Hydrogen Sulfide Inhibits Ferritinophagy-Mediated Ferroptosis in the Hippocampus of Rotenone-Exposed Rats

Abstract Our previous research has established that hydrogen sulfide (H 2 S) exerts an antagonistic effect against the hippocampal neurotoxicity induced by Rotenone (ROT). However, the underlying mechanisms are so far poorly understood. Substantial evidence corroborates the involvement of ferroptosis in ROT-induced neurotoxicity. To elucidate the protective mechanism of H 2 S against ROT-induced hippocampal neurotoxicity, this study explores its regulatory role in ferroptosis and its underlying mechanisms. We used Fluoro-Jade B staining to detect dead neurons. The levels of ferrous ions and glutathione (GSH) were measured by a kit. The ferroptosis-related proteins, including light-chain subunit (xCT), GSH peroxidase 4(GPX4), ferroptosis marker acyl-CoA synthetase long-chain family member 4(ACSL4), and ferritinophagy-related protein, including ferritin heavy chain 1 (FTH1), sequestosome 1 (p62), ferritinophagy markers autophagosome marker light-chain I/II (LC3I/II), and nuclear receptor coactivator 4 (NCOA4), were measured by Western blot. Our findings indicate that H 2 S reduces hippocampal neuron deaths in ROT-exposed rats. Meanwhile, H 2 S reverses the downregulations of xCT and GPX4, and the upregulations of ferrous ion and ACSL4 in the hippocampus induced by ROT. Furthermore, H 2 S reverses the upregulations of LC3I/II and NCOA4, and the downregulations of P62 and FTH1. Based on these findings, we concluded that the protective role of H 2 S against ROT-induced hippocampal neuronal death involves inhibiting ferroptosis triggered by ferritinophagy.