MAPK和caspase-3在异基因CD8^+T淋巴细胞诱导血管内皮细胞凋亡中的作用

Involvement of caspase-3 and p38MAPK in allogeneic CD8^+T cell-induced apoptosis of vascular endothelial cells

目的:探讨MAPK和caspase-3在异基因CD8+T细胞诱导血管内皮细胞凋亡中的作用。方法:免疫磁珠阳性分选异基因CD8+T细胞,AnnexinⅤ/FITC试剂盒检测异基因CD8+T细胞诱导的HUVECs和HD-MECs凋亡率,Western blotting检测血管内皮细胞内caspase-3、MAPK表达。观察SB203580(p38MAPK抑制剂)、SP600125(JNK抑制剂)、PD98059(ERK抑制剂)、Z-DEVD-FMK(caspase-3抑制剂)对内皮细胞凋亡的影响。结果:异基因CD8+T细胞作用24 h和48 h后,HUVECs凋亡率分别为41.7%±10.1%和29.4%±8.3%,HDMECs凋亡率分别为28.9%±7.2%和15.2%±4.8%,与对照组相比均具有显著差异(P<0.01)。异基因CD8+T细胞作用后,HUVECs和HDMECs内磷酸化p38MAPK表达、caspase-3裂解增强,而磷酸化JNK、ERK无明显变化。Z-DEVD-FMK和SB203580可显著抑制异基因CD8+T细胞诱导的HUVECs和HEMEC凋亡,并降低内皮细胞caspase-3表达。结论:p38MAPK和caspase-3介导了异基因CD8+T细胞诱导的血管内皮细胞凋亡。

AIM： To investigate the function of caspase -3 and mitogen -activated protein kinases （MAPKs） in allogeneic CD8^＋T cell - induced apoptosis of vascular endothelial cells. METHODS ： Allogeneic CD8^＋T cells were isolated from PBMC by positive selection using magnetic beads coated with anti - CD8 antibody. After coeuhured with allogeneic CD8^＋T cells, apoptosis of human umbilical vein endothelial cells （HUVECs） and human dermal mierovascular endothelial cells （HDMECs） were detected by AnnexinV - FITC labeling. Western blotting was used to examine the change of MAPK and caspase - 3 expression in the vascular endothelial cells. The influence of SB203580 （inhibitor of p38MAPK）, SP600125 （ inhibitor of JNK）, PD98059 （ inhibitor of ERK）, Z - DEVD - FMK （ a caspase - 3 - specific peptide inhibitor） on apoptosis was also examined. RESULTS： At 24 h and 48 h time - point, the apoptosis rates of HU- VECs were 41.7% ± 10. 1% and 29.4% ±8.3%, respectively （P 〈0. 01, vs untreated HUVECs） ; the apoptosis rates of HDMECs were 28.9% ± 7. 2% and 15.2%± 4. 8% , respectively （P 〈 0.01, vs untreated HDMECs）. These effects were largely prevented by Z - DEVD - FMK and SB203580 （ P 〈 0. 05 ）. Allogeneic CD8^＋T cells enhanced cleavage of caspase- 3 and led to p38MAPK phospholation. CONCLUSION： Caspase- 3 and p38MAPK mediate allogeneie CD8 ^＋T cells - induced apoptosis of vascular endothelial cells.