OX-LDL介导内皮细胞损伤及其对TLR-4和EpCAM表达的影响

Damage of endothelial cells induced by OX-LDL and its influence in expressions of TLR-4 and EpCAM

目的:研究氧化型低密度脂蛋白(OX-LDL)对血管内皮细胞的损伤效应及其对Toll样受体4(TLR-4)和上皮细胞黏附分子(EpCAM)表达的影响。方法:培养人脐静脉内皮细胞ECV-304,实验分为对照组和实验组(25、50、100和200mg·L-1 OX-LDL),处理后细胞继续培养24h,台盼蓝拒染法检测细胞活力;流式细胞术检测活性氧(ROS)水平、线粒体膜电位(MMP)、细胞周期与凋亡以及EpCAM和TLR-4表达水平。结果:与对照组比较,25、50、100和200mg·L-1 OX-LDL组ECV-304细胞内ROS水平升高,细胞活力和MMP下降(P<0.05)。与对照组比较,25和50mg·L-1 OX-LDL组S期细胞百分率升高,而G0/G1期细胞略减少,SubG1期细胞(凋亡细胞)随OX-LDL浓度增加而增加。与对照组比较,50mg·L-1 OX-LDL组24h时ECV-304细胞膜表面TLR-4和EpCAM表达水平均明显升高(P<0.05)。结论:OX-LDL具有提高内皮细胞ROS水平、降低细胞活力和MMP以及诱导细胞凋亡等损伤效应,此过程中ECV-304细胞中TLR-4和EpCAM水平升高。

Objective To study the damage effect of oxidized low density lipoprotein （OX-LDL）on the endothelial cells and its influence in the expressions of Toll like receptor 4 （TLR-4）and epithelial cell adhesion molecule （EpCAM）.Methods The human endothelial cells ECV-304 were cultured in vitro and treated by different concentrations of OX-LDL for 24 h,and the cell vitality was measured by Taipan blue rejection test.The reactive oxygen species （ROS）level,mitochondrial membrane potential （MMP）,cell cycle and apoptosis were detected by flow cytometry （FCM）.The expression of TLR-4 and EpCAM were also analyzed by FCM.Results Compared with control group,the ROS levels in ECV-304 cells in 25,50,100,and 200 mg·L-1 OX-LDL were increased, but the cell vitalities and MMP were decreased （P 〈 0.05）.Compared with control group,the percentages of S phase cells in 25 and 50 mg·L-1 OX-LDL groups were increased,the G0 G1 phase cells were slightly reduced,and the SubG1 phase cells （apoptotic cells）were increased with the increasing of OX-LDL concentration.Compared with control group,the TLR-4 and EpCAM expression levels in ECV-304 cells in 50 mg·L-1 OX-LDL group at 24 h were significantly increased （P 〈 0.05 ）. Conclusion OX-LDL could increase the ROS level and induce the apoptosis in vascular endothelial cells,while increase the cell vitality and MMP.During the damage process,the expresion levels of TLR-4 and EpCAM are up-regulated.