Blockinghepatocyte necrotic apoptosis to aggravate hepatic ischemia-reperfusion injury by inducing autophagy inhibition / 中华器官移植杂志

ABSTRACT

Objective To study the function and mechanism of receptor-interacting protein kinase 3 (RIPK3)-mediated necroptosis in liver ischemia-reperfusion injury (IRI) in mice.Methods Sixty mice were randomly divided into four groups using Stata statistical softwareWild-type (WT)-sham group,WT-IRI group,HKO (HKORIPK3 liver-specific knockout)-sham group and HKO-IRI group.Sham operation was used as a control in which only the hepatic portal blood vessels were freed after laparotomy,and blood flow was not blocked.In the WT-IRI group and the HKO-IRI group,the hepatic portal vein was freed,and the blood supply of left hepatic lobe and the mid-hepatic lobe wer blocked for 90 min,then the blood vessels were opened for 6 h.Blood and liver tissue samples of each group of mice were taken to detect liver function.Inflammatory infiltration and liver injury were detected by immunohistochemistry and hematoxylin and eosin (HE) staining,and autophagyassociated protein LC3-Ⅱ and P62 were detected by Western blotting.The primary hepatocytes of WT mice and HKO mice were extracted and divided into control group and hypoxia-reoxygenation group (HIR group).After attachment of primary hepatocytes,the HIR group was given hypoxia for 6 h and reoxygenated for 4 h.The supernatant was taken for detecting ALT and AST,and the cell extract protein was used to detect LC3-Ⅱ and P62.Results As compared with the control groups,the liver functions of the IRI groups were significantly impaired,and as compared with the WT-IRI group,the liver damage was significantly aggravated in the HKI-IRI group (P ＜ 0.05),and the LC3-Ⅱ protein content was significantly decreased and the P62 protein content was increased.Similarly,after hepatocytes were were given hypoxia and reoxygenated,HKO-derived hepatocytes were more severely damaged than WT-derived hepatocytes.Conclusions Blocking RIPK3-mediated necroptosis of hepatocytes could induce autophagy inhibition,which aggravates hepatic ischemia-reperfusion injury.