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Studies on histological changes in experimental scurvy.

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STUDIES ON HISTOLOGICAL C'HAXGES IS
EXPERIMEN TA 1, SC'URT'Y
ACI-IINTYA KUMAR MUKHERJEE AND SACHCHIDAXASDA EANERJEE
Departsient of Plzysiology, Prt.sidency Collc,q,, Calcutta, India
TWENTY-OXTC FIGURES
IKTRODUCTIOK
Different reports about the pathological changes iii scorhutic
guinea pigs hare appeared. These liare been mainly reviewed
by Hess ( '20) and Bourne ( '53). The cliaiiges iiiclude lineinorrhage into the joints, scparatioii of epipliysis, brittleness of
bones, disordered pattern of cartilage cells in the ribs, failure
of normal *dentineand enamel formation of thc teeth, differentiation of osteoblasts into fibroblasts, widespread deficient:- of
collagen in the coiincctivc tissues throughout the body and clelaped healing of wounds. S o histological studies in tissues
other than bones aiicl joints of a scorbntic animal liare appeared. As vitamin C is neccssaiy for the normrtl fmic*tionof
endocrine glands (Kaiierjee and Ghosh, '46 m d '47 ; Bnncrjee
and Deb, '51 aiid W a ) , which have profomid influence 011 other
organs of the body, histologid structures of different tissue5
of scorbutic guinea pigs and monkeys were stucliccl. Thcsc are
presented in the present communication.
MATERIALS AND METIIOUS
Male guinea pigs of weights vai*;c-iiigbetween 23@300 gni
were taken and were put on a scorhntic diet (Ranerjee, '45).
Each of the animals was also fed 5 iiig of ascorbic acid clail;.\and two drops of a concenti-ate of vitamins A and D twice a
week. Animals rvhich grew ~ w l were
l
selected for tlzc esperiment. Animals w x e divided into tliree groups each group coiltaining 8 animals. The animals of group X continued to rcceire
the scorbutic diet with daily oral supplement of 5 nig of ascor90;
bic mid. E!ach of tlre aiiim& of group B mere fed 0.23 rug of
ascorbic acid daily in addition to the scorbutic diet. This was
done to prolong the survival periods of tlie chronically scorbutic animals. The aiiinials of group C wen3 fed the scorbutic
diet without tiny supplcment of ascorbic acid. The last group
of animals ,cleveloped signs of acute scurvy at the end of three
weeks tliep m x e placed 011 scorbutic diet when they were killed
by doctipitatioil. The aiiirrials of group B dereloped signs of
scurvy at the end of 11 weeks a n d t1ic.y were killed at the end of
13 weeks when their condition became very bad. The animals
of gi*oup which grew normall:- were also killed at the end of
13 weeks along with tlic aiiim;ils of group B. .Just after decapitation tlie diflei~liittissiics of thcl animals of each ~ ~ O U
were pulled and fixed over+iiiglit iii %enker-formol solution.
After dehydration and c>learingthe tissues were embedded in
paraffin and 6 p thick s ions were cut and stained by Heidenh i i i 's iron-haematoxvli~i01' 1iaeiti:itoxplin and eosin. E'lioto~riicrogi*aplis
of difTci'(~iitseetioils w e 1 ~t1a k c ~
Thc distribution of ketostclroids and cholesterol in the
adrenals of normal and chroiiic scorbutic guinea pigs were
also studicd by the rnethotls of X 1l)cr.t and Tkbloiid ( '46) a n d
Schultz ( '24).
M o n k q s . Eight healthy female rhesus monkeys of weights
varying between 2-25 kg were fed the same scorbutic diet.
Four of thc animals received in additioii a daily oral supplement of 50 rng of ascorbic acid. All the animals were fed a
c.oncenti.ate of vitamins ~ n 1)
d twice a week. hiiimals which
did not receive a n y ascorbic acid dereloped signs of scurvy a t
the cwd of 4 months when all the 8 monkeys, normal and scorbutic, were killed hy ilwapittitioii. Histological studies of the
diffciwit t issucls T Y C ~ C c i t i ~ i c dout a s in guinw pigs. Tho
changes ohsei*ved a r c as follows.
I3 ' E 8 T T 1 ,
'IT
( ' l i a n g e ~obscrved in the scorbutic guinea pigs which did not
iwc4vc any ascorbic acid supplement a r e described under acute
sc.nr~--y
and changes observed in animals which were fed 0.25
I J
HISTOLOGICAL CHAXGES IN SCV'BVY
909
nig of ascorbic acid f o r 13 weeks a r c described under chronic
scurvy.
Adrennal glaizd. The glands were enlarged. Cellular hypertrophj- of zona fasciculata with nuclear degeneration and
haemorrhage in the zona reticularis were noticed in both acute
a r i d chronic scurvy. Fatty dqqmeration of zoiia fascicnlata was
noticed in chronic scorbutic guinea pigs. Ketosteroids whicli
were present in the cortex of adrenal gland of normal guinea
pigs were absent in scorbutic guinea pigs. The adrenal medulla
of both normal and scorbutic guinea pigs did not sliow any
ketosteroids. ('holesterol is niainlp present in the cells of zona
glomerulosa and zona fasciculata of adwnals of normal guinea
pigs. I n chronic scorbutic guinea pigs it was found only in the
zona glomerulosa and in the few upper most cells of the zona
fasciculata. In scorbutic monkeys an increase in the number
and size of cells comprising zona fas&ulata with early degenerative changes were noticed (figs. 1-9).
Testes. Complete degeneration of Leydig's cells with increased fibrosis and coagulatire nccrosis of the seminiferous
tubnles were observed in chronic scurvy. In acute scurvy the
Leydig's cells only showed early degenerative changes but
seminiferous tubules were normal (figs. 10-12).
Other endocrinc g l a d s . No changes were observed in the
,
and pitnitarj- of scorbutic animals.
thyroid, o v a r ; ~pancreas
Liver. In clironic scorbutic guinea pigs hepatic cells surronnding tlie central veins showed early f at ty degeneration,
peripheral lobular cells showed liyalinization. I n chronic
scurvy the peripheral cells were greatly degenerated. Haemorrhage was obserred in the lioer of scorhutic monkeys (figs.
13-li).
li'id+wy. In clironic scorbutic guinea pigs atrophy of the
glomeruli and hyalinc clegeneration of the proximal convoluted
tubules were observed. S o changes nrere observed in acute
scurvy (figs. 18-19).
In scorbutic monkeys the giomeruli were normal but cloudy
degeneration with nuclear pyknosis were observed in the proxi-
910
A. I<. 3lIiKHEEJEE A N D
W. BANEKJEE
ma1 convoluted tubules. The tnbulai. cpitheliuiii of other parts
was normal (figs. 20-31).
Spleen,. I n scwi%utic moiikeys the contour of nialpigliiaii
cwrpuscles vei*clost i n majority of cases. The artery to the
iiialpighiaii coiyuscles sliowcd cxrlp dcgeiici*atioii with cridothelial proliferation. S o changes wei'e observed in scoi*biitic
guinea pigs.
Other tisszrcs. N o sigiiificant changcs were observed in the
heart, aorta, utei'us, etc.
linemorrhage and degeiierative calianges a s observed in
ndrenals of scorlnutic guinea pigs and monkc:Ts indicate a liypofunction of the adrenals in scurvy. Diminution of cholesterol
and ketostcroi'ds i l l the adrenal cortcs of scorbutic guinea pigs
n i a indicate
~
either reduced transport o r diminished formation
of tliesc substances in the adrenal cortex m-hich resulted in
tiiniiiiished production of adrenal corticnl Iiormones a s cliolestclol is said to bc the raw material for the syiitl~csisof such
liorniori es,
Tlic histoc~hcmicalstudies confirm thc observation of Ranerjee and Deb who reported diminished cholesterol v a l n ~ sof
adrenals ( '51) and .diminished excretion of 17-ketosteroids h2scorbutic guinea pips ( '5%). The degenerative changes of
Lcylig's cells in the tcstes of scorhutic guinea pigs iiidicate R
lippofuiiction of the testes. Degeneratire cliaiiges in seminifcr011s tubnles w w c 0111~o1)scrrcd iii (aliroiiic scorbutic gniiica pigs
aiitl iiot in w u t e scorbutic giuitca pigs.
The degeiici*ative c4iaiiges observed in kidney in chroiiie
scurr~illdieate that ritaiiiiii (' is necessary f o r normal functioning of the organ. The rascular changes observed in the
liver am1 tlie spleen in scorbutic aninials map be a reflection of
the gciic ralized vascular clia 11ges a ssociat ed with scury-.
Tlic role of vitnmiii C' for tlir 1101~iiiiilfuiictioniiig of the tlift'oi*e~it
organs of the l~otlyis apparent froin tlic :~l)ovestnclics
h i t liom tlie vitaiiiiii a d s Iras yet to be esploi~etl.
HISTOLOGICAL CHANGES I N SCURTY
911
SUMMART
1. Haemorrliage and degenerative changes in the zona f asciculata of the adrenal cortex have been observed in scorbutic
guinea pigs and monkeys. Distribution of cholesterol and ketosteroids have been found to be greatly diminished in the
adrenals of scorbutic guinea pigs. The changes suggest hypofunction of the adrenals in scurvy.
2. Lcydig’s cells have been found degenerated in the testes
of chronic scorbutic guinea pigs which possibly led to diminished internal secretion of the testes in vitamin C deficiency.
3. Degenerative changes in the glomerulus and proximal
corn-oluted tubules have been observed in chronic scorbutic
guinea pigs and monkeys.
4. Degenerative and haemorrhagic changes have been noticed in liver and spleen of scorbutic animals.
ACKNOWLEDGMENT
The Glaxo Laboratories (India) Lt.d., kindly supplied the
vitamins A and D concentrate (Adexolin liquid) used in this
investigation.
L I T E R A T U R E CITED
ALBERT,S., AND C. P. LEBLOND
1946 The distribution of the Feulgen and 2,
4-dinitrophenylhydrazinereaction in normal, castrated, adrenaleetomized
and hormonally treated rats. Endocrinology, 39 : 386-400.
BANERJEE,
S. 1945 Relation of scurvy to the adrenalin content of the adrenal
glands of guinea pigs. J. Biol. Chem., 259 :327-331.
BANERJEE,S., AND K. C. GHOSH 1946 Adrenalin in scurry. J. Biol. Chem., 1 6 6 :
25-29.
1947 Relation of scurry to glucose tolerance test, liver glycogen, and
insulin content of pancreas of guinea pigs. J. Biol. Chem., 190: 207211.
BANERJEE,
S., AND C. DEB 1951 Effect of scurvy on cholesterol and ascorbic acid
i n guinea pigs adrenals. J. Biol. Chem., 190: 177-180.
195% Adrenal cortical activity in scurvy. Endocrinology, 51 : 572576.
19521, Urinary excretion of 17-ketosteroids in scurvy. J . Riol. Chcm.,
194: 575-577.
BOURNE,
G . H. 1953 Biochemistry and Physiology of Nutrition. Vol. 11, Acadeinir Press, h e . , Ncw Pork. Pp. 79-95.
HESS,A. 3’. 1920 Scur-q, Past and Present. .J. B. Lippincott, Co., Philadelphia.
SCHULTZ,
A. 1924 Eine Methode des mikrochemisclien Cholesterinnachn-eises am
Gewehsschnitt. Zentr. nllgem. Path. u. path. Anat., 35: 314-317.
PLATE 1
FWPLANATION
or
FIWRES
Adronal cortex of a noniial gniues pig. X 150.
Ailrenal cortex of an acute aeorbntic gniuea pig, showing haen~orrliagein eonn
rutioiilaris aud degeuarstioe changes iii eona fnwiculnta. X 150.
Aili*eiiwlcortex of n chronic worhutic guiivr pig, showing haeluorrlirigo and
degoueration of mna faacicnlata. x 150.
4
Ailrennl cortex of a normal guinea pig, slioring distribution of cholesterol in
xoiin gloniorulo8n tiiid xoim fasciculatn. x 130.
Ailroiinl colter of ti elironic scorbutic. guinea pig, showing distribution of
clialesterol on1;p in the eona glomerdosll. X 350.
Adrenal cortex of i aormul guinea pig, showing clistributioii of ketosteroids
uirlp in the eom glomerulosa. X 150.
Ailreiinl cortos of a chronic scorbutic guinea pig, showing J)Reiice of ketosteoida X 150.
.4ilreunl cortex o t a nornial monkey. X 150.
htlruiirl cortex of
retienlriris. X 150.
ti
worbutic monkey, showing haeniorrhage in tlie zonn
912
913
PLATE 2
EXPLA4NATIOhT OF YIGURES
10
Section of testcs of a nornial guinca pig.
x
150.
11 Section of testes of a n acute seorbutic guinea pig, showing disappearance of
Leydig's cells. X 150.
1.2 Section of tcstes of a chronic scorbutic guinea pig, slrowing coniplete degeneration of Leydig's cells with fibrosis. Seminiferous tubules are necrosed. X 150.
13 Section of liver of a iiorrrial monkey. X 150.
3.4 Section of livcr of a scorlmtie monkey, shorn-ing haeniorrhage. X 150.
15
Section of liver of a, iiornial guinea pig.
x
150.
If3 Section of liver of an acute scorbutic guinea pig, sliowing e a r l r dcgenei ative
clinngcs. X 150.
17
Section of liver of a chronic scorbutic guinea pig, sho~vingf a t t y degeneration
of hepatic cells. X 150.
18
Section of kidney of a normal guinea pig. X 150.
19 Section of a kidney of a chronic scorbutic guinea pig, showing atrophy of the
glomeruli and degeneration of proximal convoluted tubules. x 150.
20
Section of kidney of a normal monkey. X 150.
21
Section of kidney of a scorbutic monkey, showing normal glomeiuli but dogenerative changes in the proximal convoluted tuhulcs. X 150.
914
PLATE 2
915
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