0021-972X/79/4902-0284$02.00/0 Journal of Clinical Endocrinology and Metabolism Copyright © 1979 by The Endocrine Society
Vol. 49, No. 2 Printed in U.S.A.
Kinetics of Human Chorionic Gonadotropin-Induced Steroidogenic Response of the Human Testis. II. Plasma 17a-Hydroxyprogesterone, A4-Androstenedione, Estrone, and 17/?-Estradiol: Evidence for the Action of Human Chorionic Gonadotropin on Intermediate Enzymes Implicated in Steroid Biosynthesis* MAGUELONE G. FOREST, ANNICK LECOQ, AND JOSE M. SAEZ Unite de Recherches Endocriniennes et Me'taboliques chez I'Enfant, INSERM U. 34, and Unite de Recherches sur le Controle Hormonal des Activites Cellulaires, INSERM U. 162, Hopital Debrousse, 69322 Lyon Cedex 1, France
ABSTRACT. The time course of the rise in plasma levels of 17a-hydroxyprogesterone (OHP), A4-androstenedione (A4), estrone (Ei), and 17/J-estradiol (E2) was studied in 14 normal adult volunteers after one, two (at a 24-h interval), or four (at 5-day intervals) injections of 6000 IU hCG. The patterns were quite different among the four hormones. After a single hCG injection, OHP, A4, and Ei increased within 4 h, but E2 increased after 812 h. After this early rise, A4 decreased slightly, plateauing for 48 h, E| slightly increased, while OHP and E2 sharply rose, reaching their maximal levels at 24 h (mean, 3- and 5-fold respectively). At 48-96 h (mean, 72 h), A4 and Ei showed a paradoxical late rise (2- to 3-fold) significantly greater than the early one, while OHP and E2 continued to decline. When hCG injection was repeated 24 h later, A4 and Ei reincreased slightly within 4 h but exhibited a similar late rise delayed by about 24 h; the single maximal peak of OHP was somewhat advanced and that of E2
was amplified. When hCG was administered every 5 days, the levels of A4, Ei, and E2 found before each hCG injection increased from days 0-10 but were not different from control values for OHP. The early responses to hCG were no longer significant after the third injection for A4 and Ei, while they occurred each time for OHP. Variations in plasma E2 were not significant during the 4 h after any of the hCG injections. These findings suggest that 1) the hCG-induced steroidogenic block in testosterone biosynthesis occurs before the synthesis of A4; 2) the earlier rise and lack of late increase in OHP and E2 suggest that hCG acutely stimulates the testicular aromatase while it inhibits the 17, 20-lyase activity; and 3) the drastic rise in E2 together with previous data in humans and rats suggest that this hormone might be responsible for the latter transient enzymatic defects. (J Clin Endocrinol Metab 49: 284, 1979)
H
UMAN CG has been widely used for many years to assess the secretory reserve of interstitial cells of the testis. Various protocols and dosages of hCG have been proposed (1-20). In all of them, estimation of the concentration of testosterone has been used as the indicator of testicular function. The secretion of other androgens and estrogens by the human testis is well known (21-23) and their increase in the spermatic vein after hCG administration has been demonstrated (23-25). A4Androstenedione (A4) and testosterone (T) can also be converted into estrone (Ei) and estradiol (E2) at the
peripheral level (26, 27). Several authors have studied, ir prepubertal boys or adults, the plasma or urinary response of E2 to hCG stimulation (1-8, 25). There are verj scarce and sometimes conflicting data on the effect ol various protocols of hCG stimulation, used either ir children or in adults and often in pathological subjects on the plasma concentrations of Ei (7-9, 25), dehydro epiandrosterone (9-12, 25) or its sulfate (10), A4 (9, 1114, 25), or 17a-hydroxyprogesterone (OHP) (9,14-16, 25) They all show that hCG is able to stimulate the testicula: production of A4 and OHP. However, there is no system atic and simultaneous study of the increase in plasm* levels of the precursors of testosterone and other andro gens or estrogens after a single injection of hCG. In a previous study (28), we have shown that th< increment in T plasma levels occurring 48-72 h after i single injection of hCG is somewhat in the range of thosi observed after several injections (8,17-20). On the othe
Received January 29, 1979. Address requests for reprints to: Dr. Maguelone G. Forest, Unite de Recherches Endocriniennes et Metaboliques chez I'Enfant, INSERM U. 34, Hopital Debrousse, 29 rue Soeur Bouvier, 29322 Lyon Cedex 1, France. *This work was supported by INSERM Grant ATP-33-76-35, DGRST Grant 78-7-9364, and the Fondation pour la Recherche Medicale Francaise. 284
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 19 November 2015. at 02:14 For personal use only. No other uses without permission. . All rights reserved.
hCG AND TESTICULAR STEROIDOGENESIS IN HUMANS
hand, the early decline in T levels which follows each hCG injection was interpreted as an hCG-induced testicular steroidogenic desensitization (28). In the present study, the plasma concentrations of the two immediate precursors of T, A4 and OHP, and those of Ei ane E2 were measured after one, two, or repeated injections of hCG. From their respective dynamics of response to hCG, we present indirect evidence that hCG has specific actions on different microsomial enzymes implicated in the steroid biosynthesis of the human testis. Materials and Methods The subjects were 14 normal adult volunteers (28), ranging in age from 25-43 yr, with a mean weight of 66.2 ± 5.9 kg (SD). Eight subjects received a single im injection of 6000 IU hCG (protocol I). Three of the latter group and 4 other subjects received 2 iv injections of 6000 IU hCG at 24-h intervals (protocol II). Blood was drawn before the first injection for determination of baseline levels of plasma OHP, A4, Ei, and E2. Samples were also obtained 1, 2, 4, 6, 8, 12, and 24 h after each hCG injection, then every 24 h for 7 days. The last 4 subjects received 4 iv injections of 6000 IU hCG every 5 days (protocol III). Blood was drawn before and 2 and 4 h after each hCG injection. All samples were frozen at —20 C until analysis. All hormones were measured by RIAs. From a first plasma aliquot, A4 and OHP were separated on celite chromatography, as previously described (29, 30), while Ei and E2 were extracted by ethyl ether from a second 3-ml plasma sample and separated oh a second celite column (31). Characteristics of the anti-OHP antibody used have already been fully described (30). A4 was measured with an antiserum raised in the rabbit against lla-OH-A4-hemisuccinyl linked covalently to bovine serum albumin. This antiserum had significant (>0.5%) cross-reaction only with lla- and ll/?-hydroxyprogesterone and 11-deoxycorticosterone, which were fully separated from A4 in the celite chromatography. Ei and E2 were measured by a RIA technique already described (31) but using more specific antisera (kindly provided by Roussell laboratory) raised against their respective derivatives linked at the C6 position to bovine serum albumin. The anti-E2 antibody had cross-reactions of 3.6% with 16a-hydroxyestrone, 1.9% with E2, 0.1% with 17a-estradiol, and 0.09% with estriol. The anti-E2 antibody had 3.6% cross-reaction with Ei and estriol and less than 0.5% with other estrogenic compounds. Neither antisera had any cross-reaction (
Vol. 49, No. 2 Printed in U.S.A.
Kinetics of Human Chorionic Gonadotropin-Induced Steroidogenic Response of the Human Testis. II. Plasma 17a-Hydroxyprogesterone, A4-Androstenedione, Estrone, and 17/?-Estradiol: Evidence for the Action of Human Chorionic Gonadotropin on Intermediate Enzymes Implicated in Steroid Biosynthesis* MAGUELONE G. FOREST, ANNICK LECOQ, AND JOSE M. SAEZ Unite de Recherches Endocriniennes et Me'taboliques chez I'Enfant, INSERM U. 34, and Unite de Recherches sur le Controle Hormonal des Activites Cellulaires, INSERM U. 162, Hopital Debrousse, 69322 Lyon Cedex 1, France
ABSTRACT. The time course of the rise in plasma levels of 17a-hydroxyprogesterone (OHP), A4-androstenedione (A4), estrone (Ei), and 17/J-estradiol (E2) was studied in 14 normal adult volunteers after one, two (at a 24-h interval), or four (at 5-day intervals) injections of 6000 IU hCG. The patterns were quite different among the four hormones. After a single hCG injection, OHP, A4, and Ei increased within 4 h, but E2 increased after 812 h. After this early rise, A4 decreased slightly, plateauing for 48 h, E| slightly increased, while OHP and E2 sharply rose, reaching their maximal levels at 24 h (mean, 3- and 5-fold respectively). At 48-96 h (mean, 72 h), A4 and Ei showed a paradoxical late rise (2- to 3-fold) significantly greater than the early one, while OHP and E2 continued to decline. When hCG injection was repeated 24 h later, A4 and Ei reincreased slightly within 4 h but exhibited a similar late rise delayed by about 24 h; the single maximal peak of OHP was somewhat advanced and that of E2
was amplified. When hCG was administered every 5 days, the levels of A4, Ei, and E2 found before each hCG injection increased from days 0-10 but were not different from control values for OHP. The early responses to hCG were no longer significant after the third injection for A4 and Ei, while they occurred each time for OHP. Variations in plasma E2 were not significant during the 4 h after any of the hCG injections. These findings suggest that 1) the hCG-induced steroidogenic block in testosterone biosynthesis occurs before the synthesis of A4; 2) the earlier rise and lack of late increase in OHP and E2 suggest that hCG acutely stimulates the testicular aromatase while it inhibits the 17, 20-lyase activity; and 3) the drastic rise in E2 together with previous data in humans and rats suggest that this hormone might be responsible for the latter transient enzymatic defects. (J Clin Endocrinol Metab 49: 284, 1979)
H
UMAN CG has been widely used for many years to assess the secretory reserve of interstitial cells of the testis. Various protocols and dosages of hCG have been proposed (1-20). In all of them, estimation of the concentration of testosterone has been used as the indicator of testicular function. The secretion of other androgens and estrogens by the human testis is well known (21-23) and their increase in the spermatic vein after hCG administration has been demonstrated (23-25). A4Androstenedione (A4) and testosterone (T) can also be converted into estrone (Ei) and estradiol (E2) at the
peripheral level (26, 27). Several authors have studied, ir prepubertal boys or adults, the plasma or urinary response of E2 to hCG stimulation (1-8, 25). There are verj scarce and sometimes conflicting data on the effect ol various protocols of hCG stimulation, used either ir children or in adults and often in pathological subjects on the plasma concentrations of Ei (7-9, 25), dehydro epiandrosterone (9-12, 25) or its sulfate (10), A4 (9, 1114, 25), or 17a-hydroxyprogesterone (OHP) (9,14-16, 25) They all show that hCG is able to stimulate the testicula: production of A4 and OHP. However, there is no system atic and simultaneous study of the increase in plasm* levels of the precursors of testosterone and other andro gens or estrogens after a single injection of hCG. In a previous study (28), we have shown that th< increment in T plasma levels occurring 48-72 h after i single injection of hCG is somewhat in the range of thosi observed after several injections (8,17-20). On the othe
Received January 29, 1979. Address requests for reprints to: Dr. Maguelone G. Forest, Unite de Recherches Endocriniennes et Metaboliques chez I'Enfant, INSERM U. 34, Hopital Debrousse, 29 rue Soeur Bouvier, 29322 Lyon Cedex 1, France. *This work was supported by INSERM Grant ATP-33-76-35, DGRST Grant 78-7-9364, and the Fondation pour la Recherche Medicale Francaise. 284
The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 19 November 2015. at 02:14 For personal use only. No other uses without permission. . All rights reserved.
hCG AND TESTICULAR STEROIDOGENESIS IN HUMANS
hand, the early decline in T levels which follows each hCG injection was interpreted as an hCG-induced testicular steroidogenic desensitization (28). In the present study, the plasma concentrations of the two immediate precursors of T, A4 and OHP, and those of Ei ane E2 were measured after one, two, or repeated injections of hCG. From their respective dynamics of response to hCG, we present indirect evidence that hCG has specific actions on different microsomial enzymes implicated in the steroid biosynthesis of the human testis. Materials and Methods The subjects were 14 normal adult volunteers (28), ranging in age from 25-43 yr, with a mean weight of 66.2 ± 5.9 kg (SD). Eight subjects received a single im injection of 6000 IU hCG (protocol I). Three of the latter group and 4 other subjects received 2 iv injections of 6000 IU hCG at 24-h intervals (protocol II). Blood was drawn before the first injection for determination of baseline levels of plasma OHP, A4, Ei, and E2. Samples were also obtained 1, 2, 4, 6, 8, 12, and 24 h after each hCG injection, then every 24 h for 7 days. The last 4 subjects received 4 iv injections of 6000 IU hCG every 5 days (protocol III). Blood was drawn before and 2 and 4 h after each hCG injection. All samples were frozen at —20 C until analysis. All hormones were measured by RIAs. From a first plasma aliquot, A4 and OHP were separated on celite chromatography, as previously described (29, 30), while Ei and E2 were extracted by ethyl ether from a second 3-ml plasma sample and separated oh a second celite column (31). Characteristics of the anti-OHP antibody used have already been fully described (30). A4 was measured with an antiserum raised in the rabbit against lla-OH-A4-hemisuccinyl linked covalently to bovine serum albumin. This antiserum had significant (>0.5%) cross-reaction only with lla- and ll/?-hydroxyprogesterone and 11-deoxycorticosterone, which were fully separated from A4 in the celite chromatography. Ei and E2 were measured by a RIA technique already described (31) but using more specific antisera (kindly provided by Roussell laboratory) raised against their respective derivatives linked at the C6 position to bovine serum albumin. The anti-E2 antibody had cross-reactions of 3.6% with 16a-hydroxyestrone, 1.9% with E2, 0.1% with 17a-estradiol, and 0.09% with estriol. The anti-E2 antibody had 3.6% cross-reaction with Ei and estriol and less than 0.5% with other estrogenic compounds. Neither antisera had any cross-reaction (
