环状 RNA 0001490/miR-579-3p/FSTL1 轴调节分枝杆菌的存活以及分枝杆菌感染的巨噬细胞的活力、凋亡和炎症反应。
Circ_0001490/miR-579-3p/FSTL1 axis modulates the survival of mycobacteria and the viability, apoptosis and inflammatory response in Mycobacterium tuberculosis-infected macrophages.
机构信息
Department of Tuberculosis, Jiangxi Chest Hospital, Nanchang City, Jiangxi Province, China.
Deparment of Respratory and Critical Care Medicine, Jiangxi Chest Hospital, Nanchang City, Jiangxi Province, China.
出版信息
Tuberculosis (Edinb). 2021 Dec;131:102123. doi: 10.1016/j.tube.2021.102123. Epub 2021 Sep 3.
BACKGROUND
Macrophages play an important role in the host immune response against mycobacterial infection, and this process is regulated by various factors, including circular RNAs (circRNAs). We intended to explore the role of circ_0001490 in tuberculosis (TB) using Mycobacterium tuberculosis (M.tb)-infected THP-1 macrophages.
METHODS
Real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot assay were conducted to measure RNA and protein expression, respectively. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was conducted to analyze the viability of THP-1 macrophages. Flow cytometry was performed to analyze the apoptosis rate of THP-1 macrophages. Enzyme-linked immunosorbent assay (ELISA) was conducted to assess the release of inflammatory cytokines. Colony-forming unit (CFU) assay was conducted to analyze the survival of M.tb in THP-1 macrophages. Intermolecular target interaction was verified by dual-luciferase reporter assay and RNA immunoprecipitation (RIP) assay.
RESULTS
Circ_0001490 expression was down-regulated in the serum samples of TB patients and M.tb-infected THP-1 macrophages. Circ_0001490 overexpression suppressed M.tb survival and promoted the viability and inflammatory response of THP-1 macrophages. Circ_0001490 interacted with microRNA-579-3p (miR-579-3p), and circ_0001490 overexpression-induced protective effects in M.tb-infected THP-1 macrophages were largely overturned by the overexpression of miR-579-3p. miR-579-3p interacted with the 3' untranslated region (3'UTR) of follistatin-like protein 1 (FSTL1). FSTL1 silencing largely overturned miR-579-3p knockdown-induced effects in M.tb-infected THP-1 macrophages. Circ_0001490 acted as miR-579-3p sponge to up-regulate FSTL1 in THP-1 macrophages.
CONCLUSION
In conclusion, our results demonstrated that circ_0001490 suppressed M.tb survival and promoted the viability and inflammatory response of M.tb-infected THP-1 macrophages partly by regulating miR-579-3p/FSTL1 axis.
背景
巨噬细胞在宿主对分枝杆菌感染的免疫反应中发挥重要作用，该过程受多种因素调节，包括环状 RNA（circRNA）。我们旨在使用分枝杆菌感染的 THP-1 巨噬细胞来探索 circ_0001490 在结核病（TB）中的作用。
方法
实时定量聚合酶链反应（RT-qPCR）和 Western blot 分析分别用于测量 RNA 和蛋白质表达。3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）分析用于分析 THP-1 巨噬细胞的活力。流式细胞术用于分析 THP-1 巨噬细胞的凋亡率。酶联免疫吸附试验（ELISA）用于评估炎症细胞因子的释放。集落形成单位（CFU）分析用于分析分枝杆菌在 THP-1 巨噬细胞中的存活情况。双荧光素酶报告基因分析和 RNA 免疫沉淀（RIP）分析验证了分子间靶标相互作用。
结果
TB 患者的血清样本和分枝杆菌感染的 THP-1 巨噬细胞中 circ_0001490 的表达下调。circ_0001490 的过表达抑制了分枝杆菌的存活，并促进了 THP-1 巨噬细胞的活力和炎症反应。circ_0001490 与 microRNA-579-3p（miR-579-3p）相互作用，而 circ_0001490 过表达在分枝杆菌感染的 THP-1 巨噬细胞中的保护性作用在 miR-579-3p 的过表达下基本被推翻。miR-579-3p 与卵泡抑素样蛋白 1（FSTL1）的 3'非翻译区（3'UTR）相互作用。FSTL1 的沉默在分枝杆菌感染的 THP-1 巨噬细胞中基本推翻了 miR-579-3p 敲低引起的作用。circ_0001490 作为 miR-579-3p 的海绵，在 THP-1 巨噬细胞中上调 FSTL1。
结论
总之，我们的结果表明，circ_0001490 通过调节 miR-579-3p/FSTL1 轴抑制分枝杆菌的存活并促进分枝杆菌感染的 THP-1 巨噬细胞的活力和炎症反应。
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