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Objective Prepuberty is a critical period for Sertoli cell maturation and functional differentiation,this study aims to evaluate Sertoli cell oxidative state and possible mechanism after genistein(GEN)and Mono(2-ethylhexyl)phthalate(MEHP)single/simultaneous exposure.Methods Sertoli cells were cultured with a medium containing GEN at 10μM(G group),MEHP at 1000μM,100μM,10μM,1μM(M1000,M100,M10,M1 group),GEN and MEHP mixer(G+M1000 group,G+M100 group,G+M10,G+M1 group)as well as a control group.After treated for 48 hours,reactive oxygen species(ROS)generation,inhibition rate of superoxide dismutase(SOD inhibition rate),total antioxidant capacity(T-AOC),glutathione peroxidase(GSH-Px)and malondialdehyde(MDA)in the cell culture medium,cell proliferation,Nrf2-ARE signaling pathway and downstream antioxidant genes heme oxygenase-1(HO-1),superoxide dismutase(SOD),quinone oxidoreductase(NQO1)expression were studied.Results(1)SOD inhibition of Group G+M 1000,G+M 100 significantly increased compared with Group G(P <0.05),which were significantly lower than MEHP exposure alone(P <0.05).T-AOC of Group G+M1000,G+M100 significantly decreased than Group G(P <0.05),which were significantly higher than MEHP exposure alone(P <0.05).T-AOC of Group G+M10,G+Mlshowed no significant differences compared with Group G,but significantly increased than MEHP single exposure(P <0.05).MDA level of Group G+M1000,G+M100 were significantly decreased than M1000,M100 group(P<0.05)while Group G+M10,G+M1 showed no significant difference than GEN and MEHP single exposure(P >0.05).We also found no no significant statistical difference of GSH-PX between the GEN and MEHP co-exposure groups and groups exposed to GEN and MEHP alone(P > 0.05).(2)Con group,G group showed lower ROS levels,ROS levels of Group G+M1000,G+M100 significantly increased than GEN exposure alone(P <0.01),which were significantly lower than MEHP exposure alone(P <0.05).Group G+M10,G+M1 showed no statistical significance than GEN exposure alone(P > 0.05)but showed significant decrease than MEHP exposure alone(P <0.05).(3)Group Con,G showed a lower incidence of apoptosis,MEHP exposue induced dose-dependent gradient decreased apoptosis rate.Group G+M1000 apoptotic rate than GEN exposure alone significantly increased(P <0.05),but lower than MEHP exposure alone(P <0.05).Group G+M100,G+M10,G+M1 apoptotic rate compared to MEHP exposure alone significantly decreased(P <0.05),which showed no statistically significant difference compared to GEN exposure alone(P > 0.05).(4)Nrf2 expression of Group G+M 1000,G+M100,G+M10 group compared with GEN and MEHP exposure alone significantly increased(P <0.05),Group G+M1 significantly increased compared with GEN exposure alone(P <0.05),which showed no significant difference than MEHP exposure alone(P > 0.05).HO-1 expression of Group G+M 1000,G+M100,G+M 10 compared with GEN and MEHP exposure alone significantly increased(P <0.05),Group G+M1 compared to GEN exposure alone increased significantly(P <0.05),although higher than MEHP exposure alone but showed no significant difference(P > 0.05).SOD expression in Group G+M1000,G+M100,G+M10,G+M1 significantly increased compared with MEHP and GEN exposure alone(P <0.05).NQO 1 expression in Group G+M 1000 compared with GEN exposure alone significantly increased(P <0.05)but showed no significant difference compared with MEHP exposure alone(P > 0.05).NQO1 expression in Group G+M100,G+M10,G+M1 significantly increased compared with MEHP and GEN exposure alone(P <0.05).Conclusion Genistein could attentuates MEHP-induced Oxidative Stress in Sertoli Cell via Activating Nrf2-ARE Pathway,which regulates downstream antioxidant genes and protein expression to reduce MEHP induced Sertoli cell damage.