Bid蛋白在内质网和线粒体相关的凋亡过程中的作用

Role of Bid protein in the mitochondria and endoreticulum associated apoptotic pathway

目的研究 Bid 蛋白在内质网和线粒体相关的凋亡过程中的作用。方法以高三尖杉酯碱(HHT)诱导人骨髓增生异常综合征(MDS)细胞系 MUTZ-1细胞凋亡,用流式细胞术检测细胞凋亡率,RT-PCR 及 Western blot 法检测线粒体和内质网凋亡途径的相关基因及蛋白表达,激光共聚焦显微镜观察用药前后钙离子浓度和线粒体膜电位的改变以及凋亡相关蛋白 Bid 的转位。结果 0.05μg/ml HHT 作用后4 h胞质内 Ca^(2+)浓度增高,12 h达高峰,其后开始下降。HHT 作用后线粒体膜电位持续下降。内质网应激相关因子基因及蛋白表达增强,12 h达到高峰,其后开始下降。激光共聚焦显微镜观察发现 Bid 蛋白用药前位于内质网,用药12 h后转位至线粒体。结论 HHT 可以诱导 MDS 细胞凋亡,内质网和线粒体相关的凋亡途径在其中发挥了重要的作用,Bid 蛋白可能是两种凋亡途径的交叉点。

Objective To explore the role of Bid protein in the mitochondria and endoreticulum （ER） associated apoptotic pathway. Methods Apoptosis of MUTZ-1 cells induced by homoharringtonine （HHT） was measured by FACS. Mitochondria and ER associated apoptotic pathway was detected by RT-PCR and Western blotting. And the translocation of Bid protein was measured by laser scanning confocal microscope （LSCM）. Results After exposure of MUTZ-1 to HHT at 0.05 μg/ml for 24 h, typical ER-stress phenomenon induced apoptotic cells and release of Ca^2 ＋ from the cytosolic Ca^2＋ storage and the loss of mitochondrial membrane potential were observed. RT-PCR analysis revealed that mRNAs for ER stress-associated proapoptotic factor were markedly increased at 4 h after 0.05μg/ml HHT treatment and peaked at 12 h, then decreased steadly. Activation of caspase protein was also observed at 8 h. The translocation of Bid protein from ER to mitochondria was oberved at 12 h after HHT treatment. Conclusion HHT can induce MUTZ-1 cells apoptosis. The cell death may be likely mediated by the ER stress pathway as well as mitochondrial pathway and Bid protein may be the cross talk of the two apoptotic pathways.