INHIBITORY EFFECTS OF SUPEROXIDE-DISMUTASE AND CYCLIC GUANOSINE 3',5'-MONOPHOSPHATE ON ESTROGEN PRODUCTION IN CULTURED RAT GRANULOSA-CELLS

Superoxide dismutases (SOD) modulate oxygen free radical metabolism an d influence second messenger signaling in a variety of cell types. We have investigated the influence and possible mechanisms of action of S OD on aromatase activity in cultured rat granulosa cells. Although tre atment of granulosa cells with FSH (0.3-30 ng/ml) resulted in a dose-d ependent stimulation of estrogen levels, cotreatment of cells with SOD (10(-6) M) significantly attenuated estrogen production at the highes t doses of FSH. The effects of SOD were dose dependent between 10(-7)- 10(-5) M, with increasing amounts of SOD causing decreasing concentrat ions of estrogen. Cotreatment of cells with catalase (1500 U/ml) faile d to prevent the inhibitory influence of SOD on estrogen production, i ndicating that the effects of SOD were not due to accumulation of hydr ogen peroxide. Although incubation with either forskolin or (Bu)(2)cAM P alone stimulated estrogen production from granulosa cells, cotreatme nt with SOD significantly attenuated estrogen levels, indicating that SOD can inhibit aromatase activity at one or more post-FSH receptor si tes. Treatment of cells with SOD, FSH, or forskolin resulted in small, but significant, increases in cGMP concentrations. In contrast, cotre atment of cells with FSK plus SOD as well as forskolin plus SOD had a marked synergistic effect on cGMP content, increasing cGMP levels over 100-fold. Incubation of granulosa cells with (Bu)(2)cGMP (2 mM) signi ficantly decreased FSH-induced estrogen levels in a dose-dependent man ner (0.25-2 mM). In addition, (Bu)(2)cGMP attenuated both forskolin- a nd (Bu)(2)cAMP-induced estrogen production. In contrast to the effects of (Bu)(2)cGMP and SOD on estradiol levels, these agents had no signi ficant effect on progesterone production by cultured granulosa cells. These results demonstrate attenuated induction of aromatase activity b y FSH in cultured rat granulosa cells cotreated with SOD, suggesting a potential modulatory role of this antioxidant on granulosa cell funct ions. The findings that SOD and activators of the cAMP-dependent signa ling pathway synergistically increase the levels of the second messeng er cGMP and that (Bu)(2)cGMP treatment attenuates FSH-, forskolin-, an d cAMP-induced aromatase activity suggest a potential mechanism of SOD action and demonstrate the antagonistic action of cGMP on cAMP-mediat ed estrogen production.