Bone morphogenetic protein inhibits ovarian androgen production

Bone morphogenetic proteins (BMPs), members of the transforming growth fact or beta superfamily, were recently shown to be expressed and to regulate st eroidogenesis in rat ovarian tissue. The purpose of this study was to inves tigate the effect of BMP-I on androgen production in a human ovarian theca- like tumor (HOTT) cell culture model. We have previously demonstrated the u sefulness of these cells as a model for human thecal cells. HOTT cells resp ond to protein kinase A agonists by increased production of androstenedione and with an induction of steroid-metabolizing enzymes. In this investigati on, HOTT cells were treated with forskolin or dibutyryl cyclic AMP (dbcAMP) in the presence or absence of various concentrations of BMP-I. The accumul ation of androstenedione, progesterone, and 17 alpha-hydroxyprogesterone (1 7OHP) in the incubation medium was measured by RIA. The expression of 17 al pha-hydroxylase (CYP17), 3 beta-hydroxysteroid dehydrogenase (3 beta HSD), cholesterol side-chain cleavage (CYP11A1), and steroidogenic acute regulato ry (StAR) protein was determined by protein immunoblotting analysis using s pecific rabbit polyclonal antibodies. We also examined the expression of BM P receptor subtypes in our HOTT cells using RT-PCR. In cells treated with m edium alone, steroid accumulation and steroid enzyme expression was unchang ed. In cells treated with BMP alone there was a modest decrease in androste nedione secretion. In the presence of forskolin, HOTT cell production of an drostenedione, 17OHP, and progesterone increased by approximately 4.5-, 35- , and S-fold, respectively. In contrast, BMP-4 decreased forskolin-stimulat ed HOTT cell secretion of androstenedione and 17OHP by 50% but increased pr ogesterone production S-fold above forskolin treatment alone. Forskolin tre atment led to an increase in CYP17, CYP11A1, 3 beta HSD, and StAR protein e xpression. BMP-4 markedly inhibited forskolin stimulation of CYP17 expressi on but had little effect on 3 beta HSD, CYP11A1, or StAR protein levels. Si milar results were observed with the cAMP analog dbcAMP. In addition, BMP-4 inhibited basal and forskolin stimulation of CYP17 messenger RNA expressio n as determined by RNase protection assay. Other members of the transformin g growth factor beta superfamily, including activin and inhibin, had minima l effect on androstenedione production in the absence of forskolin. In the presence of forskolin, activin inhibited androstenedione production by 80%. Activin also inhibited forskolin induction of CYP17 protein expression as determined by Western analysis. We identified the presence of messenger RNA for three BMP receptors (BMP-IA, BMP-IB, and BMP-II) in the HOTT cells mod el. In conclusion, BMP-4 inhibits HOTT cell expression of CYP17, leading to an alteration of steroidogenic pathway resulting in reduced androstenedion e accumulation and increased progesterone production. These effects of BMP- 4 seem similar to those caused by activin, another member of the transformi ng growth factor beta superfamily of proteins.