TLR2和TLR6在鸟分枝杆菌诱导巨噬细胞凋亡中的作用  

The Effect of TLR2 and TLR6 on Macobacterium avium Induced Macrophages Apoptosis

作　　者：宁雪萍 杨东君 张依 田爽 凌敏[1] NING Xueping;YANG Dongjun;ZHANG Yi;TIAN Shuang;LING Min(Key Laboratory of Biological Molecular Medicine Research of Guangxi University,Department of Biochemistry and Moleculor Biology,Guangxi Medical University,Nanning,Guangxi,530021,China)

机构地区：[1]广西医科大学广西高校生物分子医学研究重点实验室生物化学与分子生物学教研室,广西南宁530021

出　　处：《广西科学》2018年第3期318-324,共7页Guangxi Sciences

基　　金：国家自然科学基金项目(81260245);广西自然科学基金项目(2015GXNSFAA139077)资助

摘　　要：【目的】建立鸟分枝杆菌感染巨噬细胞模型,探讨Toll样受体2(TLR2)和Toll样受体6(TLR6)在鸟分枝杆菌诱导巨噬细胞凋亡中的作用,为阐明鸟分枝杆菌致病机制提供依据。【方法】用鸟分枝杆菌感染U937细胞,于感染0h、8h、16h、24h和48h后分别提取细胞总蛋白,并通过Western blot方法检测TLR2和TLR6的表达;分别提取细胞培养上清液,利用ELISA技术检测肿瘤坏死因子(TNF-α)含量的变化情况。用鸟分枝杆菌感染阻断TLR2和TLR6后的U937细胞,通过Western blot方法分别检测其促凋亡因子BAX和抗凋亡因子BCL-2的表达量,用ELISA技术检测细胞培养上清液中TNF-α含量变化情况,用流式细胞仪检测U937细胞的凋亡变化。【结果】鸟分枝杆菌感染U937细胞可引起TLR2、TLR6和TNF-α表达上调;用鸟分枝杆菌感染阻断TLR2和TLR6后的U937细胞,可使U937细胞内促凋亡因子BAX和TNF-α表达量下降(P<0.05),抗凋亡因子BCL-2表达量升高(P<0.05),并可以明显降低感染鸟分枝杆菌后的细胞的凋亡率(P<0.05)。【结论】TLR2和TLR6与巨噬细胞凋亡相关,巨噬细胞通过TLR2和TLR6来识别鸟分枝杆菌,而鸟分枝杆菌反过来通过促进TLR2和TLR6的表达来影响巨噬细胞相关凋亡通路,从而诱导巨噬细胞凋亡。[Objective] A Mycobacterium avium infected macrophage model was established to investigate the role of Toll like receptor 2 （TLR2） and Toll like receptor 6 （TLR6） in the ap optosis of macrophages induced by Mycobacterium avium and to provide a basis for clarifying the pathogenesis of Mycobacterium avium. [Methods] U937 cells were infected with Mycobacterium avium at 0 h,8 h,16 h,24 h,and 48 h time points,respectively,and then the total proteins of the cells were used to detect the ex pression of TLR2 and TLR6 by Western blot. The cell culture supernatants were extracted re spectively and the changes of tumor necrosis fac tor （TNF-α） content were detected by ELISA. The cytokines TNF-α was measured by ELISA from U937 cells supernatant. TLR2 and TLR6 monoclonal antibody were used to treat the U937 cells 1 h before Mycobacterium avium stimu lation. The expression of BAX and BCL-2 was evaluated by Western blot. The cytokines of TNF-α was measured by ELISA from U937 cells supernatant. U937 cell apoptosis rate was detected with flow cytometry. [Results] Mycobacterium avium infection of U937 cells could cause the upregulation of TLR2,TLR6,and TNF-α expression. Blocking U937 cells after TLR2 and TLR6 infection with Mycobacterium avium could cause the expression of proapoptotic factors BAX and TNF-α in U937 cells. The amount was decreased （P 〈0.05）, the expression level of anti-apoptosis factor BCL-2 was increased （P 〈 0.05）, and the apoptosis rate of cells infected with2VIycobacterium avium was significantly decreased （P 〈 0.05）. [Conclusion]TLR2 and TLR6 are associated with macrophage apoptosis, macrophages recognize M. avium by TLR2 and TLR6,whereas Mycobacterium avium in turn affects macrophage-associated apoptosis path ways by promoting the expression of TLR2 and TLR6.

关 键 词：鸟分枝杆菌 巨噬细胞 TLR2 TLR6 细胞凋亡 

分 类 号：Q786[生物学—分子生物学]