氧化苦参碱调节Nrf2/HO-1信号通路对HUVEC细胞损伤的改善作用

Study of oxymatrine on improving HUVEC cell damage by regulating Nrf2/HO-1 signaling pathway

【目的】探讨氧化苦参碱(oxymatrine,OMT)对人脐静脉内皮细胞(human umbilical vein endothelial cell,HUVEC)氧化应激(oxidative stress,OS)损伤的保护机制,为进一步研发预防高原缺氧药物提供理论依据。【方法】将HUVEC分为以下5组:(1)对照组:不给予任何处理。(2)模型组:细胞培养液中加入终浓度450μmol/L的H2O2培养2 h。(3)、(4)和(5)OMT预处理组:分别将10、20和30μmol/L OMT加入细胞培养基中预处理12 h,再向其加入终浓度为450μmol/L的H2O2培养2 h。观察5组细胞的代谢活力、细胞形态、乳酸脱氢酶(lactate dehydrogenase,LDH)漏出量、丙二醛(malondialdehyde,MDA)含量、超氧化物歧化酶(superoxide dismutase,SOD)及谷胱甘肽还原酶(glutathione,GSH)活性、细胞凋亡情况和B细胞淋巴瘤/白血病-2(B cell lymphoma/leukemias-2,Bcl-2)、Bcl-2相关蛋白X(bcl-2 associated x protein,Bax)、含半胱氨酸的天冬氨酸蛋白水解酶3(cysteinyl aspartate specific proteinase 3,Caspase3)、心肌组织核因子相关因子2(NF-E2-related factor 2,Nrf2)、血红素氧化酶-1(heme oxygenase-1,HO-1)蛋白表达水平。【结果】与模型组比较,OMT低、中、高预处理组细胞形态逐渐恢复,代谢活力显著升高;细胞上清液中LDH漏出量降低,细胞内SOD、GSH活性增加,MDA含量下降;细胞凋亡率降低;Bax、Caspase3蛋白水平表达下降,Bcl-2、Nrf2、HO-1蛋白水平表达升高,具有统计学意义(P<0.05)。【结论】OMT通过提高细胞内源性抗氧化酶活性、上调Nrf2/HO-1信号通路及抑制细胞线粒体凋亡途径保护OS损伤的HUVEC细胞。

【Objective】To explore the protective mechanism of oxymatrine(OMT)on oxidative stress(OS)damage of human umbilical vein endothelial cells(HUVEC),and provide a theoretical basis for further research and development of drugs to prevent altitude hypoxia.【Methods】HUVEC cells were divided into the following 5 groups:(1)control group:no treatment was given;(2)model group:cell culture medium was added with a final concentration of 450μmol/L H2O2 and cultured for 2 h;(3),(4)and(5)OMT pretreatment groups:10,20 and 30μmol/L OMT were added to medium for 12 h,and then added H2 O2 with a final concentration of 450μmol/L for 2 h.The metabolic activity,cell morphology,lactate dehydrogenase(LDH)leakage,malondialdehyde(MDA)content,superoxide dismutase(SOD)and glutathione reductase(GSH)activity,cell apoptosis and protein expression levels of B cell lymphoma/leukemias-2(Bcl-2),Bcl-2 associated x protein(Bax),cysteinyl aspartate specific proteinase 3(Caspase3),NF-E2-related factor 2(Nrf2)and hemeoxygenase-1(HO-1)were observed.【Results】Compared with the model group,the cell morphology gradually recovered in the OMT pretreatment groups,and the metabolic activity was significantly increased;the amount of LDH leakage in the cell supernatant decreased,the activity of SOD and GSH in the cells increased,andthe MDA content decreased;the rate of cell apoptosis decreased;Bax,Caspase3 protein levels decreased,Bcl-2,Nrf2,HO-1 protein levels increased.The difference was statistically significant(P<0.05).【Conclusion】OMT protects HUVEC cells damaged by OS by increasing the activity of cell endogenous antioxidant enzymes,up-regulating the Nrf2/HO-1 signaling pathway and inhibiting the mitochondrial apoptosis pathway.