GPX4 Promoter Hypermethylation Induced by Ischemia/Reperfusion Injury Regulates Hepatocytic Ferroptosis

Background and Aims:Glutathione peroxidase 4(GPX4)is a key factor in ferroptosis,which is involved in ischemia-rep-erfusion injury.However,little is known about its role in he-patic ischemia-reperfusion injury(HIRI).This study aimed to investigate the role of GPX4 methylation in ferroptosis during HIRI.Methods:For the in vitro experiments,an oxygen and glucose deprivation cell model was established.For the in vivo experiments,an ischemia-reperfusion model was created by subjecting mice to simulated HIRI.Ferroptosis occurrence,GPX4 promoter methylation,and global methylation levels were then assessed.Results:Ferroptosis was observed in oxygen and glucose deprivation,characterized by a signifi-cant decrease in cellular viability(P<0.05),an increase in lipid peroxidation(P<0.01),iron overload(P<0.05),and down-regulation of GPX4(P<0.05).This ferroptosis was ex-acerbated by GPX4 knockdown(P<0.01)and mitigated by exogenous glutathione(P<0.01).Similarly,ferroptosis was evident in mice subjected to HIRI,with a down-regulation of GPX4 mRNA and protein expression(all P<0.01),and an upregulation of acyl-CoA synthetase long-chain family mem-ber 4 mRNA and protein(all P<0.01),as well as prostaglan-din-endoperoxide synthase 2 mRNA and protein expression(all P<0.05).Methylation levels increased,evidenced by upregulation of DNA methylation transferase expression(P<0.05)and down-regulation of Ten-eleven translocation fam-ily demethylases(P<0.01),along with an upregulation of GPX4 promoter methylation.Conclusions:Ferroptosis may be the primary mode of cell death in hepatocytes following ischemia-reperfusion injury.The methylation of the GPX4 promoter and elevated levels of global hepatic methylation are involved in the regulation of ferroptosis.