A forced limitation of the motor activity in rats (from 5 to 60 days) results in reduced incorporation of glycine 2-C14 in the total acid insoluble proteins of limb bones and its increase in the teeth and mandibular-maxillary bones. Daily administration of five micrograms of thyrocalcitonine together with polyvinylpyrrolidone normalizes the protein metabolism in the bone tissues during the 40 days of experimentation

Potkin, V. Y.

Stekolnikov, L. I.

Uglova, N. N.

Volozhin, A. I.

English

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INCORPORATION OF GLYCINE-2-C1 IN ACID-INSOLUBLE PROTEINS
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OF RAT BONES AND TEETH DURING HYPOKINESIA AND ADMCNISTRA-
TION OF THYROCALCITONINE
BY
A. I. Volozhin, L. I. Stekol'nikov, N. N. Uglova and
V. Ye. Potkin
Translation of "Vklyucheniye glitsina-2C14 v kislotonerastvorimyye belki
kostey i zubov krys pri gipokinezii i vvedenii tirokal ' tsitonina",
Farmakologiya i Toksikologiya, Vol. 37, No. 2, 1974, pp 223-226
(NASA-TH-75981) IDcOjpOBATI011 of
	 I80
-22959yBUCIN8-2-C-14 Ill ACID-INSOLUBLE PROTEINS OF
	 a' 1 1BAT BODES AND TEETH DOBING BTFORINE.SIA ADD
	 ' F 01ADBINISTBATION OF THISOCALCITONINE (National	 h Qnclas Aeronautics and Space Adaf aistratioa) 8 p G3/5 i 47659
	
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INOORPORATION OF GLYCINE -2-C14 IN ACID-INSOLUBIE PROTEINS OF
RAT BONES AND TEETH DURING HYPOKINESIA AND ADMINISTRATION OF
THYROCALCITONINE
by
A. I. Volozhin, Z. I. Stekol'nikov, N. N. Uglova and V. Ye.
Potkin, Department of Pathological Physiology (head--Acade-
mician of USSR Academy of Medical Sciences Professor N. A.
Fedorov) of the Moscow Medical Stomatological Institute,
Section of Technology of Endocrine Preparations (head--Can-
didate of Biological Sciences V. Ye. Razmakhain) of the
All-Union Scientific Research Institute of Antibiotics, Moscow
A forced limitation of the motor activity in rats (from
5 tg460 days) results in a reduced incorporation of glycine-
2-C in the total acid-insoluble proteins of limb bones and
its increase in the teeth and mandibula.rimaxillaxy bones.
Daily administration to these rats of 5 of thyrocalitonine
together with polyvinylpyrrolidone normalizes the protein
metabolism in the bone tissues during all the 40 days of
experimer Lation.
The thyroid gland hcrmone thryocalcitonine (TCT) can be used to
prevent disruption in the metabolic processes in the bone tissue during
prolonged limited motor activity (Alliapoulios et al., 1966), which, as is
known is accompanied by osteoporosis and a negative calcium balance
(Ye. N. Biryukov et al., 1966= Issecutz and Blizzard, 1967). We studied
the effect of TCT on certain aspects of the protein exchange in bone and
teeth tissues of animals in a normal motor pattern and in hypokinesia.
Methods of Study
Experiments were conducted on 95 albino mongrel male rats with initial
weight 1?0-130 g. All the animals were divided into 4 groups: first--
control,second--under conditions of hypokinesia, third--received TCT,
Numbers in margin indicate pagination in original foreign text.
1
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fourth--receivM TCT on the background of hypokinesia. Hypokinesia was
created by placing the rats in close cages in which movement of the animals
was sharply limited. Here the dimensions of the cages did not prevent
normal consumption of water and food by the rats. Rats in the third and
fourth groups were daily given a subcutaneous administration of 5 pg of a
complex preparation of TOT with polyvinylpyrr©lidons (PVP) obtained by the
method of L. I. Stekol'nikov at al. (1969x) and which has a prolonged hypo t
-calcemic effect (L.I. Stekol'nikov at al., 1969b).
Within 5,10, 20, 40 and 60 days after the start of the experiment
the rats (4-5 from each group) were given subcutaneous injections of 2 pCi
each of glycine-2-C14 diluted in a physiological solution. In a day after
administration of the isotope the animals were decapitated and the total
protein content was determined in the blood plasma according to Lowry. The
molars, incisors, lower and upper maxillary, humeral, femoral and tibial bones
were removed, thoroughly cleaned of soft tissues, washed in running water,
dehydrated, degreased and decalcified in 1 n. of HC1 solution according to
the accepted technique (A. A. Prokhonchukov and Lyu Din-sin', 1961).
The acid-insoluble bone protein that was dried to a constant weight
was exposed to hydrolysis by concentrated HC1 on backings made of fluoro-
plastic and dried. Radiometry-was conducted on a gas-flow counter SOT-30-
BFL. The magnitude of radiation attenuation was computed in the mass of the
preparation. The C 14 content in the protein of bones and teeth was expressed
in percents of the administered isotope dose. The results were processed
by the method of variation statistics according to Student and were con-
sidered reliable with D < 0.05.
Results and Their Discussion
The concentration of total blood protein in the rats (fig. 1) in
hypokinesia (second group) had a tendency to drop, which corresponds to
our previous data (A. I. Volozhin, 1971). The administration df TCT
Preparation to animals of the third group resulted in an increase in the
blood protein level on the fifth and tenth days, and then a decrease on the
40th and 60th days of the experiment. In rats of the fourth group the
2
IV --is
Figure 1. Content of Total Protein (in
% of Control) in Blood Plasma of Rats in
Control
	
	
Hypokinssia (1), Administration of
Thyrocalcitonine (2), as well as with
Hypokinesia. and Simultaneous Administration
of Thyrocalcitonine N
-
F I  ,
S	 IM	 40	 60; Days
changes in the level of blood plasma protein did not significantly differ
	
224
from those in the third group.
The content of acid-insoluble protein in the molars and inoisc:s of
rats in the second group increased on the 20th and 40th days respectively.
In the humeral bones the quantity of protein was increased on the 40th day
and considerably"dropped on the 60th day of hypokinesia (78%). A decrease
in the amount of protein was also noted in the femoral ( 10th and 60th days)
and in the tibial bones (on the 20th and 60th days).
The administration of TCT to the freely placed rats ( third group)
produced a reduction in the protein content in the molars ( on the fifth and
tenth days) and an increase of it in the incisors and maxillary bones (on
the 40th day). The quantity of acid-insoluble protein in the humeral (on
the 60th day), tibial ( on the 60th day) and in the femoral bones (on the
fifth, tenth and 60th days of TCT administration) was reduced. In the
humeral, femoral and tibial bones of rats in the fourth group the quantity
of protein exceeded that in the control, as well as in rats of the second
and third groups for the entire length of the experiment.
Data on incorporation of glycine-2-C 14 in the acid-insoluble proteins
	 22
of bones and teeth of rats under the conditions of our experiment are
presented in figure 2.
It is important to study the reactivity ( according to indices for
incorporation of glycine -2-C14
 in proteins) of the bone tissue and teeth of
rats under conditions of limited motor function. We speak of the reactivity
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Figure 2. Incorporation of Glycine-2-C 14 (in 1% of Control taken as 100) in
Proteins of Bones and Tooth of Rats during Hypokinesia and Administration
of Thyroca.lcitonine.
A--molars = B--incisors = C--lower maxilla; D--upper maxillas E--shoulders
F-•-femurs G--tibia. 1,2,3--the same as in fig. 1.
of the bone tissue in relation to TCT on that oasis that the bone apparently
is the "critical" organ on which the indicated hormone acts (0'Riordan and
A urbach, 1968) .
To. clarify this question the indices of protein exchange in the
mineralized tissues obtained in the second group of rats were compared with
the corresponding indices obtained in the fourth group. Here it was found
4
fthat in all the periods of observation (except the 60th day) the incorpora-
tion of C14 into the teeth and bones of the rats of the fourth group was con-
siderably higher than in the "untreated" animals (second group).	 In contra-
diction to this the level of incorporation of C14 into the proteins of the
calcified tissues of the rats in the third group is higher than in the
animals of the first group, only in the teeth and maxillary bones, and
in the humeral, femoral and tibial bones is lower than in the control, on the
t fifth, tenth and 20th days.	
Thus, a change in the reactivity of the bone
tissue to the TCT in rats under conditions of hypokinesia occurs in the bones
of the skeleton whose function is significantly diminished, i.e., in the
E
x
bones of the extremities.
The protein matrix of the mineralized tissues consists by 95% of an
acid-insoluble collagen in which about 1/3 of the amino acid residues are
glycine.	 Therefore recording of the incorporation of glycine in the acid-
` insoluble proteins of the bones and teeth is an important indicator of the
synthetic processes in the protein matrix of these tissues.
During hypokinesia in rats a considerable inhibition
	 occurs of the
incorporation of glycine-2-C 14
 in the acid-insoluble proteins of the bones
of the extremities, apparently indicating the decrease in the synthetic
processes in the protein matrix of the bones.
	 The use of TCT under con-
ditions of limited movement has a signticant normalizing effect on the
incorporation of glycine-2-C- 14
 in the bones of rats.
	 It is possible that
the
	 difference in the effect of TCT on the freely placed rats and animals
with limited movement is linked to the change in the initial condition of
the bone tissue emerging under the influence of hypokinesia.
Conclusions
1.	 Limited mobility in rats for 5
-60 days results in a suppression of
the incorporation of glycine-2-C 14
 into the total acid-insoluble proteins of
the bones of the extremities.
2. Administration of thyrocalcitonim to rats during hypokinesia
normalizes the incorporation of glycine into the total acid-insoluble
proteins of the bone tissue.
5
a_t
m
References
Biryukov, Ye. N., Kakurin, L. I. at al., Kosmicheskaya biol, No. 2 (1967),
p. 74.
Volozhin, A.I. Ibid, No. 2 (19-1), P. 17.
Prokhonchukov, A., Lyu Din-Sin', Mod. radiol, No. 7 (1961), P. 70.
Stekol'nikov, L. I., Katkovskiy, S. B., at al., Biokhimiya,  No. 1 (1969a),
P. 115.
Stekol'nikov, L. I., Tepelina, 0. M., at al., Vopr, med. khimii, Vol. 15
(1969), p. 635.	 - ---
Allia.poulios, M. A., at al. Science, Vol. 155 (1966), p. 3708.
Issecutz, B, Blizzard, J. J. apal. Physiol., Vol. 21 (1966), p. 1013.
O'Riordan, J. L. H., A urbach, G. D. Endocrinology, Vol. 82 (1968), p. 377•


Incorporation of glycine-2-C-14 in acid-insoluble proteins of rat bones and teeth during hypokinesia and administration of thyrocalcitonine

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