Neoplasms in Rats Treated with Pituitary
Growth Hormone
II. Adrenal Glands*
HENRY
D.
MOON,
M.D.,
MIRIAM
AND
E. SIMPSON,
HERBERT
M.
EVANS,
M.D.,
CHOH
HAO Li,
PH.D.,
M.D.
(From Institute of Lvperimental Biclogy and Division of Anatomy, University of California, Berkeley, California, Dirision of
Pathclogy, University of California Medical School, and Veterans Administration
Hospital, San Francisco, California)
The occurrence of many different neoplasms in
fifteen normal plateaued female rats treated for 350
to 485 days with pituitary growth hormone' was
reported in the preceding paper of this series (4).
The histogenesis of pulmonary bymphosarcoma was
reported in some detail in that paper. The present
report is concerned with the hyperplastic and neo
plastic changes in the adrenal glands of the same
The zona fascicubata was composed of bong,
straight cords of cells. The cells in this zone were
larger than those of the zona gbomerubosa, and the
cytoplasm of these cells was vacuobated in paraffin
sections and filled with lipoid in frozen sections.
TABLE I
ADRENAL GLANDS OF CONTROLS
RIGHT
animals.
OR
rfh
adrenal
glands of the rats injected
pituitary
growth hormone and their controls
dissected free of fat, weighed, and fixed. They
with
were
were
RAT
B651@
and stained
with Sudan
Orange
and
hematoxybin.
RESULTS
RATS
thickness.
The cortical
grants
from
the
tissue
U.S.
Public
Health
pituitary
growth
hormone
used
in
this
by physico-chemical
criteria (f). The initial
mg. was gradually increased to a maximum
by the end of the experiment.
Received
for publication,
February
study
GH6@6@
BH6@94
pure
Cortex
(MG.)
Medulla
48
nodular
37
38
47
30
30
30
30
30
small nests of medul
lary cells in cortex
30
38
nodular
nodular
54
R
L
48
L
40
40
36
36
34
36
88
R
L
R
L
44
G6@68
B649@
daily dose of 0.4
of 3.0 mg daily
R
L
G6460
BH6@9@
Service
was
GH6357
B6@91
was differenti
RG-409; the Research Board of the University of California;
the American Cancer Society, Inc., N.Y.; and the University of
California Cancer Grant.
1 The
R
WITH ALBUMIN
ated into the usual three zones. The zona glomeru
bosa was narrow; the cells were small, with scanty
cytoplasm, and were arranged into small groups.
by
W65@3
GH6554
INJE('TED
The adrenal glands of the control rats were
fairly uniform in appearance, and the weight of a
single gland varied from @3
to 48 mg. (Table 1 and
Figs. 1—3).These glands presented the usual lentic
ular shape and were pale yellow. Microscopically,
the glands were characterized
by cortices of uni
* Aided
B6508
R
L
R
L
R
L
R
L
GH644@
CONTROL
form
B6400
ABNORMALITIES NOTED
@\EIGHT
53
B6530
usually embedded in paraffin, sectioned serially,
and stained with hematoxylin and eosin, except in
one of the control rats and in three of the experi
mental rats—in which instance frozen sections
were prepared
LEFT
R
L
R
nodular
nodular
The reticular zone consisted of cells arranged in
irregular cords; these cells were slightly smaller
than those of the zona fasciculata. The cytoplasm
of these cells contained small amounts of brown
pigment.
The
nuclei
of the
cells
of all three
zones
were small, round, and deeply basophibic. In three
15, 1950.
364
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1950 American Association for Cancer Research.
1
.t@,
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.,@‘
S
Fios. 1—3.—Control
Rats; adrenal glands.
Fio. 3.—RatB 6400.Paraffin section; hematoxylin and
eosin. The cortical cells have finely granular, dense cyto
Fio.1.—RatB 6530.Topographicview;frozensection; plasm and small, round, deeply-staining
nuclei. The medul
hematoxylin
and Sudan Orange X 16. Note the normal
lary cells have pale or clear cytoplasm and usually vesicular,
perivascular extension of medulla to hilus.
round nuclei. A few spindle cells with hyperchromatic
nu
clei are seen. X 355.
Fia. @.—Higher
magnification
of Fig.1, X 6@.
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1950 American Association for Cancer Research.
366
Cancer
rats,
was
the
zona
nodular.
fasciculata
The
of
nodular
cells which had more
those of the adjacent
@
cells
into
adrenal
were
glands
composed
abundant
cytoplasm
cortex. These areas
“barrelstave― appearance
of the
both
areas
slightly
of
than
had a
clue to the arrangement
curved
Research
present. Frequently,
the cells had indistinct bor
ders and were in masses suggesting a syncytium.
In some of the areas of atypical medulbary hy
perplasia, the cells were larger than those of the
adjacent medulla. These cells had more abundant
cords.
TABLE
‘fhmedullae of the glands of nearly all animals
were
uniform
in shape
and
in amount.
The
ADRENAL GLANDS OF EXPERIMENTAL
medul
lary tissue was composed of polyhedral, flat or
crescentic cells usually arranged in small compact
groups.
The
predominant
cell
was
the
large
poly
hedral cell with a round vesicular nucleus and pale
or clear cytoplasm. The flat crescentic cells had
hyperchromatic
nuclei and scanty granular cyto
plasm.
In one rat (Gil
6554), several
EXPERIMENTAL
RATS
INJECTED
GROWTH
These
areas
were
to those
observed
in
the adrenal cortices of two of the control rats. In
one rat there was a cortical adenoma in the zona
reticularis (Figs. 6 and 9). In many of the rats
there was displacement and infiltration of the cor
tex by the medublary proliferation described below.
The medullae of all adrenabs of the experimental
rats were enlarged ; this was considered the basis
for the increased weight of the adrenals in many of
these rats. The medulbary cells had more abun
dant, pale cytoplasm and larger, vesicular nuclei.
Nucleoli were often prominent. Rare mitoses were
present. Most of the cells were polyhedral, but
some were spindle-shaped.
Except for their larger
size,
the
medublary
cells
were
essentially
similar
to
those of the control rats.
In addition to the medublary hypertrophy
and
hyperplasia, there were focal or nodular areas of
varying size composed of atypical cells in the
medublae of nine rats (Figs. 4 and 5). These areas
were present in one adrenal in each of six rats and
in both
adrenabs
in three
B6458
GH6368
WEIGHT
(MG.)
R
L
R
L
R
L
BH6@74*
BH6334
HORMONE
similar
G6534
OR
LEFT
ABNORMALITIES
1@1
49
R
59
L
88
R
113
rats. The cells in most of
these areas were small, with scanty cytoplasm and
hyperchromatic
nuclei. Occasional mitoses were
L
Medulla
hyperplasia
hyperplasia
hyperplasia
100
@o
NOTED
Cortex
displaced
displaced
displaced
WITH PITUITARY
The weights of the adrenals of the rats injected
with growth hormone ranged from @7to
@0mg.
(Table
@).Small nodules of translucent pink and
hemorrhagic
medullary tissue presented on the
surface of one of the adrenals in each of three rats
(GH 6368, BH 6334, and BH 6@76). In rats
BH 6334 and BH 6@76 these nodules were not re
bated to the vascular pole of the adrenal gland.
There was no consistent increase in the width of
the adrenal cortices of rats injected with growth
hormone; in some animals, the cortices were nar
rower than those of the controls. Nodular areas
were present in the zona fasciculata of six of the
rats.
RAT
small nests of
medulbary cells extended into the zona reticularis.
No neoplasms of the adrenal glands were present
in the control rats.
RATS
RIGHT
nodular and
displaced
focal atypical
hyper
plasia
hyaline degeneration of
large medullary tu
mor
bilateral
metastatic
lymp@iosercoma and
atypical hyperplasia
large medullary tumor
focal
7@
atypical
hyper
plasia
G6506
hyperplasia
R
L
95
9@
BH6@76
R
L
40
40
G6553
R
L
R
L
@7
31
61
71
R
66
L
73
nodular
R
L
R
L
40
38
40
3@
nodular
nodular
focal atypical hyper
plasia
adenoma
hyperplasia
medullarytumor grow
ing through
BH6313
W63@4
G6@55
G6361
capsule
hyperplasia
nodular
hyperplasia
hyperplasia
focal atypical
plasia
focal atypical
plasia
focal atypical
plasia
hyperplasia
hyperplasia
hyper
hyper
hyper
hyperplasia
focal
atypical
hypcr
plasia
B6@66
R
36
G6@69
L
R
L
50
44
50
R
40
L
48
G6@7l
focal atypical hyper
nodular
plasia
hyperplasia
hyperplasia
focal atypical
plasia
hyper
focal atypical
plasia
hyperplasia
hyper
a Rat BH6174, as previously
reported, bad lymphosarcoma
involving
both lungs and hilar lymph nodes, with invasion of the right
pulmonary vein and left atrium.
cytoplasm. The nuclei were larger; the nuclear
membranes were thicker, but the vesicular appear
ance of the nuclei was similar to that of the ad
jacent medulla. There were occasional mitoses.
There were transitional stages between all types
of cells described above.
In many of these rats, the zona reticularis was
displaced and invaded by nodules of atypical
medullary cells. Groups of medullary cells occa
sionally infiltrated as far as the zona fasciculata.
In two rats (BH 6334 and BH 6@76) the atypical
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1950 American Association for Cancer Research.
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Fios. 4—5.—Experimental Rats; adrenal
glands.
FIG. 4.—Rat G 6@71. There is displacement and invasion of the zona reticularis of
the cortex by small “dark―
atypical medullary cells in the center. These cells have indistinct borders, hyperchromatic
nuclei, and
syncytialmasses.There isanothernoduleof
,
atypical medullary cells at the extreme left.
There is some compression and distortion of
the zona reticularis between these two nod
ules. There are a few typical hypertrophic
medullary cells at the lower left. X 355.
FIG. 5.—Rat B 6466. There are nodular
masses of large atypical medullary cells in
vadingand compressingcortex.X 1@5.
367
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1950 American Association for Cancer Research.
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FIGS. 6—9.—Experimental Rat BH 6476; adrenal gland;
frozen section; hematoxylin and Sudan Orange.
FIG. 6.—Topographic view; note the cortical adenoma
and the irregular outlines of the medulla. X 16.
FIG. 7.—Invasion of cortex by neoplastic medullary
cells. The proliferating
medullary cells at the upper left
have grown through the cortex and have formed a nodule on
the surface of the adrenal gland. X 64.
Fio. 8.—Higher magnification
showing disruption
of
cortexand capsuleby neoplastic
medullarycells.
X 1@5.
FIG. 9.—Higher magnification
of cortical adenoma.
xi@5.
368
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1950 American Association for Cancer Research.
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FIGS. I0—1@.—Experimental Rat BH 6334; adrenal
gland;frozensection;
hematoxylinand Sudan Orange.
Fio. 10.—Topographic view: the adrenal is greatly en
larged (compare with Fig. 1). The distribution
of cortical
lipoid is altered due to the nodularity of the cortex. Neo
plastic medullary cells have broken through the cortex and
attenuated
the capsule at the right. There is a large nodular
area of abnormal medullary cells to the left. X 16.
FIG.
11.—Higher
magnification
of the
nodular
area
of
neoplastic medullary tissue which is displacing the cortex.
The pale area in the center of the picture is due to degenera
tion and old hemorrhage. X 64.
Fio. I@.—Higher magnification of neoplastic medullary
cells. The nuclei vary in size and are frequently
hyper
chromatic; the cytoplasm is finely granular. The cells are
arranged
in compact
groups.
X 355.
369
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Cancer Research
370
medullary hyperplasia with cortical displacement
and infiltration was of such proportions that the
proliferating
medublary tissue could be observed
on the surface of the adrenal gland as a macroscop
ically visible tumor (Figs. 6—1@2).
In rat BH 6@276,
the medullary cells had broken through the cap
lary tumor in a previous group of normal plateaued
female rats treated similarly was also reported (1).
In the present group of rats, as well as in those
previously reported, the thyroid glands of the rats
injected with growth hormone were morphobogi
cabby indistinguishable
from that observed in the
sule (Figs. 7 and 8). The medulbary
controls.
cells were barge
and were arrange(l in small, compact groups. The
cells were polyhedral or spindle-shaped.
The nu
clei of the polyhedral cells were moderately vesicu
lar and varied considerably in size; those of the
spin(lle-shaped cells were hyperchromatic.
The nu
clear membranes
were thick. There were occa
sionab mitoses. In one rat (G 6368), most of the
medullary tissues had undergone hyabine degener
ation. In one area, the outlines of the pre-existing
medullary cells were faintly discernible. A few
macrophages filled with hemosiderin were present
in the center and about the periphery of the hya
line mass. There was a slender rim of large, atypi
cal medullary cells along one edge of the hyaline
nodule. The lesion in this rat was interpreted as a
medullary
tumor which had undergone hemor
rhage, infarction, and hyaline degeneration.
Metastatic adrenal medullary tumors were not
observed in any of the rats.
There was no significant difference between the
ratio of heart weight to body weight in the rats in
jected with growth hormone, as compared with the
control rats injected with albumin. Nor was there
any evidence of arteriobosclerosis
in any of the
animals.
DISCUSSION
The adrenal cortices were nodular in six of fif
teen experimental
rats ; one of the other experi
mental rats had an adrenal cortical adenoma. Only
two of the control rats had nodular adrenal cor
tices. The significance of this observation is not
clear, particularly
in view of the fact that the
adrenal cortices of the experimental
rats showed
no other evidence of stimulation.
Hypertrophy
of the adrenal medullae of rats
treated
repeatedly
with
thiouracib
has
been
re
ported by Marine afl(l Baumann (3). These au
thors suggest that a direct interrelationship
be
tween the thyroid gland and the adrenal medulla
was responsible for the medullary hypertrophy
that they observed. However, their studies did not
exclude
the possibility
of such hypertrophy
mediated through the pituitary
It was reported previously
medullae
of rats
repeate(lly
being
gland.
that the adrenal
injected
with
growth
hormone were larger than those of control rats.
This was true not only for normal plateaued fe
male rats (1) receiving growth hormone but also
true for hypophysectomized
female rats (5) receiv
ing growth hormone. The occurrence of a medul
The remarkable frequency with which probifera
tive (hyperplastic
and neoplastic) lesions of the
adrenal medulla occurred in the present group of
rats injected repeatedly with growth hormone, and
the absence of similar changes in the control rats,
indicates that these changes are directly related
to the prolonged administration
of preparations of
growth hormone. The displacement and invasion
of the cortex by groups of atypical medullary cells
was very conspicuous.
There were almost all
stages
of development
between
the focal areas
of
atypical medubbary cells and the medullary tumors
which had grown through both cortex and capsule
of the adrenals. For this reason, the focal areas of
atypical medubbary cells are in all probability early
neoplastic lesions.
SUMMARY AND CONCLUSIONS
1. The adrenal cortices of rats injected chron
ically with pituitary growth hormone showed an
increase in nodularity as compared with the con
trobs. There was no other evidence of stimulation
of the adrenal cortices of the experimental
rats.
@.
The adrenal medubbarytissue was increased in
amount in all rats injected chronically with growth
hormone. The increase was due to cellular hyper
trophy and proliferation.
3. In ten of fifteen rats injected chronically with
pituitary growth hormone, there were areas of neo
plastic cells displacing and invading the adrenal
cortex. In three of the rats, the neoplastic medub
lary tissue had grown through the cortex and was
macroscopically
visible on the surface of the
adrenal gland.
REFERENCES
1. EVANS, H. M.; SIMPSON, M. E.; and LI, C. H. The Gigan
tism Produced in Normal Rats by Injection of the Pituitary
Growth Hormone. I. Body Growth and Organ Changes.
Growth, 12:15—3@,
1948.
@.
Li,C.H.;EVANS,
H.M.;andSIMPS9N,
M.E.Isolationand
Properties
of the Anterior
j. Biol. Chem.,
Hypophyseal
Growth
Hormone.
159:353—66, 1945.
3. MARINE, D., and BAUMANN, E. J. Hypertrophy
of Adrenal
Medulla of White Rats in Chronic Thiouracil Poisoning.
Am. J. Physiol., 144:69—73, 1945.
4. MooN, H. D.; SIMPSON,M. E.; Li, C. H.; and EVANS, H. M.
Neoplasms in Rats Treated with Pituitary
Growth Hor
mone. I. Pulmonary and Lymphatic Tissues. Cancer Re
search, 10:@97—308, 1950.
5. SIMpsoN, M. E. ; EVANS, H. M.; and LI, C. H. The Growth
of Hypophysectomized
Female Rats following Chronic
Treatment
with Pure Pituitary Growth Hormone. I. Gen
eral Growth and Organ Changes. Growth, 13:151—70, 1949.
Downloaded from cancerres.aacrjournals.org on June 17, 2017. © 1950 American Association for Cancer Research.
Neoplasms in Rats Treated with Pituitary Growth Hormone: II.
Adrenal Glands
Henry D. Moon, Miriam E. Simpson, Choh Hao Li, et al.
Cancer Res 1950;10:364-370.
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