芒果苷通过FoxO1/PDK4减轻肾缺血再灌注期间氧化应激损伤OA

Objective To explore the protective effects and mechanism of mangiferin(MG)against renal ischemia-reperfusion(I/R)injury in mice.Methods ①C57BL/6 mice were randomly divided into sham operation group,I/R group,I/R+high-dose MG(20 mg/kg)group,and I/R+low-dose MG(5 mg/kg)group.The mice in I/R+high-dose MG group and I/R+low-dose MG group were pretreated with the corresponding doses of MG for three days before renal ischemia-reperfusion.The mice were euthanized 24 h after reperfusion,and serum and renal tissue samples were collected.The levels of serum creatinine,blood urea nitrogen,and malondialdehyde(MDA)were measured.HE staining was used to observe the histopathological changes of renal tissue.Immunohistochemistry and Western blot were employed to detect the expression of pyruvate dehydrogenase kinase 4(PDK4).②Renal tubular epithelial cells(HK-2)were divided into control group,hypoxia/reoxygenation(H/R)group,H/R+high-dose MG(10 μmol/L)group,and H/R+low-dose MG(5 μmol/L)group.The cells in H/R+MG groups were pre-treated with the corresponding doses of MG for 1 h before H/R.Cell viability was assessed using CCK-8 assay,and the intracellular reactive oxygen species(ROS)content was measured using the DCFH-DA fluorescent probe.③HK-2 cells were divided into H/R+empty vector group,H/R+empty vector+MG group,and H/R+PDK4 overexpression+MG group.The intracellular ROS level was detected using the DCFH-DA fluorescent probe.Additionally,the cells were divided into H/R group,H/R+MG group,H/R+MG+leptomycin B(LMB)group,and H/R+MG+importazole(Imp)group.Immunofluorescence and Western blot were used to detect the subcellular localization of Forkhead box O1(FoxO1)and the effects of the nuclear export inhibitor LMB and import inhibitor Imp on its localization.Results ①Compared with sham operation group,the mice in I/R group exhibited signifi-cantly elevated levels of serum creatinine,blood urea nitrogen,MDA,and PDK4 protein expression(P<0.05);the renal tubules showed extensive loss of brush borders,severe tubular dilation,protein casts,and tissue necrosis.Compared with I/R group,the mice in I/R+MG groups demonstrated significantly reduced levels of serum creatinine,blood urea nitrogen,MDA,and PDK4 protein expres-sion(P<0.05);the renal tubular epithelial cells were neatly arranged,and tissue necrosis was diminished.Compared with I/R+MG low-dose group,the mice in I/R+MG high-dose group showed a significantly lower expression of PDK4 protein(P<0.05).②Compared with control group,the cell viability decreased in H/R group(P<0.05),and the expressions of PDK4 and nuclear FoxO1 protein were significantly elevated(P<0.05).Compared with H/R group,the cell viability increased while the levels of PDK4 and nuclear FoxO1 protein expression significantly reduced in H/R+MG groups(P<0.05).Compared with H/R+MG low-dose group,the cell viability significantly increased while the levels of PDK4 and nuclear FoxO1 protein expression significantly reduced in H/R+MG high-dose group(P<0.05).③Compared with H/R+empty vector+MG group,the cells in H/R+PDK4 overexpression+MG group showed signifi-cantly higher ROS levels(P<0.05).Compared with H/R+MG group,the levels of FoxO1 and PDK4 protein expression,and ROS produc-tion significantly increased in H/R+MG+LMB group(both P<0.05);the above indexes showed a converse trend in H/R+MG+Imp group(both P<0.05).Conclusion Mangiferin can alleviate renal I/R injury-induced oxidative stress injury by inhibiting the nuclear translo-cation of FoxO1 and down-regulating PDK4 expression.