Regulation of 3 beta-hydroxysteroid dehydrogenase delta 5/delta 4-isomerase and cholesterol side-chain cleavage cytochrome P450 by activin in rat granulosa cells.

Activin is a dimeric protein implicated in the local control of follicular steroidogenesis. Using cultured rat granulosa cells, we previously showed that the effect of activin on FSH-induced progesterone synthesis changes with preovulatory follicular development, from positive regulation in nondifferentiated (immature) granulosa cells to negative regulation in preovulatory (mature) granulosa cells. The aim of the present study was to assess development-related effects of activin on the expression of enzymes crucial to progesterone synthesis: cholesterol side-chain cleavage cytochrome P450 (P450scc) and 3 beta-hydroxysteroid dehydrogenase/delta 5-4-isomerase (3 beta HSD). Nondifferentiated granulosa cells were isolated from the ovaries of estrogen-pretreated immature female rats that received no other treatment; differentiated granulosa cells were obtained from similar animals treated for 48 h with human FSH to induce preovulatory follicular development. Cells were cultured for 48 h in serum-free medium with and without human FSH and/or recombinant activin-A, and medium was collected for measurement of progestagens (progesterone, pregnenolone, and 20 alpha-dihydroprogesterone). In cultures of nondifferentiated granulosa cells, activin augmented the FSH-induced production of all three steroids. In differentiated granulosa cells, activin suppressed the FSH-stimulated production of progesterone and 20 alpha-dihydroprogesterone, but had no effect on pregnenolone. The presence of exogenous pregnenolone increased the overall production of progesterone, but did not alter qualitative steroidogenic responses to activin. To assess the interaction between FSH and activin on 3 beta HSD and P450scc messenger RNA (mRNA) expression, Northern blot analyses were performed on total RNA isolated from cultured granulosa cells. Treatment in vitro with FSH alone markedly enhanced the abundance of both the 3 beta HSD and P450scc mRNA transcripts in nondifferentiated and differentiated granulosa cells. FSH-stimulated expression of P450scc mRNA was further enhanced by cotreatment of nondifferentiated granulosa cells with activin. However, activin had no consistent effect on FSH-stimulated expression of 3 beta HSD mRNA in nondifferentiated cells. In differentiated granulosa cells, both mRNAs were suppressed more than 50% by the presence of activin. We conclude that the in vitro effects of activin on FSH-induced expression of 3 beta HSD and P450scc mRNAs in rat GC are similar: initially stimulatory (P450scc) or without effect (3 beta HSD), then becoming completely inhibitory. The mechanism of this development-dependent change in the granulosa cell response to activin remains to be elucidated.