Studies have revealed that ferroptosis is non apoptotic
Studies have revealed that ferroptosis is non-apoptotic and peroxidation-driven form of cell death that requires abundant and accessible cellular Fe2+, and thus, the balance of Fe2+ metabolism is of great significance in regulating the process of ferroptosis (Cao and Dixon, 2016; Xie et al., 2016; Yang and Stockwell, 2016). In this study, we observed that the levels of Fe2+ were increased remarkably by arsenite in both whole cell and mitochondria. IREB2 is the master regulator of Fe2+ homeostasis (Kühn, 2015), we further found that P7C3 of IREB2 was enhanced by low level of arsenite, but sharply reduced by arsenite at the doses of 5.0 and 50 mg/L. These results indicate that, at relative low concentration of arsenite, the overload of Fe2+ may adaptably cause the overexpression of IRBE2, while the level of IREB2 can be sharply depleted with increasing doses of arsenite. Given ferroptosis can be driven by loss of activity of the lipid repair enzyme GPX4 and subsequent accumulation of lipid-based ROS and MDA (Yang et al., 2014), we further tested the level of GPX4 in the tissue of cerebral cortex. Our result showed that both the mRNA and protein expression of GPX4 as well as the contents of GSH were all decreased by chronic arsenite exposure, suggesting that arsenite treatment may induce ferroptoisis via GSH depletion by inactivation of GPX4. In addition, the intracellular GSH level can be reduced by ferroptosis inducer, such as erastin, through directly inhibiting cystine/glutamate antiporter system Xc− activity (Dixon et al., 2014). Moreover, disruption of system Xc−-mediated cystine uptake is sufficient to induce ferroptosis (Dixon et al., 2014). Thus, we determined the levels of SLC7A11 in the tissues. We observed that the mRNA and protein expression SCL7A11 were also reduced by different concentrations of arsenite, indicating that arsenite may disrupt the system Xc− to trigger ferrroptosis. Increasing evidence has demonstrated that VDAC3 has a positive role in ferroptosis (Do Van et al., 2016; Wang et al., 2016). Cells with more VDAC3 protein are more sensitive to the ferroptosis inducer (Xie et al., 2016). The elevated mRNA and protein expression of VDAC3 observed in arsenite-treated mice indicate that oxidative mitochondrial metabolism and limit aerobic glycolysis occur through disrupting the interaction between VDAC3 and tubulin within the cells (Xie et al., 2016). To further confirm the role of ferroptosis in arsenite-induced neurotoxic effects, we used the PC-12 cells as the in vitro model. Our data revealed that, the cytotoxicity and the production of mitochondrial ROS could be attenuated by iron chelators, DFO. These findings together suggest arsenite is capable to trigger ferroptotic cell death.
Conflicts of interests
Acknowledgements This research was supported by National Science Foundation of China (Grant Number: 81602820), Foundation and Frontier Research Program of Chongqing Municipal Science and Technology Commission (Grant Number: cstc2016jcyjA0223 and cstc2016jcyjA0435), Science and Technology Research Program of Chongqing Education Commission (Grant Number: KJ1600204), China Postdoctoral Science Foundation funded project (Grant Number: 2017M612925), and Postdoctoral Research Project of Chongqing Municipality Grant Number (XM2017003).