Autophagy Contributes to Oxidative Stress-Induced Apoptosis in Porcine Granulosa Cells.

Oxidative stress-induced granulosa cell (GC) death is a major cause of follicular atresia. As the major types of programmed cell death, autophagy and apoptosis have been observed in response to HO-mediated oxidative stress and have been demonstrated to be responsible for porcine GC death. To date, however, the cellular reactions linking autophagy to the apoptosis of porcine GC under oxidative stress are still poorly understood. Porcine GC were treated with HO, and autophagic flux was examined by western blotting. Cell viability and cell death assays were performed after cotreatment of porcine GC with autophagy activator (rapamycin) or inhibitor (3-methyladenine, 3-MA) together with HO. We revealed that short exposure (1-3 h) of porcine GC to HO dramatically increased autophagic flux (1.8- to 2.5-fold over that in the control), whereas 6-12 h prolonged treatment decreased autophagy but elevated the caspase-3 activity and GC apoptotic rate. Furthermore, we showed that pretreatment with rapamycin exacerbated HO-mediated cytotoxicity and caspase-3 activation but that 3-MA or siRNAs specific for Beclin 1 and Atg7 genes ameliorated HO-mediated GC apoptosis. Together, our results indicate that autophagy plays a pivotal role in HO-mediated porcine GC apoptosis. Importantly, we show that the early induction of autophagic flux contributes to oxidative stress-induced apoptosis in porcine GC. The results also suggest that regulating the autophagy response in porcine GC under oxidative stress might be a new strategy for abnormal follicular atresia.