Effects of astragaloside Ⅳ on oxidative damage of vascular endothelial cells by regulating Nrf2/HO-1 signaling pathway
Objective To explore the effects of astragaloside Ⅳ(AST-Ⅳ)on vascular endothelial oxidative damage by modulating the nuclear factor-erythroid 2-related factor 2/heme oxygenase-1(Nrf2/HO-1)signaling pathway,based on cell experiments.Methods An endothelial cell oxidative damage model was established by stimulating vascular endothelial cells with 1-palmitoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine(POVPC)for 24 hours.The cells were randomly divided into model group(35 μmol/L POVPC),low-dose AST-Ⅳ(AST-L)group(35 μmol/L POVPC+50 μmol/L AST-Ⅳ),high-dose AST-Ⅳ(AST-H)group(35 μmol/L POVPC+100 μmol/L AST-Ⅳ),and high-dose AST-Ⅳ+ML385(Nrf2 inhibitor)(AST-H+ML385)group(35 μmol/L POVPC+100 μmol/L AST-Ⅳ+5 μmol/L ML385).Untreated normal cells were used as the control group.Rat aortic vascular endothelium cells(AVECs)were identified by immunofluorescence.The proliferation of AST-Ⅳ cells was tested by CCK-8 assay.Transwell chambers were utilized to evaluate AVEC migration,while Matrigel matrix gel was employed to assess angiogenic function of the cells.The cytoskeletal structure was determined by phalloidin staining,reactive oxygen species(ROS)levels were measured by the DCFH-DA fluorescence probe,and superoxide dismutase(SOD)levels in cell culture supernatants were measured by kit method.The mRNA and protein expressions of Nrf2 and HO-1 were determined by RT-qPCR and Western blot.Results The expression of extracted cell surface marker von willebrand factor(vWF)was determined by immunofluorescence and was identified as AVEC.Compared with control group,the cell viability,migration ability,and angiogenic function in Model group decreased significantly(P＜0.01),with obvious cytoskeleton disruption,a significant increase in intracellular ROS level(P＜0.01),a significant decrease in SOD content in the cell culture supernatant(P＜0.01),and downregulated mRNA and protein expression levels of Nrf2 and HO-1 in cells(P＜0.01 or P＜0.05).Compared with model group,AST-L group and AST-H group showed significantly increased cell viability,migration ability,and angiogenic function(P＜0.01),with better repair of cytoskeleton disruption,a significant decrease in intracellular ROS level(P＜0.01),a significant increase in SOD content in the cell culture supernatant(P＜0.01),and significantly upregulated mRNA and protein expression levels of Nrf2 and HO-1 in cells(P＜0.01),with more pronounced effects in AST-H group(P＜0.01).Compared with AST-H group,AST-H+ML385 group showed significantly decreased migration ability and angiogenic function(P＜0.01),increased cytoskeleton disruption,a significant increase in intracellular ROS level(P＜0.01),a significant decrease in SOD content in the cell culture supernatant(P＜0.01),and significantly downregulated mRNA and protein expression levels of Nrf2 and HO-1 in cells(P＜0.01).Conclusion AST has protective effects on vascular endothelial oxidative damage,with a certain dose dependence,and its mechanism may involve the activation of Nrf2/HO-1 signaling pathway.
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