Effect And Mechanism Of Oxidative Damage To Human Umbilical Vein Endothelial Cells Caused By Di(2-ethylhexyl) Phthalate And The Repairing Function Of Resveratrol

Objective: To establish an in vitro injury model of human umbilical vein endothelial cells(HUVEC)induced by di-2-ethylhexyl phthalate(DEHP),and to verify the in vitro repair effect of resveratrol on this injury,and to investigate the mechanism of DEHP on HUVEC functional injury and resveratrol on its repair.Methods: The damage concentration of DEHP and the repair concentration of resveratrol were screened by CCK-8 assay;HUVEC were divided into control,injury and repair groups;the changes of proliferation and migration ability of HUVEC cells in different treatment groups were observed by CCK8 assay,Transwell assay and wound healing assay;the in vitro tube formation ability of HUVEC in different treatment groups was detected by in vitro tube formation assay;the apoptosis rate of HUVEC was measured by cell flow assay;expression of m RNA of Bax and Bcl-2 in different treatment groups was detected by real-time fluorescence quantitative PCR;the expression of proteins Bax and Bcl-2 in HUVEC cells was observed by western blotting;the different expressions of malondialdehyde and superoxide dismutase(SOD)in HUVEC in different treatment groups were detected by chemical spectrophotometric colorimetric assay;the different expression of reactive oxygen species(ROS)and mitochondrial membrane potential in HUVEC were detected by cellular immunofluorescence;.Results: The cell proliferation capacity of HUVEC was significantly reduced after treated with 80 μmol/L DEHP for 24 h(P<0.01).Resveratrol was not significantly toxic to cells at 40 μmol/L and below;the multiple cell capacity of cells in the injury group was significantly decreased compared with the control group(P<0.01),and significantly increased in the repair group compared with the injury group(P<0.01);the apoptosis rate of cells in the injury group was significantly increased(P<0.01)compared with the control group,and the apoptosis rate of cells in the repair group was significantly decreased compared with the injury group(P<0.01).There was no significant difference in the expression of m RNA of Bax and Bcl-2 between the different treatment groups;the protein expression level of cells in the injury group showed an increase in Bax expression,a decrease in Bcl-2 expression,and a decrease in Bcl-2/Bax ratio compared with the control group(all P<0.05),and the protein expression Bax expression decreased,Bcl-2 expression increased and Bcl-2/Bax ratio increased in the repair group compared to the injury group(all P<0.05);malondialdehyde content and SOD activity increased significantly in the injury group compared to the control group(both P<0.01)and decreased significantly in the repair group compared to the injury group(both P<0.05);ROS level increased significantly in the injury group compared to the control group(P<0.01)and decreased in the repair group compared with the injury group(P<0.01);the mitochondrial membrane potential in the injury group decreased significantly compared with the control group(P<0.01)and increased significantly in the repair group compared with the injury group(P<0.01).Conclusion: DEHP was able to inhibit multiple abilities of HUVEC and induce apoptosis through oxidative stress pathway,while resveratrol could repair this damage by alleviating oxidative stress.