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目的研究肺泡Ⅱ型(AECⅡ)细胞不同毒力结核分枝杆菌感染后炎症因子及Toll样受体(TLRs)信号通路的表达变化,为结核分枝杆菌的免疫治疗提供参考。方法建立结核分枝杆菌强毒力株H37Rv与弱毒力株BCG菌株感染的AECⅡ细胞模型,在感染12 h后采用荧光定量PCR、Western blot技术等分析TLRs信号通路及炎症因子的表达。结果强毒力H37Rv株感染12 h时细胞TLR2 mRNA、TLR4 mRNA相对表达量分别为(1.73±0.35)、(1.87±0.52),弱毒力BCG株感染分别为(1.18±0.34)、(1.33±0.28),比较差异有统计学意义(P<0.05);强毒力H37Rv株感染12 h时IL-6、IL-8及TNF-α蛋白表达水平(ODI值)分别为(2.74±0.57)、(2.95±0.43)、(5.12±0.71),弱毒力BCG株感染分别为(0.73±0.21)、(0.87±0.22)、(1.85±0.30),比较差异有统计学意义(P<0.05);两种毒力结核分枝杆菌菌株感染12 h后细胞TLR2 mRNA、TLR4mRNA表达量与IL-6、IL-8及TNF-α蛋白表达量之间均存在正相关性(P<0.05)。结论强毒力较弱毒力结核分枝杆菌感染更能活化AECⅡ细胞TLRs信号通路,促进炎症细胞因子分泌,介导机体炎症反应及影响机体免疫功能。
Objective To study the expression changes of inflammatory cytokines and Toll-like receptors (TLRs) signaling pathways after exposure to different virulence of Mycobacterium tuberculosis in alveolar type Ⅱ (AECⅡ) cells and provide a reference for the immunotherapy of Mycobacterium tuberculosis. Methods AEC Ⅱ cell model infected with Mycobacterium tuberculosis virulent strain H37Rv and attenuated BCG strain was established. The expression of TLRs signaling pathway and inflammatory cytokines were analyzed by fluorescence quantitative PCR and Western blot 12 h after infection. Results The relative expression levels of TLR2 mRNA and TLR4 mRNA were (1.73 ± 0.35) and (1.87 ± 0.52), respectively, in the virulent H37Rv strain at 12 h after infection. The attenuated BCG strains were (1.18 ± 0.34) and (1.33 ± 0.28 ) (P <0.05). The protein expression levels of IL-6, IL-8 and TNF-α in severe H37Rv strain were (2.74 ± 0.57) and 2.95 ± 0.43 and 5.12 ± 0.71, respectively. The attenuated BCG strains were (0.73 ± 0.21), (0.87 ± 0.22) and (1.85 ± 0.30) respectively, with significant difference (P <0.05) The positive expression of TLR2 mRNA and TLR4mRNA in IL-6, IL-8 and TNF-αprotein were detected at 12 h after virulent M. tuberculosis infection (P <0.05). Conclusion Mycobacterium tuberculosis infection, which is more virulent and virulent, can activate the TLRs signaling pathway of AEC Ⅱ cells, promote the secretion of inflammatory cytokines, mediate the inflammatory reaction and affect the immune function.