HIV-1Nef调节内皮细胞黏附分子ICAM-1的表达

HIV-1Nef up-regulates the expression of adhesion molecule ICAM-1 on endothelial cells

目的研究HIV-1的调节基因Nef对ECV304细胞ICAM-1表达的影响,从而为分析HIV-1感染引起内皮细胞生物学活性的变化,以及为阐明Nef参与HIV-1致病的分子机制奠定基础。方法应用本实验室已经建立保存的HIV-1Nef基因在内皮细胞的稳定表达细胞株ECV304-Nef和其阴性对照细胞株ECV304 pcDNA 3.1(+),通过RT-PCR、实时定量PCR(real-time PCR)、Western blot、FCM和细胞黏附试验分析ECV304-Nef细胞ICAM-1的表达水平。结果 RT-PCR、real-time PCR结果显示ECV304-Nef细胞ICAM-1 mRNA表达水平明显升高,为对照组的(4.3±0.2)倍;Western blot结果示ECV304-Nef细胞I-CAM-1蛋白的表达水平高于对照组;FCM分析显示ECV304-Nef细胞和对照组细胞ICAM-1阳性细胞百分率分别为(35.3±2.2)%和(12.5±0.8)%(P<0.01),两组间ICAM-1表达有显著差异。细胞黏附实验观察到ECV304-Nef细胞黏附的Jurkat细胞数明显多于对照组,荧光仪定量分析结果显示ECV304-Nef细胞黏附的Jurkat细胞的荧光强度值显著高与对照组(P<0.05)。结论本实验证实了HIV-1Nef基因可以上调血管内皮细胞细胞黏附分子ICAM-1的表达。

This study aimed to explore the effects of HIV-1 Nef on ICAM-1 over expression on the endothelial cells for investigating the molecule mechanism of HIV-1 Nef in AIDS pathogenesis. We utilized a stable Nef expressing cell line ECV304-Nef and its negative control cell line ECV304 pcDNA 3.1 （＋）, which have been established and preserved in our laboratory, to measure 1CAM-1 expression of ECV304-Nef by real-time PCR, Western blot, FCM and cellular adhesion assay. Compared with the control cells, the 1- CAM -1 mRNA expression level was up-regulated by （4.3±0.2） folds in ECV304-Nef ceils; cellular adhesion assay indicated that adhesion of Jurkat cells to ECV304-Nef cells was significantly increased （P 〈 0.05）; FCM results showed that the percentage of ICAM-1 positive cells in ECV304-Nef and ECV304 pcDNA 3.1（＋） cells were （35.3±2.2）% and （12.5±0.8）%, respectively （P 〈 0.01）; ICAM-1 protein expression level increased in ECV304-Nef cells. Results above mean that HIV-1 Nef could enhance ICAM-1 gene expression in vascular endothelial ceils, which could take part in HIV-1 pathogenesis.