[IOX1 alleviates hepatic ischemia-reperfusion injury by inhibiting mitogen-activated protein kinase signaling...

Objective: To explore and discuss the protective effects of 5-carbonyl-8-hydroxyquinoline (IOX1) potential mechanisms in the mouse model of hepatic ischemia-reperfusion injury (IRI) and in vitro hepatocyte hypoxia-reoxygenation (H/R). Methods: In vivo and in vitro models of liver injury were established using a mouse hepatic ischemia-reperfusion model and a primary hepatocyte hypoxia-reoxygenation (H/R) model. In vivo experiments were divided into three groups: sham group, IRI group, and IRI+IOX1 group. In vitro experiments were divided into four groups: negative control group (CTRL), CTRL+IOX1 group, H/R group, and H/R+IOX1 group. Liver injury was assessed by measuring serum liver enzyme levels, hematoxylin-eosin (HE) staining, and immunohistochemistry. Apoptosis and oxidative stress were evaluated using terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay, flow cytometry, real-time quantitative PCR, and reactive oxygen species (ROS) fluorescence staining. Differentially expressed genes were screened by RNA sequencing of liver tissues, and the expression levels of differential proteins were validated by western blotting. Data were processed using GraphPad Prism 10.0. Results: Compared with the IRI group, the IRI+IOX1 group showed significantly lower levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST). At an IOX1 dose of 2 mg/kg, ALT and AST levels decreased by 87.05% and 94.49%, respectively (P<0.000 1), and hepatic histopathological damage was markedly ameliorated. Immunohistochemical staining results revealed less inflammatory cell infiltration in the IRI+IOX1 group. TUNEL assay, western blotting, and real-time quantitative PCR results demonstrated that IOX1 treatment reduced apoptosis after hepatic I/R injury. In the in vitro model, flow cytometry and western blotting results showed that IOX1 reduced apoptosis in primary hepatocytes after H/R injury, and ROS staining results indicated that IOX1 decreased ROS production following H/R injury. RNA sequencing and western blotting results revealed that IOX1 regulated the mitogen-activated protein kinase signaling pathway in hepatocytes by inhibiting AlkB homolog 5 (ALKBH5). Conclusion: IOX1 regulates the extracellular signalregulated kinase (ERK)/p38 mitogen-activated protein kinase (MAPK) signaling pathway by inhibiting ALKBH5 in hepatocytes, thereby reducing hepatocyte apoptosis and inflammatory responses and attenuating liver injury following ischemia-reperfusion.