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目的:比较不同浓度过氧化氢(hydrogen peroxide,H2O2)对人外周血来源的内皮祖细胞(endothelial progenitor cells,EPCs)生存能力的改变及其对凋亡相关蛋白表达的影响。方法:采用密度梯度离心法和差速贴壁法从人外周静脉血中分离培养内皮祖细胞。选取传代后第3代EPCs作为研究对象,以终浓度分别为50μmol/L、100μmol/L、150μmol/L、200μmol/L、300μmol/L和400μmol/L的过氧化氢处理内皮祖细胞12 h,同时设立正常处理对照组。CCK-8法检测各组内皮祖细胞生存能力的差异;Western blot分析各组内皮祖细胞中凋亡相关蛋白Bax、Bcl-2和p53的蛋白表达情况。结果:与对照组相比,在浓度为50μmol/L、100μmol/L和150μmol/L过氧化氢处理组中细胞存活能力逐渐增强,促凋亡蛋白Bax、p53随之下调,抗凋亡蛋白Bcl-2则显著上调;在浓度为200μmol/L、300μmol/L和400μmol/L过氧化氢处理组中细胞生存能力相对于正常对照组逐渐减弱,促凋亡蛋白Bax、p53表达水平逐渐增加,抗凋亡蛋白Bcl-2表达水平下降。结论:过氧化氢对内皮祖细胞存活能力和凋亡相关蛋白Bax、Bcl-2、p53表达的影响均呈双相性变化。
OBJECTIVE: To compare the effects of different concentrations of hydrogen peroxide (H2O2) on the viability of human peripheral blood endothelial progenitor cells (EPCs) and their effects on the expression of apoptosis-related proteins. Methods: Endothelial progenitor cells were isolated and cultured from human peripheral blood by density gradient centrifugation and differential adhesion method. Endothelial progenitor cells (EPCs) were selected as the third generation of EPCs after passage, EPCs were treated with 50μmol / L, 100μmol / L, 150μmol / L, 200μmol / L, 300μmol / L and 400μmol / L H2O2 for 12h, At the same time set up a normal control group. CCK-8 method was used to detect the difference of viability of endothelial progenitor cells. Western blot was used to analyze the protein expression of Bax, Bcl-2 and p53 in endothelial progenitor cells. Results: Compared with the control group, the cell survival ability gradually increased in the groups of 50μmol / L, 100μmol / L and 150μmol / L hydrogen peroxide, the pro-apoptotic proteins Bax and p53 were down-regulated and the anti-apoptotic protein Bcl -2 was significantly up-regulated. Compared with the normal control group, the cell viability in 200μmol / L, 300μmol / L and 400μmol / L H2O2-treated groups decreased gradually, and the expression of pro-apoptotic proteins Bax and p53 increased gradually. The apoptosis protein Bcl-2 expression decreased. CONCLUSION: Hydrogen peroxide has a biphasic effect on the viability of EPCs and the expression of Bax, Bcl-2 and p53.