MicroRNA-495对人脐静脉内皮细胞ICAM-1表达的影响

目的:探讨MicroRNA-495(miR495)对人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)中细胞间粘附分子-1(intercellular adhesion molecule-1,ICAM-1)表达的影响。方法:体外分离培养HUVECs,将HUVECs铺至6孔板中,待细胞融合至80%时,将miR495模拟物(miR495 mimics)、miR495抑制剂(miR495 inhibitor)及其相应的对照negative control(NC)、inhibitor NC分别转染到6孔板的HUVECs中,于转染后不同时间点(12 h、24 h和48 h)收集细胞进行RNA及蛋白提取。荧光定量PCR方法检测HUVECs中miR495及ICAM-1基因mRNA表达。Western blotting检测HUVECs中ICAM-1蛋白表达。结果:(1)与NC相比较,miR495 mimics组中miR495水平显著升高;与inhibitor NC组比较,miR495 inhibitor组中miR495表达明显下降。(2)与NC组比,miR495 mimics组明显降低HUVECs中ICAM-1的mRNA及蛋白表达;与inhibitor NC组比,而miR495 inhibitor组能显著增加HUVECs中ICAM-1的m RNA及蛋白表达。结论:MiR495能降低HUVECs中ICAM-1基因mRNA和蛋白表达。

Effect of MicroRNA-495 on the Expression of ICAM-1 in the HUVECs

ABSTRACT Objective: To investigate the effect of microRNA-495 (miR495) on the expression of intercellular adhesion molecule-1 (ICAM-1) in human umbilical vein endothelial cells (HUVECs). Methods: HUVECs cells cultured in vitro were plated into 6-well plate, when cells were grown to 80 %, miR495 mimics, miR495 inhibitor or negative control (NC), inhibitor NC was transfected into HUVECs. At different time points after transfection (12 h, 24 h and 48 h), HUVECs were collected for RNA and protein extraction. The mRNA ex- pressions of miR495 and ICAM-1 were examined using quantitative real time RT-PCR. ICAM-1 protein expression was examined using western blotting. Results: (1) Compared with NC group, miR495 expression was significantly higher in miR495 mimics group; Compared with inhibitor NC group, miR495 expression was significantly decreased in miR495 inhibitor group. (2) Compared with NC, miR495 sig- nificantly inhibited the mRNA and protein expressions of ICAM-1 in HUVECs; Compared with inhibitor NC, miR495 inhibitor signifi- cantly increased the mRNA and protein expressions of ICAM-1 in HUVECs. Conclusion: MiR495 could affect ICAM-1 expression in HUVECs.