Effect of Human Menopausal Gonadotropin and Follicle Regulatory Protein(s) on 3β-Hydroxysteroid Dehydrogenase in Human Granulosa Cells*

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Abstract
Recently, a protein fraction [follicle regulatory protein (FRP)] which inhibits FSH-induced granulosa cell aromatase activity was isolated from both human and porcine follicular fluid. In this study, the actions of FRP on 3β-hydroxysteroid dehydrogenase (33β-HSDH) activity were examined using granulosa cells obtained from hyperstimulated patients undergoing oocyte aspiration for in vitro fertilization. Granulosa cells were cultured with 0,167, or 500 μ g/rril FRP with or without human menopausal gonadotropin (hMG; 10 mlU/ml). After 48 h, the medium (S) was removed and stored. Cells then were mechanically lysed and centrifuged at 10,000 × g. The supernatant was further centrifuged (100,000 × g) to obtain a microsomal fraction (M) and cytosol (C). The M fraction was resuspended in medium 199 with 10-6 M pregnenolone plus 5 μM NAD+ and incubated for 2 h to determine 3β-ol dehydrogenase activity. The S, C, and M fractions were all assayed for progesterone (P) by RIA. hMG markedly increased P concentrations in the S and C fractions. The M fraction demonstrated a hMG-dependent enhancement in 3β-HSDH activity. However, the hMG-associated S, C, and M P levels were decreased in granulosa cells coincubated with FRP. In conclusion, ovarian steroidogenesis may be dependent on the integrated interactions of both gonadotropins and local nonsteroidal paracrine/autocrine modulators of granulosa cell function.

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