摘要目的 探讨氧化型低密度脂蛋白(oxidized low-density lipoprotein, oxLDL)对人脐静脉内皮细胞(human umbilical vein endothelial cells, HUVECs)自噬和Akt/mTOR/p70S6K信号通路的影响.方法 将培养的HUVECs分为oxLDL组和对照组,分别用100 μg/ml oxLDL和等体积磷酸盐缓冲液处理.在培养6 h和12 h后收集细胞.应用透射电子显微镜观察自噬小体,实时荧光定量聚合酶链反应检测微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3, LC3)和p62 mRNA表达,应用蛋白质印迹法检测LC3、p62、p-Akt/Akt、p-mTOR/mTOR和p-p70S6K/p70S6K蛋白表达.结果 与对照组相比,oxLDL处理组细胞内的自噬小体数量明显增多(P<0.05),LC3 mRNA及蛋白表达水平均显著增高(P均<0.05),而p62 mRNA和蛋白表达水平均显著降低(P均<0.05).此外,oxLDL处理组Akt、mTOR、p70S6K磷酸化蛋白表达水平均较对照组显著降低(P均<0.01),但Akt、mTOR和p70S6K总蛋白表达水平与对照组无显著差异.结论 oxLDL可通过抑制Akt/mTOR/p70S6K信号通路诱导HUVECs自噬.

abstractsObjectiveTo investigate the effects of oxidized low density lipoprotein (oxLDL) on autophagy and its effect on Akt/mTOR/p70S6K signaling pathway in human umbilical vein endothelial cells (HUVECs).MethodsThe cultured HUVECs were divided into either an oxLDL or a control group, and treated with 100 μg/ml oxLDL and equal volume phosphate buffer solution respectively.The cells were collected after 6 h and 12 h.Transmission electron microscopy was used to observe the autophagosome.Real-time fluorescence quantitative polymerase chain reaction was used to detect expression levels of microtubule associated protein 1 light chain 3 (LC3) and p62 mRNA.Western blot was used to detect the expression levels of LC3, p62, P-Akt/Akt, P-mTOR/mTOR, and p-p70S6K/p70S6K.Results Compared with the control group, the number of intracellular autophagosome increased obviously (P<0.05), LC3 mRNA and protein expression levels increased significantly (all P<0.05), and p62 mRNA and protein expression levels decreased significantly (all P<0.05) in the oxLDL group.In addition, the phosphorylated protein expression levels of Akt, mTOR and p70S6K in the oxLDL group were significantly decreased than those in the control group (all P<0.01).However, total protein levels of Akt, mTOR, and p70S6K were not significantly different between the oxLDL group and the control group.Conclusion oxLDL may induce the autophagy of HUVECs via inhibiting the Akt/mTOR/p70S6K signaling pathway.