BPDE诱导GC-1细胞凋亡及褪黑素的保护作用

发布时间：2018-04-25 07:04

本文选题：-二羟- + -环氧苯并[a]芘　；参考：《第三军医大学学报》2017年23期

【摘要】：目的探讨7,8-二羟-9,10-环氧苯并[a]芘(benzo[a]pyrene-7,8-diol-9,10-epoxide,BPDE)对小鼠精原细胞株GC-1的细胞毒性机制及褪黑素的保护作用。方法不同浓度BPDE处理GC-1细胞,采用Annexin V/PI染色及流式细胞仪检测细胞凋亡,JC-1探针染色检测线粒体膜电位,Western blot检测细胞质Cyt C及细胞总蛋白中caspase-9/3的活化水平,DCFH-DA探针及流式细胞仪检测细胞内的活性氧(reactive oxygen species,ROS)水平。褪黑素预处理细胞后再进行BPDE染毒,采用上述方法检测褪黑素对BPDE细胞毒性的影响。结果 BPDE可提高GC-1细胞凋亡率,存在剂量-效应关系。同时,BPDE可降低细胞线粒体膜电位,促进线粒体Cyt C的释放,提高细胞中caspase-9和caspase-3蛋白的活化水平,并诱导细胞中ROS水平升高。褪黑素预处理则可降低BPDE诱导的细胞凋亡,抑制Cyt C的释放及caspase-9/3的活化,并激活Nrf-2/ARE抗氧化通路,降低细胞ROS水平。结论BPDE可诱导小鼠精原细胞GC-1线粒体途径细胞凋亡和氧化应激,而褪黑素在这一过程中起保护作用。
[Abstract]:Objective to investigate the cytotoxic mechanism of 7-dioxy-8-diol-8-diol-9 10-epoxide (BPDE) on mouse spermatogonia cell line GC-1 and the protective effect of melatonin on the cytotoxicity of pyrene benzo [a] pyrene benzo (a) on mouse spermatogonia cell line GC-1. Methods GC-1 cells were treated with different concentrations of BPDE. Annexin V/PI staining and flow cytometry were used to detect apoptosis and JC-1 probe staining. Mitochondrial membrane potential was detected by Western blot to detect the activation level of Cyt C in cytoplasm and caspase-9/3 in total cell protein. DCFH-DA probe was used to detect the reactive oxygen species-ROSs in cells by flow cytometry. The effect of melatonin on the cytotoxicity of BPDE cells was detected by the method mentioned above after the cells were pretreated with melatonin and then treated with BPDE. Results BPDE could increase the apoptosis rate of GC-1 cells in a dose-effect relationship. At the same time, BPDE could decrease the mitochondrial membrane potential, promote the release of mitochondrial Cyt C, increase the activation of caspase-9 and caspase-3 protein, and induce the increase of ROS level. Melatonin pretreatment could decrease the apoptosis induced by BPDE, inhibit the release of Cyt C and the activation of caspase-9/3, activate the antioxidant pathway of Nrf-2/ARE, and decrease the level of ROS. Conclusion BPDE can induce apoptosis and oxidative stress in GC-1 mitochondria pathway of mouse spermatogonia, and melatonin plays a protective role in this process.
【作者单位】：第三军医大学军事预防医学院毒理学研究所;
【基金】：国家自然科学基金重点项目(81130051)~~
【分类号】：R114

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