lncRNA LINC00943通过调节TLR4/NF-κB通路对结核杆菌感染的巨噬细胞活性的机制研究

Mechanism of lncRNA LINC00943 regulating TLR4/NF-κB pathway against macrophage activity in Mycobacterium tuberculosis infection

目的 探讨LINC00943在结核杆菌感染的巨噬细胞自噬中的作用。方法 收集活动性结核病(tuberculosis, TB)患者15例和健康志愿者15例的外周血样本。实时定量反转录聚合酶链式反应检测LINC00943的表达。免疫荧光和透射电子显微镜检测自噬。蛋白质免疫印迹法检测LC3-Ⅰ、 LC3-Ⅱ以及Toll样受体4(Toll-like receptor 4,TLR4)/核因子κB(nuclear factor kappa-B,NF-κB)通路相关蛋白的表达。细胞计数盒8(cell counting kit-8,CCK-8)检测细胞活力;流式细胞术检测细胞凋亡。结果 与健康对照组相比,TB患者的单核细胞凋亡减少、自噬增强。与健康对照的单核细胞相比,TB患者单核细胞中LINC00943、TLR4和MyD88的表达显著上调。结核病患者单核细胞中LINC00943的表达上调。过表达LINC00943可诱导巨噬细胞自噬。LINC00943通过调节TLR4/NF-κB通路调节巨噬细胞自噬。TLR4/NF-κB通路抑制剂处理可以逆转LINC00943对巨噬细胞自噬的促进作用。结论 LINC00943通过调节TLR4/NF-κB通路抑制结核病中的巨噬细胞自噬,这意味着治疗由结核分枝杆菌免疫逃逸引起的活动性肺结核发生的新策略。

Objective To explore the role of macrophage autophagy induced by Mycobacterium tuberculosis(TB)infection.Methods Peripheral blood samples were collected from 15 active TB patients and 15 healthy volunteers respectively.The expression of LINC00943 was detected by real time quantitative reverse transcription polymerase chain reaction(qRT-PCR).Autophagy was detected by immunofluorescence and transmission electron microscopy.Expressions of LC3-Ⅰ,LC3-Ⅱand Toll-like receptor 4(TLR4)/nuclear factor kappa-B(NF-κB)pathway-related proteins were detected by Western blot.Cell viability was detected by cell counting kit-8(CCK-8),and apoptosis was detected by flow cytometry.Results Compared with the healthy control group,monocyte apoptosis was reduced and autophagy was enhanced in TB patients.Compared with the monocytes of healthy controls,the expression of LINC00943,TLR4 and MyD88 in the monocytes of TB patients was significantly upregulated.Overexpression of LINC00943 induced macrophage autophagy,and LINC00943 regulated macrophage autophagy by modulating the TLR4/NF-κB pathway.TLR4/NF-κB pathway inhibitor treatment could reverse the promotion of macrophage autophagy by LINC00943.Conclusion LINC00943 inhibits macrophage autophagy in tuberculosis by modulating the TLR4/NF-κB pathway,suggesting a new strategy for treating the occurrence of active TB caused by immune escape from Mycobacterium tuberculosis.