Involvement of oxidative stress in ZnO NPs-induced apoptosis and autophagy of mouse GC-1 spg cells
Dan Yang, Meijuan Zhang, Yu Gan, Siyu Yang, Jinglei Wang, Yu Mei, Jie Wei, Jiaxiang Chen
Abstract
Zinc oxide nanoparticles (ZnO NPs) have been extensively used in various industries and reported to inhibit spermatogenesis, however, ZnO NPs-induced spermatogenesis failure is yet to be fully elucidated. Herein, mouse-derived spermatogonia cell line GC-1 spg cells were treated with ZnO NPs for 24 h in the presence or absence of radical scavenger N-acetyl-L-cysteine (NAC) or autophagy inhibitor 3-methyladenine (3-MA), then cell viability was observed by MTT assay; apoptosis was observed by western blotting analysis and AnnexinV-FITC/PI assay, respectively; autophagy was detected by western blotting analysis and transmission electron microscopy, respectively; and the contents of MDA and GSH and the activities of SOD and GSH-PX were measured by oxidative stress kits. The present study showed that ZnO NPs exposure inhibited viability and induced apoptosis of mouse GC-1 spg cells. Intriguingly, ZnO NPs markedly increased the protein content of LC3-II, the ratio of LC3-II/LC3-I, and the protein levels of ATG 5 and Beclin 1 in the cells. Furthermore, transmission electron microscopy (TEM) showed that autophagic vesicles in the cytoplasm increased significantly in the ZnO NPs-treated cells, indicating that ZnO NPs could induce autophagy of the cells. Oxidative stress could be induced by ZnO NPs; moreover, inhibition of oxidative stress could alleviate the induction of apoptosis and autophagy by ZnO NPs. Inhibition of autophagy by 3-MA could rescue the inhibition of cell viability and induction of apoptosis by ZnO NPs, which indicated that autophagy might have cytotoxic effect on ZnO NPs-induced apoptosis. In summary, oxidative stress was involved in ZnO NPs-induced apoptosis and autophagy of mouse GC-1 spg cells, and autophagy might play a cytotoxic role in ZnO NPs-induced apoptosis.