摘要
目的 探讨线粒体及其细胞色素C释放 ,在过氧化氢诱导的人脐静脉内皮细胞凋亡信号途径中的作用。方法 人脐静脉内皮细胞和 10 0 μmol/LH2 O2 共同培养 2 4h ,或内皮细胞先与环孢霉素A(cyclosporinA ,CsA)孵育 30min后加入H2 O2 ;在不同时相点取细胞作DAPI染色计数凋亡细胞数量 ;同时用Rhoadmine12 3聚集法观察线粒体通透性的变化 ;Westernbolt检测早期线粒体和血浆中细胞色素C浓度的变化。 结果 H2 O2 处理细胞后 ,细胞呈现出典型的凋亡形态 :细胞皱缩 ,胞核致密 ,凋亡小体形成。从处理后 4h起凋亡细胞数量迅速增加 ,至 12h达峰值。CsA可以明显抑制H2 O2 诱导内皮细胞凋亡的作用。H2 O2 可诱导线粒体细胞色素C浓度的下降 ,使得胞浆中细胞色素C浓度升高 ;同样线粒体中Rhoadmine12 3浓度下降 ,线粒体通透性增加 ,而CsA则可以明显抑制二者的变化。 结论 H2 O2 促进细胞色素C释放进入胞浆 ,导致内皮细胞的凋亡。
Objective To explore the role of mitochondria and from which released cytochrome C in the apoptosis of HUVECs induced by H 2O 2. Methods HUVECs were cultured with H 2O 2 of l00mol/L for 24 hrs. H 2O 2 was added after that HUVECs were precultured with CsA (cyclosporin A) for 30 mins. The cell samples were collected at different time points for DAPI staining and counting of apoptotic cell number. Simultaneously,the changes of mitochondrial permeability was observed by Rhoadmine 123 accumulation. The changes of the cytochrome C concentrations in plasma and mitochondria were determined by western blot. Results HUVECs exhibited obvious apoptosis after being processed by H 2O 2. The apoptotic cell number increased since 4 hrs of culturation of HUVECs with H 2O 2, and reached peak level at 12 hrs. And the HUVEC apoptosis induced by H 2O 2 could be inhibited significantly by CsA. H 2O 2 could lead to the decrease of mitochondrial cytochrome C concentration, which in turn lead to the increase of cytoplasmic cytochrome C concentration. Similarly,mitochondrial Rhoadmine 123 concentration decreased and the mitochondrial permeability increased. But CsA could obviously inhibit the changes of both mitochondrial Rhoadmine 123 and permeability. Conclusion H 2O 2 could induce cytochrome C releasing to cytoplasma which lead to endothelial apoptosis. And CsA could inhibit the apoptosis by maintaining the normal function of mitochondrial membrane.
出处
《中华烧伤杂志》
CAS
CSCD
2001年第6期360-363,共4页
Chinese Journal of Burns
基金
国家重点基础研究发展规划资助项目 (G19990 5 42 0 3)
关键词
细胞培养
过氧化氢
细胞色素C
线粒体
Apoptosis
Endothelium,Hydrogen Peroxide
Cytochrome C
Mitochondria
