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目的:探讨CC类趋化因子配体2(C-C motif ligand 2,CCL2)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细胞,将HUVECs铺至6孔板中,待细胞融合至80-90%时,将CCL2过表达载体[pc DNA3.1(+)-CCL2]及CCL2小分子干扰RNA(si-RNA)分别转染到HUVECs中,于转染后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CCL2及ICAM-1基因m RNA表达。Western blotting检测HUVECs中CCL2及ICAM-1蛋白表达。结果:(1)与pc DNA3.1(+)组相比较,pc DNA3.1(+)-CCL2组中CCL2基因m RNA和蛋白水平均显著升高;与si-Control组相比较,si-CCL2组中CCL2基因m RNA和蛋白表达均明显下降。(2)与对照组比较,pc DNA3.1(+)-CCL2组明显增加HUVECs中ICAM-1的m RNA及蛋白表达,而si-CCL2组显著抑制HUVECs中ICAM-1的m RNA及蛋白表达。结论:CCL2能增加HUVECs中ICAM-1基因m RNA和蛋白表达,为深入认识动脉粥样硬化的发病机制提供了理论依据。
Objective: To investigate the effect of CC motif ligand 2 (CCL2) on the expression of intercellular adhesion molecule-1 (ICAM) in human umbilical vein endothelial cells (HUVECs) -1) expression. Methods: HUVECs were isolated and cultured in vitro. HUVECs were plated in 6-well plates. When the cells were fused to 80-90%, CCL2 overexpression vector pcDNA3.1 (+) - CCL2 and CCL2 small interfering RNA si-RNA) were transfected into HUVECs respectively. The cells were harvested for RNA and protein extraction at 12 h, 24 h and 48 h after transfection. Fluorescent quantitative PCR was used to detect the mRNA expression of CCL2 and ICAM-1 in HUVECs. Western blotting was used to detect the expression of CCL2 and ICAM-1 in HUVECs. Results: (1) Compared with pcDNA3.1 (+) group, the m RNA and protein levels of CCL2 gene in pcDNA3.1 (+) - CCL2 group were significantly increased. Compared with si-Control group, CCL2 group CCL2 gene m RNA and protein expression were significantly decreased. (2) Compared with the control group, pcDNA3.1 (+) - CCL2 group significantly increased the m RNA and protein expression of ICAM-1 in HUVECs, while si-CCL2 significantly inhibited the expression of m RNA and protein of ICAM-1 in HUVECs . Conclusion: CCL2 can increase the mRNA and protein expression of ICAM-1 in HUVECs and provide a theoretical basis for further understanding of the pathogenesis of atherosclerosis.