Over-activation of cAMP signaling increases vulnerability to hydrogen peroxide via actin rearrangement, which is inhibited by prostaglandin E₂ in a mouse granulosa cell line.

Abstract

Apoptosis of granulosa cells (GC) contributes to ovarian follicular atresia, and has been implicated to depend on the oxidant status of GC within follicles. Here, we investigated the effects of cAMP and prostaglandin E₂ (PGE₂) on sensitivity to oxidative stress using 4B2 cells with cAMP-dependent, steroidogenic and differentiated properties. This cell line was isolated from mouse GC co-transfected with genes for SV40 large T antigen and Ad4BP/SF1 transcription factor (Kamei et al. 2005). Treatment of serum-starved cells with 8-BrcAMP caused 30 to 40% of the cells to become polygonal within 2 h through actin rearrangement. Interestingly, H₂O₂ treatment showed that these polygonal cells were vulnerable to oxidative stress that led to cell death, which was inhibited by pretreatment with phalloidin, an F-actin stabilizing agent. Although PGE₂ alone had no effect, cotreatment with PGE₂ and 8-Br-cAMP completely inhibited the effects of 8-Br-cAMP on cell shape change and oxidative stress vulnerability through phosphatidylinositol 3-kinase (PI3K)-dependent manner. Notably, PGE₂ and 8-Br-cAMP cooperated additively to increase progesterone secretion. These data suggest that cAMP signaling in GC may enhance oxidative stress risk through actin rearrangement, and PGE₂ may reduce such risk through activating PI3K, while cooperating with cAMP signaling in steroidogenesis.