脂联素抑制t-BHP诱导的内皮细胞凋亡作用及其可能机制探讨

The Effect of recombinant adiponectin on apoptosis induced by t-BHP in human umbilical vein endothelial cells

目的:探讨重组脂联素对第三丁基过氧化氢(t-BHP)诱导的人脐静脉内皮细胞(HUVECs)凋亡的影响及其相关的分子机制。方法:以HUVECs作为研究对象,给予t-BHP处理,模拟体外HUVECs氧化损伤细胞凋亡模型。在此基础上,用携带重组脂联素基因的腺病毒转染HUVECs,观察重组脂联素对t-BHP诱导的HUVECs凋亡的影响。用MTT法检测细胞增殖活力。Hochest/PI荧光染色检测细胞凋亡率。Western blot检测细胞凋亡相关蛋白p-JNK、JNK和Caspase 3表达水平的变化。结果:100μmol/L的t-BHP作用8 h可诱导HUVECs发生凋亡。与对照组相比,t-BHP组p-JNK、active caspase 3表达增多(P<0.01)。HUVECs高表达重组脂联素基因后,可明显抑制t-BHP诱导的HUVECs凋亡(P<0.01),下调t-BHP诱导的p-JNK、active caspase 3表达。结论:持续t-BHP氧化损伤可诱导HUVECs发生凋亡。重组脂联素可有效抑制t-BHP诱导的HUVECs凋亡,其机制与p-JNK、active caspase3的表达下调有关。

Objective： To investigate the protective effect and possible mechanism of recombinant adiponectin on apoptosis in Human Umbilical Vein Endothelial Cells（ HUVECs） induced by tert-butyl hydroperoxide（ t-BHP）. Methods： HUVECs were cultured in vitro and apoptosis was induced by t-BHP. On this basis,HUVECs were transfected with adenovirus carrying adiponectin prior to exposure to t-BHP,to further explore the protective effect of adiponectin on apoptosis induced by t-BHP. The percentage of cell viability was determined by MTT assay. The apoptotic rate was evaluated by fluorescence microsopic analysis with Hochest / PI staining. The protein levels of p-JNK,JNK and Caspase 3 were detected by Western blot. Results： Following t-BHP 100 μmol / L administration for8 h,the ratio of apoptotic cells was increased. Western blot revealed that the protein levels of p-JNK and active caspase 3 were increased（ P〈 0. 01） compared to the control group. When cells were pretreated by adenovirus with adiponectin,the apoptosis rate and protein levels of p-JNK and active caspase 3 were decreased significantly（ P〈 0. 01）. Conclusion： Continuous exposure to t-BHP induced apoptosis in HUVECs. Recombinant adiponectin protected HUVECs from apoptosis induced by t-BHP,which was correlated with the downregulation of p-JNK and active Caspase 3.