OVINE FETAL ADRENAL SYNTHESIS OF CORTISOL - REGULATION BY ADRENOCORTICOTROPIN, ANGIOTENSIN-II AND TRANSFORMING GROWTH-FACTOR-BETA

An increase in cortisol production by the fetal adrenal cortex is an important prepartum event. The increase in ovine fetal adrenal synthesis of cortisol appears to rely in part on the ACTH induction of 17-alpha-hydroxylase cytochrome P-450 (P-450(17-alpha)) which occurs before parturition. In the present study we examined the effect of ACTH treatment on cortisol production and P-450(17-alpha) expression using primary cultures of ovine fetal adrenal cells. In addition, we examined the effects of angiotensin II (A-II) and transforming growth factor-beta (TGF-beta) on ACTH-treated cells. We have demonstrated previously that these factors modulate P-450(17-alpha) levels in adult ovine and bovine adrenal cells. Fetal ovine adrenal cells were isolated from 126- to 130-day fetuses (term = 144 +/- 3 days) and placed in monolayer cell culture. After 1 day in culture the cells were treated with ACTH (10 nM) with or without A-II (0.1-100 nM) or TGF-beta (1-100 pM). Medium content of cortisol was low under basal conditions, whereas ACTH-stimulated cortisol production by 10- to 100-fold. A-II and TGF-beta inhibited ACTH-induced cortisol production by 70-90%. In addition, 3 days of treatment with ACTH caused a greater than 10-fold induction of P-450(17-alpha) enzyme activity in fetal adrenal cells. A-II and TGF-beta inhibited the ability of ACTH to induce P-450(17-alpha) activity by at least 75%. Using an antibody to P-450(17-alpha) and immunoblotting techniques the effects of ACTH, A-II, and TGF-beta on enzyme activity were observed to correspond to cellular levels of P-450(17-alpha) protein. The inhibitory effects of TGF-beta and A-II could not be overcome by the cAMP analog (Bu)2cAMP. Interestingly, the expression of the enzyme 3-beta-hydroxysteroid dehydrogenase was much less sensitive to inhibition by A-II or TGF-beta. The ability of A-II and TGF-beta to suppress P-450(17-alpha) expression could play a role in determining the pathway of steroidogenesis and specifically the amount of cortisol produced by fetal adrenocortical cells in vivo.