中文摘要：

肺泡Ⅱ型上皮细胞（alveolar epithelial typeⅡcells,AECⅡ）是由呼吸道吸入的结核病原菌最先侵染的靶细胞。本研究通过建立结核分枝杆菌（Mycobacterium tuberculosis,Mtb）强毒株（H37Rv）、弱毒株（Bacillus Calmette-Guérin,BCG）感染AECⅡ细胞模型,分析不同感染时间Toll样受体（toll-liking receptors,TLRs）信号通路活化和炎症细胞因子分泌的差异,探讨AECⅡ细胞在Mtb感染时的免疫应答和持续感染的分子机制。利用H37Rv和BCG分别感染AECⅡ细胞系A549,通过荧光定量PCR和Western-blot等技术在核酸和蛋白水平分析感染6、12和24 h时TLRs信号通路信号分子和促炎细胞因子的表达变化。结果表明,m RNA表达水平检测发现,在H37Rv感染A549细胞中,TLR2、TLR4、My D88和NFκB在6 h时显著高于BCG感染组;在感染24 h时,两组TLR2、TLR4表达均上调,且差异不显著,而H37Rv感染组NFκB呈上调表达;在蛋白水平分析发现,在H37Rv感染引起A549细胞中My D88和磷酸化NFκB表达呈显著下调趋势,且高于BCG感染组;而促炎细胞因子IL-1a、IL-6、IL-12a、IL-8、TNF-α和CSF2 m RNA表达水平随着感染时间升高,且H37Rv感染组IL-1a、IL-6 IL-8、TNF-α、和CSF2的表达显著高于BCG感染组,IL-12a差异不显著。本研究发现,Mtb强毒株感染AECⅡ细胞对TLRs信号通路表现为抑制趋势,且引起细胞的炎症反应明显强于弱毒株,这为阐明AECⅡ细胞在不同毒力结核分枝杆菌感染中的免疫调控机制研究和临床肺结核的发病机制研究提供了参考依据。

英文摘要：

Alveolar epithelial typeⅡcells（AECⅡ） are the first infected target cells by tuberculosis pathogens inhaled from respiratory tract. The activation of toll-liking receptors（TLRs） signal pathway and expression of pro-inflammatory cytokines were analyzed by the different virulence Mycobacterium tuberculosis（Mtb）infected alveolar epithelial Type Ⅱ cells（AEC Ⅱ） in different time,such as H37 Rv and Bacillus CalmetteGuérin（BCG） Mycobacterium tuberculosis strains,and which was used for finding the molecular mechanisms and immune response of AEC Ⅱ cells infected with Mtb. The AEC Ⅱ cell lines of A549 were infected by H37 Rv and BCG strains respectively. The expression of key molecules and proinflammatories related to TLRs signal pathway was detected by q RT-PCR and Western-blot in the infected cell at different time points（6,12 and 24 h）. The result showed that,the m RNA expression of TLR2,TLR4,NF- κB and My D88 etc. were upregulated significantly in the presence of H37 Rv compared with BCG infected group at 6 h time point. TLR2/4was increased in both H37 Rv and BCG infected groups and there was no statistically significant difference between the 2 infected groups at 24 h time point. The expression of NF- κB was up- regulated in H37 Rv infected groups at 24 h time point. Furthermore,the protein expression of TLRs signal molecules was analyzed. The expression of interleukin-1 receptor-associated kinase 4（IRAK4） and TNF receptor-associated factor 3（TRAF3） in the infected A549 cells with BCG were significantly higher than with H37 Rv infection.The expression of IRAK4 was up-regulated at the time of 6 and 12 h of BCG infection,but suppressed at 24 h.However,the expression of IRAK4 was all suppressed at those 3 time points of H37 Rv infection. When the A549 cells infected with H37 Rv,the expression of My D88 and phosphorylated NF- κB were significantly increased at 6 h and down-regulated at 12 and 24 h. But in BCG infection,both of them were no significant difference. The expressi