温石棉通过活性氧激活线粒体通路诱导A549细胞凋亡的机制研究

摘要

In this study,the authors investigated the role of reactive oxygen species (ROS) in inducing A549 apoptosis by activated mitochondria apoptosis pathway in chrysotile asbestos.Human lung epithelial cell (A549) cells were exposed to different concentrations of chrysotile asbestos dust and,on such a basis,the authors conducted detection of cell viability by MTT assay and detection of cell apoptosis rate,intracellular ROS level and mitochondrial membrane potential by flow cytometry before and after pretreatment of N-acetyl-l-cysteine (NAC) of ROS inhibitor,and then performed determination of caspase-3 and p53 protein expression level by Western blot.The experimental results show that chrysotile asbestos dust can reduce the survival rate of A549 cells and present the effectiveness relationship between the time and the dose.NAC could significantly reduce intracellular ROS level,prevent mitochondrial membrane potential declining,lower the expression level of caspase-3,p53 protein,and inhibit the occurrence of some cell apoptosis.The results indicate that ROS could activate mitochondrial apoptotic pathway by raising p53 protein expression and thus induce apoptosis of A549 cells.%探讨活性氧(reactive oxygen species,ROS)在温石棉激活线粒体通路诱导A549凋亡中的作用.采用不同浓度温石棉粉尘染毒人肺上皮细胞(A549)后,MTT法检测细胞存活率,流式细胞术检测ROS抑制剂N-乙酰半胱氨酸(N-acetyl-L-cysteine,NAC)预处理前后的细胞凋亡率、胞内ROS水平及线粒体膜电位,Western Blot法检测各组细胞半胱天冬氨酸蛋白酶3(caspase-3)、p53蛋白表达水平.实验结果显示,温石棉可降低A549细胞存活率,呈现时间-剂量效应关系;NAC可显著减少胞内ROS水平、阻止线粒体膜电位下降,并下调caspase-3、p53蛋白的表达水平,抑制部分细胞凋亡的发生.表明ROS可通过诱导p53蛋白过表达,激活线粒体凋亡通路,诱导A549细胞发生凋亡.