Impact of IRS-2 on the regulation of hepatocyte pyroptosis via the PI3K/AKT pathway
Objective To explore the role and molecular mechanism of insulin receptor substrate(IRS)-2 in hydrogen peroxide(H2O2)-induced hepatocyte pyrotosis.Methods HepG2 and L02 cells were stimulated with H2O2,and the expressions of IRS-2 and pyroptosis-related proteins were assessed by Western blot analysis.IRS-2 siRNA was synthesized and employed to suppress the expression of IRS-2 gene in both HepG2 and L02 cell lines.The expression levels of IRS-2 and pyroptosis-related proteins were subsequently evaluated using Western blot analysis.Cell viability was determined using the CCK-8 assay,while changes in mitochondrial membrane potential were analyzed via flow cytometry.Cell morphology,mitochondria structure,and pyroptosomes were visualized under an electron microscope,with mitochondria morphology and quantity observed using Mito-Track Green staining.HepG2 and L02 cells were treated with IRS-2 siRNA alone or in combination with a PI3K/AKT pathway agonist to assess the expression levels of PI3K/AKT pathway proteins and pyroptosis-related proteins,Data analysis was conducted using independent sample t tests or one-way analysis of variance(ANOVA)where appropriate.Results In comparison to the control group,exposure to H2O2 led to decreased viability of hepatocytes,downregulated expression of IRS-2 protein,and increased expression of pyroptosis-related proteins.Reduced expression of IRS-2 resulted in mitochondrial dysfunction,disrupted hepatocyte morphology,increased pyroptosome numbers,and up-regulate expression of pyroptosis-related proteins.Additionally,the ratio of P-PI3K/PI3K and P-AKT/AKT was decreased.Activation of the PI3K/AKT pathway reversed the expression of pyroptosis-related proteins induced by IRS-2 downregulation.Conclusion Stimulation with H2O2 can decrease the expression of IRS-2 protein and induce pyroptosis in hepatocytes.Inhibiting IRS-2 expression may induce mitochondrial dysfunction by reducing PI3K/AKT pathway activation,ultimately leading to hepatocyte pyroptosis.
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