结核分枝杆菌eis基因表达对人THP-1细胞细胞因子分泌的影响

Effect of Mycobacterium tuberculosis eis gene on cytokines production of THP-1 cell

目的研究结核分枝杆菌eis基因表达对人单核巨噬细胞Thp-1细胞的影响,筛选出差异性表达细胞因子,探讨结核分枝杆菌eis基因在单核巨噬细胞致持留的免疫机制。方法 eis基因重组耻垢分枝杆菌MS-pmv261-eis及对照组细菌MSpmv261感染经PMA诱导分化的人单核巨噬细胞Thp-1细胞;细胞信号传导通路抑制剂用于分析作用机制;ELISA方法检测细胞上清中IL-4、IL-6、IL-10、IL-12、INF-γ表达水平;筛选差异性表达的细胞因子,RT-PCR进一步确定该细胞因子的基因表达。结果结核分枝杆菌eis基因可引起IL-10表达增加(P<0.05),而对细胞因子IL-4、IL-6、IL-12、INF-γ的表达无显著影响(P>0.05),RT-PCR结果也进一步确证了IL-10基因的高表达。2-AP和U0126能明显抑制MS-pmv261-eis(MS-eis)感染引起的IL-10的释放(P<0.05)。结论结核分枝杆菌eis基因可上调Thp-1细胞IL-10在蛋白水平和基因水平的表达,可能通过MEK1/2或PKR信号通路刺激IL-10的释放来增强机体的免疫功能,其作用可能与结核分枝杆菌持留性有关。

This study designed to investigate the impact of eis gene of Mycobacterium tuberculosis on monocyte-macrophage Thp-1 cells of human, and screen differential expression of cytokine for exploring the immune mechanism of the persistence of mycobacterium tuberculosis eis gene in the monocyte-macrophage. The human macrophages induced by PMA were infected with MS-pmv261-eis and MS-pmv261, respectively; cell signaling pathway inhibitor was used to analyze the mechanism; the levels of the expression of IL-4, IL-6, IL-10,IL-12, INF-γ in cell liquid supernatant were measured by ELISA. Furthermore, differential expression of cytokine was screened and the cytokine gene expression was determined by RT-PCR. Data showed Mycobacterium tuberculosis eis gene could increase the specific expression of IL-10（P〈0.05）, while the expression of IL-4, IL-6,IL-12, and INF-γ demonstrated no significant change（P〈0.05）; the RT-PCR results further confirmed the high expression of IL-10 gene. 2-AP and U0126 could significantly inhibit the production of IL-10 induced by MS-pmv261-eis（MS-eis）. In conclusion, eis gene of Mycobacterium tuberculosis can upregulate the expression of IL-10 in the protein and gene levels of Thp-1 cells through MEK1/2 or PKR signaling pathway, which may be related to the persistence of Mycobacterium tuberculosis.