ADRENAL STEROIDS AND BODY COMPOSITION*

Transcription

1 ADRENAL STEROIDS AND BODY COMPOSITION* BY CHARLES D. KOCHAKIANt AND EVANGELINE ROBERTSON (From the Department of Physiology and Vital Economics, School of Medicine and Dentistry, The University of Rochester, Rochester, New York) (Received for publication, August 11, 1950) In a previous report (1) the effect of adrenal cortical steroids on body weight and nitrogen metabolism in correlation with enzymic changes has been presented. In this study, an analysis of the tissues of the mice has been made in order to obtain a further insight of the mechanism and site of action of these steroids. Procedure Animals and Treatment-The mice were those of a previous report (1). They were castrated at 17 to 19 gm. in body weight and were maintained usually two, occasionally one or three, in a glass jar in an air-conditioned room at They were fed ad libitum the following diet: casein 16.7, sucrose 61.2, hydrogenated vegetable oil 7.4, dry brewers yeast (Fleischmann s No. 2019) 9.2, Cellu flour 1.8, Wesson s salt mixture 3.7 (a), and three times per week a supplement of 1 drop of cod liver oil and 1 drop of a 34 per cent tocopheroll concentrate of wheat germ oil diluted lo-fold with Wesson oil. The animals were weighed three times per week. The experiments were begun 40 to 60 days after castration. The mice in each group of experiments were from a single batch and were studied at the same time, except those of the 21 day experiment with cortisone acetate and 11-desoxy-17-hydroxycorticosterone acetate, which were studied in two equal groups at an interval of 1 week. The mice were paired aecording to body weight at the time of implantation of the corticoid pellets, which weighed approximately 14 mg. each (3). 14 sham operation of implantation of the pellet was performed on the control animals. Preparation of Tissues and Determination of Moisture-The analyses of * This investigation was supported by a grant from the American Cancer Society on recommendation of the Committee on Growth of the National Research Council. The 11-dehydrocorticosterone was provided by Dr. E. C. Kendall and Merck and Company, Inc., the cortisone acetate and the Il-dehydrocorticosterone acetate were provided by Merck and Company, Inc., and the 11-desoxy-17-hydroxycorticosterone acetate by Dr. K. Miescher. t Present address, Oklahoma Medical Research Institute, 825 North East 13th Street, Oklahoma City, Oklahoma. 1 The tocopherol concentrate was provided by Distillation Products, Inc., through the courtesy of Dr. P. L. Harris. 495

2 496 ADRENAL STEROIDS AND BODY COMPOSITION the tissues were essentially as previously reported (4). At autopsy approximately 0.5 gm. of the liver and one kidney were saved for the enzyme determination (cf. (I)). The remaining liver and the other kidney were saved for analyses. The rest of the mouse, including the previously weighed thymus and spleen, is designated as carcass. The tissues were dried to constant weight at in an electric oven attached to a water aspirator. The dried carcass was dissolved with warming in 30 ml. of 50 per cent potassium hydroxide and 30 ml. of redistilled ethyl alcohol, and then made t,o 200 ml. with distilled water. The liver and kidneys were dissolved with warming in 10 ml. of 50 per cent potassium hydroxide and 10 ml. of redistilled alcohol. The liver digest was diluted to 50 ml. and the kidney to 25 ml. with redistilled wat)er. Nitrogen Determination-The micro-kjeldahl procedure was used for all nitrogen determinations on 1 ml. aliquots of the above dilutions. Total Fat Determination-The previously report ed modification (4) of the Leathes and Raper method (5) was used on aliquots of 10 ml. for the carcass and 15 to 25 ml. for the liver and kidney. Results Effect of Cortisone Acetate on Carcass-The consistent decreases in body weight produced by 11-dehydro-17-hydroxycorticosterone acetate (cf. (1)) were not accompanied by any major alterations in the per cent composition of the various constituents of the carcass (Table I). Thus, the amounts of the different substances changed in proportion to the total weight of the carcass (Fig. I), except that at 2 days no loss in fat occurred and at 30 days the amount of water loss was decreased sufficiently to compensate for the further loss in fat. The ratio of the lost water to that of nitrogen (Fig. 1) was higher than normal (6), even after 30 days. The composition of the smaller loss in body weight obtained at 7 days with the lower dose of cortisone acetate (Table I) was the same as at the higher dose except that the fat was slightly, but not significantly, increased instead of decreased. Effect of Cortisone Acetate on Liver and Kidney-The weight of the liver (Table II, Fig. 2) is slightly increased after 2 days of 11-dehydro-17- hydroxycorticosterone acetate (cortisone acetate) and then consistently decreased. The change in t.he weight is due mainly to changes in fat and water (Fig. 2) especially after 21 and 30 days (Table II, Fig. 2). I1 -Desoxy-I 7-hydroxycorticosterone Acetate-The subcutaneous implantation of a pellet of 11-desoxy-17-hydroxycorticosterone acetate for 21 days did not alter the weight or the composition of the body. II-Dehydrocorticosterone-The decrease in carcass weight (Table III) was

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4 498 ADRENAL STEROIDS AND BODY COMPOSITION due to a loss of water and protein (Table V). The increase in fat was small (Tables III, V) and not sufficient (Table V) to compensate for the loss of the other constituents. The lower dose of the corticoid produced similar changes, but to a much lesser degree. The changes in the constituents of the liver were very small (Tables IV, V). 11-Dehydrocorticosteone Acetate-The daily subcutaneous injection for 100 IO 20 DAYS FIG. 1. Changes produced by cortisone acetate in the carcass of castrated mice. The values represent the differences between the respective values of the control and treated mice. See Table I. 25 days of 0.5 mg. of ll-dehydrocorticosterone acetate, dissolved at 5 mg. per ml. in sesame oil containing 10 per cent benzyl alcohol, had no effect on the weight or composition of the tissues of castrated mice. 1. DISCUSSION The production by cortisone acetate of a loss in body protein and fat is in general agreement with the previous observations by Kendall and Heilman (7) and this laboratory (8). It is significant that the total loss in protein occurs within the first few days and parallels the involution of the lymphatic tissue and the extra nitrogen excretion. Furthermore, the loss in body protein agrees very well with the nitrogen balance data (1).

5 TABLE II Effect of Cortisone Acetate on Composition of Liver and Kidney of Castrated Mice* Treatment Liver Kidney Weight water Protein Fat Weight 1 Water (Protein/ Fat 2 day experiments w per cetat per cent per cent mg. $%I cent per cent,2$ Control i f f i f P 5.7 Cortisone acetate i f f f zk g 8 7 day experiments h d - Control & f f A i E Cortisone acetate i f f f f I I (1:l) i i f f f m 21 day experiments Control.. Cortisoneacetate day experiments Control f rt It A f Cortisoneacetate f f f 0.6 / 7.9iO:2/403-t12/75:Ojl6:415:5 * See Table I for experimental details, number of animals, and statistical treatment of the data. The tissues of two mice were prepared as a sample, except in the odd numbered groups, in which one of the samples necessarily contained the tissue of a single animal.

6 500 ADRENAL STEROIDS AND BODY COMPOSITION The extremely high loss of water compared to nitrogen indicates that the type of tissue lost is embryonic in nature (6). This was specially true during the phase of protein catabolism and loss of lymphatic tissue (1). The subsequent reduction is suggestive of a readjustment in internal metabolic processes. An initial extra excretion of sodium and chloride (and water) has been noted also in rats (9, 10). It is significant that the tissues involved in the so called collagen diseases are mesenchymal derivatives (11) and that these diseases respond rapidly to cortisone administration (cf.(12)). It is specially noteworthy that during the height of protein catabolism fat was being spared. The depletion of the protein stores, however, was IO 20 DAYS FIG. 2. Composition of the changes produced by cortisone acetate in the liver of castrated mice. The values represent the differences between the respective values of the control and treated mice. See Table II. P = protein, F = fat, W = water, WGT = weight. accompanied by a progressive mobilization and utilization of the fat depots. It is quite likely that if the duration of treatment had been extended, with a resulting depletion of fat stores, vital protein structures would be mobilized. On the other hand, a small dose of the corticoid would favor a sparing and accumulation of fat, in agreement with the reports (13-15) that the adrenal cortex is essential for maintenance of peripheral fat. The effect of ll-dehydrocorticosterone on the carcass constituents was qualitatively the same as that of cortisone acetate, but only about onetwentieth as effective. The amount of fat deposited was quite small compared to the previous observations (7,8). It seems premature to attribute the r61e of fat deposition specifically to ll-dehydrocorticosterone and corticosterone and fat catabolism to cortisone until these compounds become available for more detailed study.

7 C. D. KOCHAKIAN AND E. ROBERTSON 501 The liver changes are noteworthy, for during the phase of most rapid loss in body weight this organ increased all of its constituents. A decrease in weight did not occur until the intense protein catabolic phase had ended. Furthermore, the loss in fat and water preceded the loss in protein, which TABLE E$ect of II-Dehydrocorticosterone on Composition of Carcass of Castrated Mice after 7 Days* Treatment Control Dehydrocorticosteronet... (l:l)$. T rto. of mice -- 6 III Steroid absorbed mg. per day Initial body weight CaICaSS Weight Water Protein Fat ~- -- w. &m. ger cc?&1 per cent per cent zko.3 f0.2 ho.6 f0.4 f f0.2 f0.3 zkl.1 zko.1 f f0.3 f0.4 rto.7 lto.4 f0.3 * dba mice generously provided by Dr. S. R. Warner, Biological Station, Springville, New York. See Table I for statistical treatment of the data. t Provided by Dr. E. C. Kendall. $ Pellets prepared from an equal mixture of 11-dehydrocorticosterone (synthetic, Merck) and cholesterol. TABLE IV Effect of il-dehydrocorticosterone on Composition of Liver of Castrated Mice after 7 Days* Treatment No. of I mice I Weight Water Proteint Fatt w. ger cent per celtt per cent Control f f Dehydrocorticosterone A f (1:l) f i * See Table III for experimental details and Table I for statistical treatment of the data. t These analyses were obtained on the pooled samples. was not evident until after 21 days. Adrenocorticotropic hormone at 3 mg. per day for 10 days has been reported (16) to increase the fat and decrease the water and protein of the liver of adult rats. At least one tissue, the kidney, did not give up any of its constituents. Indeed, anabolism occurred in this organ. The increased utilization of fat has now been observed in experiments with protein anabolic steroids (4, 17) and growth hormone (4, 18-22) and by the protein catabolic steroids (23) and appears after the protein ana-

8 502 ADRENAL STEROIDS AND BODY COMPOSITION bolic or catabolic phase has worn off and when the dosage is higher than the physiological level. Is the extra utilization of fat for energy or to provide carbon residues and assist in the reshuffling of endogenous metabolic materials? It is pertinent that protein anabolic processes in certain tissues, accompanied by catabolic processes in other tissues, continue in the above experiments without any detection in the nitrogen balance. Therefore, it is not surprising to note the growth of implanted tumors (24, 10) while the animal is in negative nitrogen balance as a result of corticoid stimulation (10). TABLE V Composition of Changes Produced by II-Dehydrocorticosterone in Castrated Mice after 7 Days* - Treatment Il-Dehydrocorticosterone Total Il-Dehydrocorticosterone (1:l) Carcass Liver Carcass Liver Tissue Total - * See Tables III and IV for experimental details. SUMMARY.- Weight Water Protein Fat - ~ P. m. gm. gm _ ~ ~ Cortisone acetate implanted as a pellet in adult castrated mice stimulated an immediate and rapid loss in carcass protein during the first 7 days, after which no further loss occurred. A parallel but excess loss in water occurred, which diminished by the 30th day. The fat was spared during the period of intense protein catabolism, but thereafter progressively decreased. The constituents of the liver increased after 2 days and then decreased. The constituents of the kidney increased in proportion to the increase in weight. 11-Dehydrocorticosterone was only one-twentieth as effective as cortisone acetate. 11-Desoxy-17-hydroxycorticosterone acetate had no effect on tissue composition. BIBLIOGRAPHY 1. Kochakian, C. D., and Robertson, E., J. Biol. Chem., 190, 481 (1951). 2. Wesson, F., Science, 76, 339 (1932). 3. Kochakian, C. D., Am. J. Physiol., 142, 315 (1944).

9 C. D. KOCHAKIAN AND E. ROBERTSON Koch&an, C. D., and Stettner, C. E., Am. J. Physiol., 166, 255 (1948). 5. Leathes, J. B., and Raper, H. S., The fats, Monographs on biochemistry, London and New York (1925). 6. Moulton, C. R., J. Biol. Chem., 67, 79 (1923). 7. Kendall, E. C., and Heilman, F. R., Conference on metabolic aspects of convalescence, Josiah Macy, Jr., Foundation, New York, 10th meeting, 81 (1945). 8. Kochakian, C. D., Conference on metabolic aspects of convalescence, Josiah Macy, Jr., Foundation, New York, 5th meeting, 146 (1943); 6th meeting, 13 (1944). 9. Ingle, D. J., Sheppard, R., Evans, J. S., and Kuiaenga, M. H., Endocrinology, 37, 341 (1945). 10. Ingle, D. J., Prestrud, M. C., and Rice, K. L., Endocrinology, 46, 510 (1950). 11. Aegerter, E., and Long, J. H., Am. J. Med. SC., 218, 324 (1949). 12. Sprague, R. G., Power, M. H., Mason, H. L., Albert, A., Mathieson, D. R., Hench, P. S., Kendall, E. C., Slocumb, C. H., and Polley, H. F., Arch. Int. Med., 86, 199 (1950). 13. Reiss, M., Epstein, H., Fleischmann, F., and Schwarz, L., Endokrinologie, 17, 302 (1936). 14. Schiffer, F., and Wertheimer, E., Endocrinology, 6, 147 (1947). 15. Wertheimer, E., and Shapiro, B., Physiol. Rev., 28, 481 (1943). 16. Li, C. H., Ingle, D. J., Evans, H. M., Prestrud, M. C., and Nezamis, J. E., Proc. Sot. Exp. BioZ. and Med., 70, 753 (1949). 17. Kochakian, C. D., Robertson, E., and Bartlett, M. N., Am. J. Physiol., 163, 332 (1950). 18. Downs, W. G., Jr., J. Dent. Res., 10, 601 (1930). 19. Wadehn, F., Biochem. Z., 256, 188 (1932). 20. Lee, M. O., and Shaffer, N. K., J. Nub., 7, 337 (1934). 21. Li, C. H., Simpson, M. E., and Evans, H. M., Endocrinology, 44, 71 (1949). 22. Szego, C. M., and White, A., Endocrinology, 44, 150 (1949). 23. White, A., in Pincus, G., Recent progress in hormone research; Proceedings of the Laurentian Hormone Conference, New York, 4, 153 (1949). 24. Heilman, F. R., and Kendall, E. C., Endocrinology, 34, 416 (1944).

10 ADRENAL STEROIDS AND BODY COMPOSITION Charles D. Kochakian and Evangeline Robertson J. Biol. Chem. 1951, 190: Access the most updated version of this article at Alerts: When this article is cited When a correction for this article is posted Click here to choose from all of JBC's alerts This article cites 0 references, 0 of which can be accessed free at tml#ref-list-1