活性氧激活线粒体凋亡通路介导高糖诱导心肌细胞凋亡的研究

目的探讨氧化应激和p53介导的线粒体凋亡通路在高糖诱导乳鼠心肌细胞凋亡的作用。方法将培养的SD大鼠乳鼠心肌细胞随机分为四组：对照组、高糖组（葡萄糖浓度为25mmol/L）、高糖＋抗氧化剂组（抗氧化剂浓度为10mmol/L）和高糖＋p53抑制剂组（p53抑制剂浓度为50μmol/L）。荧光分光光度计检测细胞内的活性氧,分光光度计检测上清液中的超氧化物歧化酶,流式细胞术检测细胞凋亡,Western blot对各组细胞p53和Caspase-9进行半定量分析。结果与对照组相比,高糖组心肌细胞凋亡率（P〈0.01）和ROS明显升高（P〈0.01）,上清液中的SOD含量明显降低（P〈0.01）,Caspase-9（P〈0.01）和线粒体蛋白p53表达增多（P〈0.01）。与高糖组相比,高糖＋抗氧化剂组心肌细胞凋亡率（P〈0.01）和ROS明显降低（P〈0.01）,上清液中SOD明显升高（P〈0.01）,Caspase-9（P〈0.01）和线粒体蛋白p53表达减少（P〈0.01）。与高糖组相比,高糖＋p53抑制剂组的心肌细胞凋亡率（P〈0.01）和ROS明显降低（P〈0.05）,上清液中SOD含量升高（P〈0.05）,Caspase-9（P〈0.01）和线粒体蛋白p53表达减少（P〈0.01）。结论高糖可能通过产生的活性氧介导p53转位到线粒体激活线粒体凋亡通路,引起心肌细胞凋亡。

The study that active oxygen activeates mitochondrial opoptosic pathway in cardiac myocytes apoptosis by high glucose

【Objective】To investigate the effect of oxidative stress and P53 in cardical myocytes apoptosis by high glucose.【Methods】The cardiac myocytes of rat were divided into four groups as followings： control group（Mannitol 25 mmol/L）、high glucose（25 mmol/L）、high glucose＋ antioxidant（10mmol/L）、high glucose＋ Pifithrin-α（50μmol/L）.The Active Oxygen of cardiac myocytes was measured by fluorospectrophotometer,the SOD of supernatant was measured by spectrophotometer,apoptosis was detected by flow cytometry,intracellular expression of caspase-9 and the level of p53 were analyzed by Western blotting.【Results】Compared with the control group,in the high glucose group,the results suggested that the cardiomyocyte apoptosis rate,ROS（P0.01）as well as intracellular expression of caspase-9（P0.01）and p53 i（P0.01）ncreased,while SOD activity decreased significantly（P0.01）.However,in the antioxidant group,the cardiomyocyte apoptosis rate,ROS（P0.01） as well as intracellular expression of caspase-9（P0.01）and p53 decreased（P0.01）,SOD activity increased remarkably（P0.01） compared with the high glucose group.The cardiomyocyte apoptosis rate,ROS（P0.05） as well as intracellular expression of caspase-9（P0.01）and p53 decreased（P0.01）,SOD activity increased（P0.05） in the Pifithrin-α group in comparison with that in the high glucose group.【Conclusion】The transposition of p53 to mitochondrion induced by active oxygen activeates mitochondrial opoptosic pathway in cardiac myocytes apoptosis by high glucose.