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目的观察艾溴利平对高糖诱导人脐静脉内皮细胞凋亡和蛋白激酶B(PKB)磷酸化的影响,并探讨其分子机制。方法应用终浓度为33.3 mmol·L-1葡萄糖诱导内皮细胞HUVEC-12细胞株高糖模型,实验分为正常对照组、高糖模型组、艾溴利平组和抑制剂组,Hoechst33258染色及AnnexinV和PI双染流式细胞术检测细胞凋亡;Western blotting检测PKB总蛋白、PKB苏氨酸308(PKB-Thr308)及PKB丝氨酸473(PKB-Ser473)的磷酸化水平。结果高糖模型组内皮细胞凋亡指数(AI)增加为(14.07±s 0.25)%,高于正常对照组(P<0.01),艾溴利平8μmo·lL-1组AI降至(5.42±0.13)%,与高糖模型组相比有非常显著差异(P<0.01),抑制剂组AI增加至(12.00±0.10)%。低浓度艾溴利平(2、4、8μmo·lL-1)短时间(6、12、24、48 h)增加PKB磷酸化(P<0.01);而阻断PKB活性可显著抑制艾溴利平的抗凋亡作用。结论艾溴利平能抑制高糖引起的人脐静脉内皮细胞凋亡,可能与信号分子PKB的激活有关。
Objective To observe the effect of abirapril on high glucose-induced apoptosis of human umbilical vein endothelial cells and phosphorylation of protein kinase B (PKB), and to explore its molecular mechanism. Methods The HUVEC-12 cells were induced with 33.3 mmol·L-1 glucose at final concentration. Hoechst33258 staining, Annexin V and Hoechst33258 staining were used to observe the changes of HUVEC- PI double staining and flow cytometry. The phosphorylation level of PKB total protein, PKB threonine 308 (PKB-Thr308) and PKB serine 473 (PKB-Ser473) were detected by Western blotting. Results The apoptosis index (AI) in hyperglycemia model group was (14.07 ± 0.25)% higher than that in normal control group (P <0.01), AI in 8 μmo · lL-1 group was decreased to (5.42 ± 0.13) )%, Which was significantly different from that in high glucose group (P <0.01). The AI in inhibitor group increased to (12.00 ± 0.10)%. PKB phosphorylation was increased in a short time (6, 12, 24, 48 h) at a low concentration of 2, 4, 8μmo · L-1, while blockade of PKB activity inhibited the release of abirapril Anti-apoptotic effect. Conclusion AiBuliLing can inhibit the apoptosis of human umbilical vein endothelial cells induced by high glucose, which may be related to the activation of PKB.