摘要
目的探讨热休克蛋白90(heat shock protein 90,HSP90)在对抗氯化钴(cobalt chloride,CoCl2)诱导H9C2心肌细胞损伤中的作用。方法应用不同浓度的CoCl2处理H9C2心肌细胞,建立化学性缺氧诱导心肌细胞损伤的实验模型。应用CCK-8比色法检测细胞存活率;Western-blot法检测HSP90的表达;双氯荧光素染色荧光显微镜照相检测细胞内活性氧(reactive oxygen species,ROS)水平;超氧化物歧化酶(superoxidedismutase,SOD)检测试剂盒分析SOD活性抑制率。结果在400~1000μmol/L浓度范围内,CoCl2处理H9C2心肌细胞36 h,呈剂量依赖性地抑制细胞存活率。在0.5~36 h时间范围内,600μmol/L CoCl2呈时间依赖性地促进H9C2心肌细胞HSP90的表达。2~16μmol/L HSP90抑制剂17-丙烯胺基-17-去甲氧基格尔德霉素(17-allylamino-17demethoxygeldanamycin,17AAG)呈剂量依赖性地加重600μmol/L CoCl2对H9C2细胞存活率的抑制。2μmol/L 17AAG本身不损伤H9C2心肌细胞,但明显地加强CoCl2增加H9C2心肌细胞内ROS生成的作用,并增加CoCl2对SOD活性的抑制。结论HSP90表达上调可能是H9C2心肌细胞对抗化学性缺氧的内在防御机制之一。
Objective To investigate the protection of heat shock protein 90 (HSP90) against H9C2 cardiac cell injuries induced by cobalt chloride (COCl2). Method H9C2 cells were treated with COCl2 at different concentrations to set up the chemical hypoxia-induced cardiomyocyte injury model. Cell viability was measured by using cell counter kit (CCK-8). The expression of HSP90 was tested by Western-blot assay. Intracellular reactive oxygen species (ROS) was evaluated by 2′,7′- dichlorfluorescein-diacetate (DCFH-DA) staining and photofluorography. The inhibitory rate of superoxide dismutase (SOD) was detected by commercial kit. Result H9C2 cell viability was dose-dependently damaged by COCl2 at concentrations from 400 to 1000 μmol/L for 36 h. The expression of HSP90 in H9C2 cells was unregulated by 600 μmol/L COCl2 in a time-dependent manner. 17-allylamino-17-demethoxygeldanamycin (17AAG) (an inhibitor of HSP90) at 2-16 μmol/L promoted dose-dependently the inhibition of 600 μmol/L COCl2 on the cell viability. 17AAG at 2 μmol/L alone did not damage H9C2 cells, but significantly enhanced overproduction of ROS induced by COCl2, and increased the CoCl2-induced inhibition of SOD activity. Conclusion Upregulation of HsP90 expression may be one of the endogenous defense mechanisms for H9C2 cells against chemical hypoxia.
出处
《热带医学杂志》
CAS
2009年第8期871-874,共4页
Journal of Tropical Medicine
基金
广东省科技计划资助项目(No.2006B60501024
No.2007B080701030)
关键词
热休克蛋白
氯化钴
心肌细胞
活性氧
heat shock protein
cobalt chloride
cardiomyocyte
reactive oxygen species