Inhibitory Effect Of Genistein On Tert-butyl Hydroperoxide Induced Senescence Of H9c2 Cells And Its Mechanism

Objective:To investigate the effect of Genistein on the antioxidation of senescent H9c2 cells induced by tert-butyl hydroperoxide and its mechanism.Methods:H9c2 cells of the 5～13 passage were.used in the experiment and treated with different concentrations of t-BHP.The optimal concentration of t-BHP was screened by MTT and beta-galactosidase staining to establish the aging model of H9c2 cell.The cytotoxicity of Gen was detected by CCK-8 method.H9c2 cells were divided into normal control group and model group and Gen low,medium and high dose groups.MTT assay was used to detect the survival rate of H9c2 cells.β-galactosidase staining was used to observe the degree of cell senescence.The level of T-AOC,the activities of SOD,GSH-Px,CAT and the content of MDA in H9c2 cells were measured by colorimetric method.The expression of P53,P21,Keapl,Nrf2,HO-1 and nuclear Nrf2 protein were detected by Western blotting.Results:1.200μmol/L was used as the optimal concentration of t-BHP for the establishment of H9c2 cell senescence model.2.The results showed that Gen had no significant effect on the survival rate of H9c2 cells in the concentration range of 0～5μmol/L.H9c2 cells pretreated by Gen were stained with β-galactosidase and the number of positive cells in the model group increased significantly compared with that in the normal control group(P<0.01).Compared with the model group,the number of positive cells in each dose group of Gen reduced significantly(P<0.01).3.Compared with the normal control group,the T-AOC level and the activities of SOD,GSH-Px and CAT in the model group decreased(P<0.01),and the content of MDA increased(P<0.01);compared with the model group,Gen can significantly increase the level of T-AOC and the activities of SOD,GSH-Px and CAT in H9c2 cells(P<0.05 or P<0.01),and reduce the content of MDA(P<0.01).Western Blotting results showed that compared with the normal control group,the protein expression levels of P53 and P21 in the model group were significantly up-regulated(P<0.01);compared with the model group,Gen could significantly down-regulate the protein expression levels of P53 and P21(P<0.01)).Compared with the normal control group,the expression level of Keapl protein in the model group was significantly up-regulated,while the expression levels of Nrf2,HO-1 protein and Nrf2 protein in the nucleus were down-regulated(P<0.01);compared with the model group,Gen could significantly down-regulate the expression of Keapl protein Level,up-regulate the expression levels of Nrf2,HO-1 protein and Nrf2 protein in the nucleus(P<0.01).Conclusion:Gen inhibits the senescence of H9c2 cells by increasing the antioxidation of H9c2 cells.The mechanism may be related to the down regulation of p53,p21,Keap 1 protein and the up regulation of Nrf2 and HO-1 protein expression.