Protective Effects Of Genistein On The Injury Induced By Hydrogen Peroxide In Vascular Endothelial Cells

Objective:The injury of vascular endothelial cells and oxidative stress are the basic lesions of cardiovascular diseases.The oxidative stress injury model was established by the effect of hydrogen peroxide(H₂O₂)on human umbilical vein endothelial cells(HUVEC).Genistein was used as an antioxidant to compare the cell viability,intracellular reactive oxygen species,apoptosis rate,the activities of intracellular enzymes such as glutathione peroxidase(GPx),superoxide dismutase(SOD),and glutathione peroxidase(GSH),as well as the expression of protein NRF2,HO-1,SOD1,BAX,BCL-2 AND CASPASE-3 to explore the regulatory effect of genistein on cell oxidative stress injury and apoptosis,so as to lay a foundation for the study of prevention of oxidative stress injury and provide an experimental basis for the clinical application of genistein.Method:After the preliminary experiments,the cell viability was determined by CCK-8,and10?mol/L GEN pretreatment for 1 h and 130?mol/L H₂O₂ treatment for 1 h were determined as the conditions for the subsequent experiments.The cells were divided into four groups,namely group control,group GEN,group GEN+H₂O₂ and group H₂O₂.Flow cytometry was used to determine the intracellular reactive oxygen species(ROS)levels and apoptosis rate in the different groups.Total glutathione peroxidase(GPx)activity,superoxide dismutase(SOD)activity and glutathione(GSH)content were measured using enzyme activity assay kits.The expression of m RNA Nrf2,Ho-1,Sod1,Bax,Bcl-2,Caspase-3 were determined by q PCR and the expression of the above proteins was measured by Western Blot.Graph Pad Prism 8.0 was applied for statistical analysis,and comparisons between multiple groups were made by ANOVA,corrected by Bonferroni method.Two independent samples t-test was used for comparison between two groups,and differences were considered statistically significant at P<0.05.Result:Compared with the H₂O₂ alone treatment group,the intracellular ROS in the combined treatment group decreased significantly(P<0.05);the early and total apoptosis rate decreased significantly(P<0.05);The activities of GPx,SOD,and GSH were increased(P<0.05).The results of q PCR showed that compared with the H₂O₂alone treatment group,the m RNA expression of Nrf2,Ho-1 and Sod1 increased(P<0.01),the expression of Bax/Bcl-2 decreased(P<0.01)and the expression of Caspase-3 decreased(P<0.001).Western blot showed that compared with the H₂O₂ alone treatment group,the protein expression of NRF2,HO-1 and SOD1 increased(P<0.05),and the ratio of BAX/BCL-2 decreased(P<0.01)and the expression of CASPASE-3decreased(P<0.001).Conclusion:1.GEN ameliorates the decrease in cell viability caused by H₂O₂ on HUVEC.2.GEN attenuated H₂O₂-induced intracellular ROS accumulation in HUVEC.3.GEN can reduce the increase of early and total apoptosis rate of HUVEC caused by H₂O₂ treatment.4.GEN could increase the decrease of antioxidant enzyme expression in HUVEC caused by H₂O₂.5.GEN may protect HUVEC from vascular endothelial injury caused by H₂O₂ through regulating the NRF2/HO-1 signaling pathway.