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目的观察热休克蛋白70(Hsp70)对缺糖引起的HeLa细胞凋亡的影响,探讨其与凋亡相关的Bcl-2家族成员的相互作用关系。方法用人正义Hsp70重组质粒稳定转染HeLa细胞获得Hsp70过表达细胞。采用Hsp70过表达和正常HeLa细胞无糖培养建立两组细胞损伤模型,并均取3个平行样本。四甲基偶氮唑盐(MTT)法测细胞活率;Hoechst染色法和Giemsa染色法检测细胞凋亡率;RT-PCR法检测Bax和Bcl-2的表达变化,并计算其灰度比值;细胞免疫化学和荧光免疫法检测Bax构象改变情况。结果缺糖48h内,Hsp70明显抑制了HeLa细胞凋亡并增强其活力,同时也抑制了凋亡细胞中Bax/Bcl-2的增高以及Bax的构象改变。结论缺糖诱导下,热休克蛋白70在HeLa细胞中能够通过与Bcl-2家族成员作用而抑制凋亡的发生。
Objective To investigate the effect of heat shock protein 70 (Hsp70) on HeLa cell apoptosis induced by glucose deprivation and to explore the interaction between Hsp70 and apoptosis-related Bcl-2 family members. Methods HeLa cells were stably transfected with recombinant human Hsp70 plasmid to obtain Hsp70 overexpression cells. Two models of cell injury were established by Hsp70 overexpression and normal glucose-free culture of HeLa cells. All three parallel samples were taken. The cell viability was measured by MTT assay; the apoptosis rate was detected by Hoechst staining and Giemsa staining; the expression of Bax and Bcl-2 was detected by RT-PCR; Immunocytochemistry and fluorescence immunoassay to detect changes in the conformation of Bax. Results Within 48 hours after hypoglycemia, Hsp70 significantly inhibited HeLa cell apoptosis and enhanced its viability, and also inhibited the increase of Bax / Bcl-2 and the conformational change of Bax in apoptotic cells. Conclusion Hypoxia-induced heat shock protein 70 inhibits apoptosis in HeLa cells by interacting with Bcl-2 family members.