scatterer recoil, 43
Scattering, a-particle, 42

coherent, 104

Compton, 37, 38, 44

angular distribution of, 39
distribution of energy, 38

of electromagnetic radiation, 37, 45-48

probability of, 39

Rutherford, 41, 43, 44, 51
Schwann cells, 996
Sciara, 484, 689, 690
Scilla, 776, 799, 804
Scintillation counters, 233
Scintillations, 4
Sclerosis in skin, 1122
Sea urchins (Arbacia), 554, 766, 769, 771,
772, 784, 785, 789, 796, 803, 812, 814

sperm, 300, 301, 304
Sea water, 576
Sebaceous glands, 1119, 1123

carcinogenesis in, 1176
Secede, 363
Secondary radiation, 72

SUBJECT INDEX

1259

Secondary sex characters, 1118
Sedimentation rate of blood, 964

effect of radiogold on, 1061
Sedimentation velocity, 295
Seedlings, pea, 341

sunflower, 340

Vicia, 340, 341

wheat, 339, 935
Seeds, 939

Segmental interchange, 367, 368, 847
Segregation, Mendel's law of, 355
Semilethal effects, 1005
Semipermeable membrane, 301
Semisterility, 825, 840, 842-850, 854
Sensitive area, 140, 283, 284
Sensitive volume, 130, 140
Sensitivity of cells, differences in, to

ionizing radiations, 681-691
Separation of cartilage from bone, 1113
Sequoias, 583

Sertoli cells, 828, 831, 1114, 1116, 1117
Serum, horse, 935
Serum phosphatase, 299
Sex and radiosensitivity, 933
Sex determination, 455
Sex ratio, 839, 840

following prenatal irradiation, 877-878,
912
Shielding, 926

of parts of pregnant females during
irradiation, 864, 901, 911

spleen (see Spleen shielding)
Showers, cascade, 63, 70, 95, 111
Shwartzman phenomenon, 1003
Silkworm, 457, 510
Single-hit, 132

Single-hit curves (see Dose effect, expo-
nential curves)
Situs inversus following prenatal irradia-
tion, 886, 888, 892
Skeleton, skeletal abnormalities, effects
of prenatal irradiation on, 882-887,
909

(See also specific abnormalities)
Skin, 325, 337-338, 1119

cysteine protection, 943

damage following prenatal irradiation,
881, 886, 896-898, 912

derma, 1119, 1120

epidermis, cellular polymorphism, 1121
degeneration of, 1121

epilation of, 1120

epithelium, 1120
atypical, 1120
cessation of mitosis, 1120

erythema of, 926, 976-977, 1120

inflammatory reaction, 1121, 1122

latent period in, 1121

Skin, loss of elastic fibers in, 1122
mammary gland, 1123
necrosis, 1120

nuclear polymorphism of, 1120
papilloma, 1175
pigmentation of, 1120
radiation effects, 997
plutonium, 1122
radium, 1122
yttrium91, 1122
radiosensitivity of, 929
sclerosis of, 1120
sebaceous glands, 1123
sweat glands, 1123
temperature of, 977
tumors, 1151, 1153, 1155, 1159, 1175-

1177
ulceration, 1120, 1149
Skin glands and X rays, 1123
Skull abnormalities following prenatal
irradiation, 879, 881, 883, 887, 889,
909
Slow neutrons (see Neutrons)
Small intestines, 306, 307, 1124
Smear techniques in radiobiology, 632,

635
Sodium, excretion of, 989
of plasma, 974
retention of, 971
of tissue, 972
Sodium fluoride, 941
Sodium hydrosulfite, 580
Sodium nitrite, 941, 945
Sodium space of intestine, 989
Sodium24, 1093
effect of, 1058

on intestine, 1127
on spleen, 1 107
on thymus, 1105
toxicity of, 923
Somnolence, 995

Sonic vibration, effect on radiation-
induced chromosome aberrations,
737
and mutation, 572
Soret band, 287

Spatial distribution of ions, 194, 197
Species, radiosensitivity of, 930-934
Species heterozygotes, 456
Specific ionization, 87, 315, 920, 923,
1093
and carcinogenesis, 1146, 1147
as related, to oxygen effect on chromo-
some-aberration production, 752
to radiation efficiency in chromo-
some-aberration production, 732
Spectral analysis, 18

1260

RADIATION BIOLOGY

Spectrum, 18
absorption, 47
albumin, 294
bands, 47
continuous, 47
line, 47

mutational, 590, 594, 596
nuclei, 49
optical, 20
Sperm, fertilization power of, 301
formation of, 998-999
motility of, 455
restitution of breaks, 483
Spermatids, 828-831, 842, 1114
Spermatocytes, 828-830, 842, 844, 1114,
1116, 1133
radiosensitivity of, 929
Spermatogenesis, 829, 830, 842, 843, 854,
1114, 1116, 1136
cessation of, 1116
Spermatogonia, 829, 830, 837-839, 842,
843, 846, 847, 852, 855, 1114, 1116,
1133
radiosensitivity of, 929, 999
Spermatozoa, 826, 828, 830-832, 834,
835, 837, 839, 842-844, 846-848, 855,
856
Spermia, 1114

Sphaerocarpus, 476, 531, 540, 553
Spiderwort, 484
Spin-conservation rule in collisions, 205,

208, 220
Spindle, mitotic, destruction of, ultra-
violet-induced, 814, 815
suppression of, X-ray-induced, 804
Spleen, 306, 919, 926, 960-966, 979, 982,
1106
ectopic erythropoiesis, 963
ectopic myelopoiesis, 1106
effect on, of clamping off circulation
in shielding-irradiation procedure,

1077, 1078

of cutting distal vessel in shielding-
irradiation procedure, 1069, 1070,
1076
of transplants, 963, 965
effect of increments to, on survival,

1078, 1079
erythropoiesis, 1107, 1108
germinal centers, 1106
hemocytoblasts, 1107
heterophil leukocytes, 1106
homogenates, 926, 963, 965
lymphatic tissue, 1106
lymphocytes, 1106
lymphopoiesis, 1106

mast cells in, 1107
megakaryocytes, 1106, 1108

Spleen, myelocytes, 1106
radiation effect on, 919
Ba140-La140, 1107
fast neutrons, 1 107
gamma rays, 1107
plutonium, 1107
radioactive isotopes, 1107
radium, 1107
slow neutrons, 1107
sodium24, 1107
strontium89, 1107
yttrium91, 1107
Zr93-Cb93 (Zr95-Nb95), 1107
radiation exposure, 982

prenatal, sensitivity to, 880, 897-898
red pulp, 1106, 1108
regeneration of, 1106
retention of colloids, 979
reticular cells, 1106
sarcoma of, 1155, 1156
white pulp, 1106

(See also Spleen shielding; Spleen
transplants)
Spleen shielding, 926, 963, 965, 1068-1073
comparison of, with head shielding,
1073
with intestine shielding, 1072
with limb shielding, 1073
with liver shielding, 1072
effect of, on peripheral blood, 1069-
1070
on survival of mice, 1068
and cysteine, 1079

plus estrogen, 1079, 1080
and estrogen, 1079
hematopoietic recovery, 1069-1076
splenectomy effects after, 1077-1081
on antibody formation, 1080, 1081
before radiation and with spleen

transplants, 1073-1074
on survival of mice, 1073, 1074, 1077,
1081
Spleen transplants, 926

modification of radiation injury by,

1074-1075
quantity of transplanted tissue on
survival, 1076
Splenic suspensions, 1074
Staphylococcus (Micrococcus aureus), 537,

545, 548-552, 562
Stationary states, 29-30

within a molecule or crystal, 32
oscillatory or rotary, 32
Stem cells in testis, 1117
Sterility, 826-831, 840-843, 854, 855
following prenatal irradiation, 897-898
(See also Reproductive system, effects
of prenatal irradiation on)

SUBJECT INDEX

1261

Sterility, and ovary, 1117

partial, 825, 840, 842-850, 854
production of, 999
treatment of, 999
Sterilization, 337
Slichococcus, 322
Sticky chromosomes (see Chromosome

stickiness)
Stillbirths, 835, 836, 839

following prenatal irradiation, 874, 898,

900, 911
{See also Mortality, neonatal)
Stimulation of blood-forming tissue by

radiation, 1064
Stock, "sifter," 458
Stomach, 1124

radiation effects, 985-986
ulcers, 1125
Stopping number, 52, 53
Stopping power, 52, 87, 158, 165, 166
of different materials, 88
of water, electron, 54
proton, 54
Straggling, 88

Stratified squamous epithelium, 1124
Streptomyces, 543, 544
Streptomycin, 585, 1003
Stresses, nonspecific, 1004
Stromal cells of thymus, 1104
Strontium, radioactive, 586
Strontium89, 1093

effect of, on bone, 1110
on bone marrow, 1059
on cartilage, 1113
on intestine, 1127
on peripheral blood, 1059, 1060
on spleen, 1107
on testis, 1117
on thymus, 1105
and splenectomy, effect on erythro-
cytes, 1059, 1060
toxicity of, 963, 970, 973
Styrene, 291

Submaxillary glands, 306
feminizing changes, 999
Succinate, 306
Succinic dehydrogenase, 987
Succinoxidase, 298
Sugars, 307

(See also individual sugars)
Sulfhydryl compounds, 551, 579

effect on radiation injury, 1067
Sulfhydryl content of plasma, 975
Sulfhydryl groups, 937-938, 941

oxidation, 287, 290, 292
Sulfhydryl enzymes, 937, 941, 987
Sulfide, 945

Sulfur, 285, 576, 586

radioactive, and mutation, 585
Supersonics and mutation, 589
Supralethal irradiation, 1006
Survival, mean time of, 931, 932
Sweat glands, 1119, 1123
Swelling, of cells, 1095

of nuclei, 1096, 1127
Swine, LD50 for, 930
Symbiosis, 444
Synapsis, 355
Synchrotron, 12

Tadpole, 776, 786, 804-806, 810
Tail abnormalities following prenatal
irradiation, 881, 884, 892, 902, 905,
907
Target theory (hit theory, Treffertheorie) ,
129, 130, 132, 140, 245, 248, 252, 258,
272, 283, 284, 301, 921
and mutation, 525-531, 552
Telangiectasias, 1151
Telomeres, 366, 446

Temperature, effect of, on chromosome
aberrations in absence of oxygen,
743
on radiation-induced chromosome
aberrations, 741
and mutation, 412, 565
and oxygen effect, 742
and radiation effects, 1093
and radiosensitivity, 927, 935-937,
945-946
Temperature changes in skin, 977
Tendon, 1097
Termites, 444
Terphenyl, 290
Terramycin, 1003

Testes, 306, 828, 829, 831, 842, 843, 1114
basophilic outstretched cells in, 1114,

1116
carcinogenesis in, 1168-1171, 1190
interstitial cells of, 1116, 1117
radiation effects, 998-999
Ba140-La140, 1117
fast neutrons, 1114, 1116, 1117
gamma rays, 1114, 1116
phosphorus32, 1116, 1117
plutonium, 1117
radium, 1117
repeated irradiation, 1114
strontium89, 1117
yttrium91, 1117
Zr93-Cb93 (Zr95-Nb96), 1117
spermatogenesis, 1136
Tetanus toxin, 941

1262

RADIATION BIOLOGY

Tetraborohydride, 942, 945

Tetrads, 355

Tetraploid cells, 642, 643, 683

Thiocyanate space, 971

Thiol compounds, 307

Thiol groups, 298, 304

Thiols, 286-289, 303, 308

Thiosulfate, 942, 945

Thiouracil, 935, 946, 993

Thiourea as protection, 942, 945

Thorax abnormalities following prenatal

irradiation, 884, 906
Thorotrast, 970

Threshold dose in carcinogenesis, 1167
Thrombi, platelet, 977
Thrombocytes (see Platelets)
Thrombocytopenia, 966
Thrombosis, vascular, 967
Thymic cell suspensions, 935, 939, 942,960
Thymonucleic acid, 289
Thymus, 306, 1104

effect on, of cerium-praseodymium,
1105
of external beta rays, 1105
of fast neutrons, 1105
of plutonium, 1105
of radioactive isotopes, 1105
of slow neutrons, 1105
of sodium24, 1105
of strontium89, 1105
of yttrium91, 1105
Hassall's bodies, 1105
lymphocytes, 1104
mast cells in, 1105
sensitivity of, to prenatal irradiation,

880, 897-898, 912
stromal cells, 1104
Thyroid, 1132

administration of, 929, 935, 937, 946
carcinogenesis in, 1167, 1171-1172
iodine131 and, 1132
radiation effects, 942
Thyroidectomy, 935, 993
Time-intensity factor, 134, 1091, 1130
Tissue breakdown, 1000-1002
Tobacco mosaic virus, 510
Toluidine blue, 967-968

effect on radiation-induced hemor-
rhage, 1046
Tonus of intestine, 985
Tooth primordia, effect of P32 on, 899-900
Total-body vs. local irradiation, 1091
Totipotency, 868, 904
Toxins, bacterial, 1000, 1003

produced by radiation, 245, 877, 890,
900
Trachea, carcinogenesis in, 1180
cartilage, 1000

Tracks of high-energy particles, 77, 82,

83, 315
Tradescantia, 363, 385, 386, 484-488, 490,
491, 493, 516-518, 522, 524, 525, 533,
534, 559, 561, 562, 566-568, 570-573,
578, 584, 774, 799, 804, 807, 814, 940,
941
chromosome number in, 631
microspores and chromosomes, 631,
632, 659, 668, 669, 675, 679
diploid, 683
haploid, 683
radiation-induced aberrations in chro-
mosomes of, 713-761
Transfusions, blood, 966, 967, 1006, 1081,

1082
Transition effect (X rays), 111
Translocations, aneucentric, 367, 386
chromosome, 999

in animal cells (see Chromosomes)
deletion-insertion type, 376
eucentric, 367, 369, 386
in mammals, 842-844, 846-850, 854
mutual or reciprocal, 367, 368
rate with dose, 480
and temperature, 565
whole-arm, 369
X-ray induced, 484, 487
Transport of small lymphocytes, 1104
Trauma (see Radiation injury)
Treatment of radiation syndrome, 1006
Treffertheorie (see Target theory)
Tricarboxylic acid cycle, 946
Trichophyton, 531, 535, 540. 570
Trigeminal ganglia, 995
Trillium, 689, 691, 695, 722, 799, 802
Triols, 291

Triploids, 456, 643, 683
Triticum, 589, 798
Tritium, 587

Triton, lethal effects, 924
Triturus, 807
Trypan blue, effects of, on mammalian

embryo, 907
Trypsin, 299

Tryptophane metabolism, 988
Tumor cells, 935, 939-940
Tumor induction (tumorigenesis), 922
latency of, 1150, 1158, 1171. 1178. 1181
pathogenesis of, 1174
by radioscopy, 1148, 1163
(See also Carcinogenesis; Carcinogenic
agents)
Tumors, 305

carcinoma (see specific animals and

sites)
chondrosarcoma (see Bone)

SUBJECT INDEX

1263

Tumors, chorioepithelioma induced by
X rays, 1169
chromosome abnormalities in. 338
embryoma, 1158
endothelioma, 1157, 1169, 1184
hemangioendothelioma, 1157, 1160,

1169, 1176, 1184
incidence of, in painters of radioactive

watch dials, 1158, 1182-1183
inhibition of "takes," 339
lymphoma (see Leukemia)
mammary, 1153, 1161, 1164, 1172-
1174, 1188
milk agent, 1173
neoplasms (see Carcinogenesis; Car-
cinogenic agents; specific sites)
osteosarcoma, 1111
papilloma, 1175
plasmocytoma, induction by meso-

thorium, 1157
radiosensitivity of, 929
regression of, 325, 339
sarcoma (see specific animals and sites)
Turbidity of plasma, 974-975
Two-hit aberrations, 723
Tyndall effect, 119
Tyrosine, 292, 293, 296

dissociation of hydroxyl group, 293
effect of alpha radiation on, 271

U

Ulcers, intestinal, 983
Ultraviolet radiation, 362, 390, 399
action spectrum for mutation, 540, 541,

545
and chromosome changes, 531-535, 738
chromosome effects, 738, 814
cleavage furrow, change in position,

815. 816
in combination with X rays, 738
and deletions, 532, 533
and dominant lethal mutations, 532
energy absorption in tissues, 764
excitations, 539
and inversions, 533
lethal effect on cells, 816. 817
mitotic effects (see Mitotic effects)
mutagenesis, by irradiation of medium,

548-552
repair and enhancement in, 542-548
and mutation, 504, 592-595

intensity, 538
nucleolar spheration, 816
and point mutations, 531-542
spindle effects, 814
and terminal deletions, 532
and translocations, 532, 533
(See also Light)

Uranium, 295, 574
Uranyl nitrate, 575
Urea of blood, 973, 989
Urease, 299
Urethane, 922, 937, 946

ethyl, 416, 424
Uric acid, of blood, 925, 973, 1000

of liver, 988
Uricemia, 925
Urinary bladder, 1129
Urine composition, 989
Urobilinogen, 988

Urogenital-system abnormalities follow-
ing prenatal irradiation, 886-888,
892, 896-897, 907, 909
Uterus, 1117, 1118

carcinogenesis in, 1151, 1153, 1174-
1175

V

Vacuolization of cartilage cells, 1113
Vagina, 1117, 1118
Vagotomy, 980-981, 985
van der Waals force, 32
Variability, biological, 27
Variance, 80

Vascular calcification and radium, 1122
Vascular changes in skin, 1121
Vascular damage, 966
Vascular fragility, 968, 979
Vascular permeability, 978-979
Vascular thrombosis, 967
Vascularity and radiosensitivity, 940
Velocity, sedimentation, 295
Velocity gradient, 289
Vertebral-column abnormalities follow-
ing prenatal irradiation, 884, 887,
902, 906-907, 909
Viability and mutations, 395, 853-856
Viability effects, cell, 804-810, 816-817
chromosome losses and, 807, 808
degenerative changes described, 804-

807
DNA formation and, 808
dosage rate and, 810
kind of radiation and, 809, 810, 816,
817
a rays, 810
j3 rays, 809
7 rays, 809, 810
neutrons, 809
ultraviolet radiation, 816
X rays, 809
mitotic stage and, 805, 806
radiosensitivities of different kinds

of cells, 808
temperature and, 810, 816
in vivo and in vitro, 806

1264

RADIATION BIOLOGY

Vicia faba, 363, 391, 558, 577, 589,
736, 744, 748, 755, 774, 776-778, 782,
784, 786, 799, 801, 811, 929, 935, 940,
941
chromosome aberrations, 306
Vinyl acetate, 291
Virus, 527

inactivation of, 324, 329-330
Visceral abnormalities and retarded devel-
opment following prenatal irradia-
tion, 886, 888, 896-898
Viscosity, 289
anomalous, 289
of cytoplasm, 1095
of gelatin solutions, 295
Vitamin- A absorption, 986
Vitamin C, 942, 945, 967, 975, 990
Vitamin deficiency, 934
Vitamin K, 967
Vitamin P, 968

Vitamins, 934, 965, 967-968, 986
Vomiting, 969, 982, 984

W

Water, 256, 273

"activated," 284, 937

and cell organization, 273

effects on, 324, 328

extracellular, 1001

influence on high-energy mutagenesis,

555-564
metabolism, 969
"pure," 285
radicals from (see Radicals from

water)
radiodecomposition of, 750
role of, in radiation effects, 244n., 273,
274
on chromosomes, 751^".
and radiosensitivity, 754
Water content, effect on radiosensitivity,
927
of organs, 1001
of tissues, 971
Wave length, 16
Wave length effect, 137
Weeds, noxious, 444
Weight, at birth (see Body size)
loss of, 984
of organs, 1001
Wheat seedlings, 935
Whole-body irradiation, 1136
of pregnant females, 864, 901
(See also Embryo ; Fetus)
Wines, 445

X radiation, anemia induced by, bio-
logical effectiveness of fast neu-
trons and, 1048
comparative effect of fast neutrons
and, 1044
effect of, on adrenal glands, 1131

in mammalian prenatal develop-
ment, 861-918
effect on peripheral blood, of acute
exposure, 1031-1041
of chronic exposure, 1047-1049
(See also Bone marrow)
effect of tissue shielding on injury in-
duced by acute exposure, 1068-1073
genetic effects, 629

in mammals, 825-859
mitotic inactivity, 628
primary effect, 628, 694
secondary effects, 628, 629
X-ray fluorescence, 47, 69, 107
X rays, 3, 14

absorption of, 49, 107, 108
absorption coefficient, 103, 110
asters, induced supernumerary, 803
build-up, 108

chromonema effects, 801, 802
chromosome breakage, 801
chromosome change in form, 801
chromosome stickiness (see Chromo-
some stickiness)
crystal diffraction, 105
cytosome effect, 803
deamination by, 265-270
definition of, 20
distribution of, 86
effect of intensity of, 727
effects of, on chromosomes, 723Jf.
effects on, of modifying factors, 736jf.
electronic equilibrium of, 111
emission, 44
energy distribution, 109
energy release in tissues, 764, 786
lethal effect on cells, 805-810
measurement of, 318-319
mitotic effects, 768-791, 794-798

(See also Mitosis; Mitotic effects)
penetration of (see Penetration)
probability of different effects, 70
production of, 23

relation of aberration yield to dose, 723
relative efficiency, 732
scattered, 107
secondary, 106
spindle suppression, 804
thymonucleic acid depolymerization,
800, 801

SUBJECT INDEX

1265

X rays, transition effect, 111

translocations induced by, 484, 487
(See also Chromosome aberrations;
Mutations; Radiation; X radia-
tion)

Yeasts, 321, 324, 325, 331-332, 445, 595,
940

diploid, 303, 321, 324

haploid, 302, 303, 321, 324

hexokinase, 298

Sac char omyces cerevisiae, 304, 325
Yttrium, radioactive, 586
Yttrium90 colloid, 978

Yttrium91, effect of, on bone, 1110, 1112
on intestine, 1127
on liver, 1128

on peripheral blood, 1058, 1059
on skin, 1122
on spleen, 1107
on testis, 1117
on thymus, 1105

Zr93-Cb93 (Zr95-Nb96), effect of, on intes-
tine, 1127

on spleen, 1107

on testis, 1117
"Zone sensible," 301