【摘要】 目的探索C5a是否与LPS、毒源性大肠杆菌O157产生的外毒素—V毒素(verotoxin)具有相似的可以导致血管内皮细胞(VEC)凋亡的作用。方法流式细胞仪检测细胞凋亡发生率,首先探讨rhC5a刺激的质量浓度,分别为0.2μg/mL、0.5μg/mL、1μg/mL和1.5μg/mL刺激12 h检测VEC凋亡发生率;在rhC5a质量浓度均为1μg/mL条件下,时间分别为2 h、6 h、12 h、18 h和24 h,检测VEC凋亡发生率。Western blot方法检测凋亡相关蛋白。结果在rhC5a质量浓度为0.2μg/mL、0.5μg/mL、1μg/mL和1.5μg/mL刺激12 h诱导的凋亡发生率分别为4.58%、7.87%、17.94%和19.03%。在确定rhC5a质量浓度1μg/mL的条件下,VEC发生凋亡呈时间依赖关系,在2 h、6 h、12 h、18 h和24 h的时相,细胞凋亡发生率分别为4.58%、12.78%、18.12%、19.08%和19.96%。rhC5a刺激VEC细胞,calpain-2蛋白表达呈时间依赖性,而calpain-1蛋白的表达为浓度依赖性。结论C5a可以导致VEC细胞发生凋亡,calpain参与了这类细胞的凋亡进程。更多还原

【Abstract】 Objective To investigate the apoptotic effects of C5a on vascular endothelial cells(VECs).Methods The VECs were treated with rhC5a at different concentration(0.2,0.5,1 and 1.5 μg/mL,respectively) for 12 hours or with 1 μg/mL rhC5a at different time(0,2,6,12,18,and 24 h).Flow cytometry analysis with Annexin V/PI staining was used to identify the undergoing apoptosis and necrosis of VECs after rhC5a treatment.The calpain-1 and calpain-2 proteins were detected with Western blotting.Results The percentage of apoptotic cells were 4.58%,7.87%,17.94% and 19.03%,respectively after VECs treated with rhC5a at 0.2,0.5,1,and 1.5 μg/mL respectively for 12 hours.Apoptotic cells were 4.58%,12.78%,18.12%,19.08%,and 19.96% respectively in VECs treated with 1 μg/mL rhC5a for 2,6,12,18 and 24 h.Western blotting shown the calpain-2 protein was time-dependently induced by rhC5a, however,the calpain-1 proteins were dose-dependently induced by rhC5a.Conclusion Calpain contributes to cellar injury in VECs with rhC5a.更多还原