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目的:肺脏上皮细胞在抵抗外源微生物入侵中发挥着重要保护作用,最近研究揭示,在器官组织发育中发挥调控作用的经典Wnt信号具有免疫调节功能。因此,有必要研究肺脏上皮细胞A549中Wnt信号经典途径在抗结核分枝杆菌(Mtb)感染中的免疫调节作用。方法:用牛结核分枝杆菌卡介苗(BCG)刺激转入Wnt信号报告基因质粒BATflash的A549细胞,利用双荧光素酶报告系统、Western blot和ELISA方法分析细胞中Wnt信号经典途径主要相关信号因子和免疫反应炎症因子的表达变化。结果:BCG感染A549细胞后抑制了Wnt信号报告荧光素酶的活性,Western blot结果表明,随着细胞浆内磷酸化β-catenin含量增加,Wnt信号的抑制基因GSK3β和Axin2表达上调,而细胞核中的效应子β-catenin基因活性组分和下游转录因子TCF4与Lef-1的表达下调;BCG感染过表达β-catenin的细胞时导致IL-6、NF-κB、MyD88和TRAF6蛋白表达下调,但不影响TNF-α的表达。结论:肺脏上皮细胞在抗结核分枝杆菌感染过程中通过下调Wnt信号活性而抑制细胞中的MyD88/TRAF6/NF-κB信号通路来降低免疫反应以保护宿主细胞免受感染造成的免疫损伤。
PURPOSE: Pulmonary epithelial cells play an important protective role against invasion of foreign microorganisms. Recent studies have revealed that classical Wnt signaling, which plays a regulatory role in organ tissue development, has immunomodulatory functions. Therefore, it is necessary to study the immunomodulatory effect of the Wnt signaling pathway in lung epithelial cells A549 in anti-Mtb infection. Methods: A549 cells transfected with Wnt signaling plasmid BATflash were stimulated with BCG (bovine Mycobacterium tuberculosis) bacilli. Dual-luciferase reporter assay, Western blot and ELISA were used to analyze the expression of major signal pathways related to Wnt signaling pathway and Changes in immune response to inflammatory cytokines. Results: Wnt signaling inhibited luciferase activity after BCG infection in A549 cells. Western blot results showed that the expression of GSK3β and Axin2 were up-regulated in the cytoplasm with the increase of phosphorylated β-catenin in the cytoplasm, The expression of the active components of β-catenin gene and the downstream transcription factors TCF4 and Lef-1 were down-regulated. The expression of IL-6, NF-κB, MyD88 and TRAF6 was down-regulated when BCG was overexpressed in β-catenin cells Does not affect the expression of TNF-α. Conclusion: The lung epithelial cells can reduce the immune response by down-regulating the Wnt signaling activity and inhibiting the MyD88 / TRAF6 / NF-κB signaling pathway in the process of anti-MTB infection, so as to protect the host cells against the immunological damage caused by the infection.