Based on the Nrf2-ARE Signaling Pathway,the Molecular Mechanism of Oxidative Stress Damage Induced by DJ-1 Against Cardiomyocyte Hypoxia/Reoxygenation was Investigated

QIAN Ruping; MA Jianjun; GU Weimin

doi:10.13919/j.issn.2095-6274.J202403090

Abstract：Objective To investigate the molecular mechanism of oxidative stress damage induced by hy-poxia/reoxygenation of anti-cardiomyocytes by DJ-1 based on the Nrf2-ARE signaling pathway.Methods H9c2 cardiomyocytes were transfected with DJ-1 siRNA,NC siRNA,pFlag-DJ-1,and pFlag,and the expressions of DJ-1 and antioxidant enzymes(MnSOD,CAT and GPx)were detected by Western blot.The H/R model in cardiomyocytes was established,and the cell viability,LDH activity,MDA levels,and ROS content in each group were accessed.The DJ-1's effect on Nrf2-Keapl dissociation,the Nrf2 nuclear translocation,the binding of Nrf2 to MnSOD,CAT,and GPx-ARE,and the influence of transcriptional activity of Nrf2 were observed,in order to observe the effect of DJ-1 on the Nrf2 signaling pathway.H9c2 cells were transfected with pFlag-DJ-1/pFlag and then transfected with Nrf2 siRNA/NC siRNA,and the changes in the expression of MnSOD,CAT,and GPx were detected by Western blot;H/R damage model was established to detect the sur-vival rate of cardiomyocytes,LDH activity,and MDA and ROS contents in each group,and observe the effect of inhibition of Nrf2 signaling pathway on antioxidant stress in DJ-1.Results The expression levels of DJ-1,MnSOD,CAT,and GPx were significantly increased after transfection with pFlag-DJ-1 with H9c2.The survival rate of H/R-damaged cardiomyocytes was significantly im-proved after H9c2 transfection with pFlag-DJ-1,but the LDH activity,and ROS and MDA production were signifi-cantly reduced,while the above effects were reversed when transfected with DJ-1 siRNA.After transfection of H9c2 cells with pFLAG-DJ-1,the dissociation of Nrf2-Keapl,the entry of Nrf2 into the nucleus,the binding of Nrf2 to ARE in the nucleus were improved,while the above effects were reversed when DJ-1 is silenced..When the intracel-lular Nrf2 signaling pathway was inhibited,the effect of DJ-1 overexpression which up-regulated the expression of MnSOD,CAT,and GPX was reversed.DJ-1 overexpression significantly improved the survival rate of H/R-damaged cells,inhibited LDH activity,and the production of ROS and MDA,but these effects were significantly inhibited by Nrf2 silencing.Conclusion DJ-1 may induce the expression of antioxidant enzymes by activating the Nrf2-ARE signaling pathway,thereby exerting anti-cardiomyocyte H/R-induced oxidative stress damage.