The original 'two-cell mechanism' explained the endocrine regulation of follicular oestrogen synthesis and implied paracrine signalling in the follicle wall. It is now known that the CYP17 gene encoding 17-hydroxylase/C17-20-lyase activity crucial to androgen synthesis, is expressed exclusively in thecal cells. 17-Hydroxylase/C17-20-lyase activity is regulated by LH and subject to local modulation by a factor(s) emanating in FSH-stimulated granulosa cells. The FSH receptor gene is expressed exclusively in granulosa cells, where FSH acts directly to induce cytoproliferation and differentiation via cyclic AMP/protein kinase-A mediated post-receptor signalling. Granulosa cells also express androgen receptors, and theca-derived androgen has the potential to modulate locally differentiative responses to FSH. When follicles are recruited to preovulatory development by FSH, their granulosa cells develop LH receptors functionally coupled to aromatase activity and inhibin production. Thereby they simultaneously undertake LH-responsive aromatization and inhibin synthesis. Inhibin has the potential to potently enhance LH-stimulated thecal androgen synthesis. Granulosa-derived inhibin may therefore participate in a paracrine mechanism that locally amplifies androgen synthesis, and hence oestrogen formation, in the preovulatory follicle(s).
In mammalian ovaries, luteinizing hormone (LH) induces androgen biosynthesis by theca interna cells, whereas both follicle-stimulating hormone (FSH) and LH stimulate aromatase activity by granulosa cells. The joint action of these two types of cell and pituitary hormones forms the basis of a 2-cell, 2-gonadotropin hypothesis for biosynthesis of estrogen. We tested the synergism between these cell types using antisera against the estrogen precursors progesterone and testosterone. Ovaries were obtained from immature rats two days after a single injection of pregnant mare serum gonadotropin (PMSG). Granulosa cells were obtained by puncturing the follicles, and ovarian remnants were dissociated with collagenase to derive cells from the theca interstitium. Granulosa and theca interstitial cells were cultured alone or in combination for 20 h. Granulosa cells secreted negligible amounts of estrogen and testosterone, but contained high aromatase activity. Treatment with both FSH and human chorionic gonadotropin (hCG) increased production of progesterone in granulosa cells. In contrast, theca interstitial cells had negligible aromatase activity. Treatment with hCG, but not FSH, increased androgen production by theca interstitium; the hCG action was further augmented by the inclusion of exogenous progesterone. Furthermore, co-culture of gonadotropin-treated granulosa and theca interstitial cells resulted in substantial increases in estrogen production, indicating synergism between the two types of cell. The increases in estrogen production in the co-cultures were accompanied by decreases in progesterone content. To test the possibility that progesterone released by granulosa cells may serve as a substrate for production of androgen in theca cells, specific antiserum was used to adsorb progesterone present in the medium. Addition of the progesterone antibody inhibited gonadotropin-stimulated production of testosterone (39% decrease) and estrogen (64% decrease) by the combined cell cultures. The inhibitory effect on estrogen production could be reversed by the addition of progesterone (10(-6) M) or testosterone (10(-6) M) but not by the addition of a synthetic progestin, R5020. Since testosterone released from theca cells may serve as the substrate for aromatases in granulosa cells, specific testosterone antiserum was also used. Production of estrogens by the combined cell cultures was inhibited (78%) by the testosterone antiserum, but the inhibitory effect was completely reversed by exogenous testosterone or progesterone. Thus, synergistic interactions between granulosa and theca interstitial cells are important in effecting maximal estrogen biosynthesis.(ABSTRACT TRUNCATED AT 400 WORDS)