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目的 研究曲霉菌素诱导人胚肾细胞毒性作用机制。方法 结晶甲紫法用于细胞存活率研究。琼脂糖凝胶及Burton法研究细胞核DNA断裂片段。以水解特异性底物Ac DEVD AMC活性为研究指标 ,测定胞浆半胱天冬酶 (caspase) 3类蛋白酶活性。采用蛋白质印迹法检测细胞半胱天冬酶 3蛋白表达。采用荧光标记探针、流式细胞仪技术研究细胞核DNA核型及细胞活性氧的产生。结果 曲霉菌素浓度依赖性地诱导人胚肾细胞凋亡 ,最大效应浓度为 1 .0mg·L-1 。BAF ,半胱天冬酶 3蛋白抑制剂和N 乙酰半胱氨酸 (活性氧抑制剂 )能显著性抑制曲霉菌素诱导人胚肾细胞凋亡作用。结论 半胱天冬酶类及活性氧调节曲霉菌素诱导的人胚肾细胞凋亡。
Objective To study the mechanism of streptozotocin-induced human embryonic kidney cell toxicity. Methods Crystalline purple method was used to study cell viability. Agarose gel and Burton method to study nuclear DNA fragmentation. The Ac DEVD AMC activity of hydrolyzate-specific substrate was used as a research index to determine the activity of the caspase 3 protease. Western blotting was used to detect the expression of caspase 3 protein. Fluorescently labeled probes and flow cytometry were used to study the nuclear DNA and the production of reactive oxygen species. Results The concentration of amikacin induced human embryonic kidney cell apoptosis in a concentration-dependent manner. The maximum effect concentration was 1.0 mg · L -1. BAF, caspase 3 inhibitor and N-acetylcysteine (ROS inhibitor) could significantly inhibit the apoptosis of human embryonic kidney cells induced by streptomycin. Conclusions Caspases and reactive oxygen species regulate the apoptosis of human embryonic kidney cells induced by streptomycin.