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目的研究染料木黄酮(genistein,Gen)对叔丁基过氧化氢(tert-butyl hydroperoxide,t-BHP)诱导的人血管内皮细胞株EA.hy926凋亡的保护效应并探索其相关分子机制。方法以t-BHP建立氧化应激损伤体外模型,设立空白对照组、Gen和t-BHP处理组,CCK-8法、流式细胞仪检测Gen对内皮细胞活力、凋亡的影响;Western blot法检测Gen对内皮细胞Caspase-3表达和PPARγ蛋白表达的影响;免疫细胞化学法检测Gen对细胞PPARγ表达定位。结果 t-BHP能明显抑制EA.hy926生长,至最大浓度240μmol/L时,抑制率达到(93.66±4.66)%,与对照组相比,差异有统计学意义(P<0.05),半数抑制浓度(IC50)约为100μmol/L;Gen(5～5000nmol/L)能显著抑制t-BHP诱导的内皮细胞凋亡,并呈明显的剂量-效应关系,500nmol/L预处理组总凋亡率为(12.20±1.28)%,与对照组相比,差异有统计学意义(P<0.05);Gen能够显著激活PPARγ蛋白的表达,并诱导其核易位。结论 Gen对t-BHP诱导的内皮细胞凋亡有保护效应,其保护效应可能与PPARγ的活化有关。
Objective To investigate the protective effect of genistein (Gen) on the apoptosis of human vascular endothelial cell line EA.hy926 induced by tert-butyl hydroperoxide (t-BHP) and explore its related molecular mechanism. Methods The in vitro model of oxidative stress injury was established by t-BHP. The blank control group, Gen and t-BHP treatment groups were established. The effects of Gen on the viability and apoptosis of endothelial cells were detected by CCK-8 and flow cytometry; The effects of Gen on Caspase-3 expression and PPARγ protein expression in endothelial cells were detected. The expression of PPARγ in cells was detected by immunocytochemistry. Results t-BHP could significantly inhibit the growth of EA.hy926, and the inhibition rate reached (93.66±4.66)% when the maximum concentration was 240 μmol/L. Compared with the control group, the difference was statistically significant (P<0.05). (IC50) was approximately 100 μmol/L; Gen (5-5000 nmol/L) significantly inhibited t-BHP-induced endothelial cell apoptosis in a dose-response relationship, and the total apoptosis rate was 500nmol/L preconditioning. (12.20±1.28)%, compared with the control group, the difference was statistically significant (P<0.05); Gen can significantly activate the expression of PPARγ protein and induce its nuclear translocation. Conclusion Gen has a protective effect on t-BHP-induced apoptosis of endothelial cells, and its protective effect may be related to the activation of PPARγ.