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.

中文翻译：

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Circ_0001490/miR-579-3p/FSTL1轴调节分枝杆菌的存活以及结核分枝杆菌感染的巨噬细胞的活力、细胞凋亡和炎症反应

 背景

巨噬细胞在宿主针对分枝杆菌感染的免疫反应中发挥着重要作用，该过程受到多种因素的调节，包括环状RNA（circRNA）。我们打算使用结核分枝杆菌(M.tb) 感染的 THP-1 巨噬细胞探索 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）和蛋白质印迹测定分别测量RNA和蛋白质表达。进行3-(4,5-二甲基噻唑-2-基)-2,5-二苯基溴化四唑(MTT)测定来分析THP-1巨噬细胞的活力。流式细胞术分析THP-1巨噬细胞的凋亡率。进行酶联免疫吸附测定（ELISA）来评估炎症细胞因子的释放。进行菌落形成单位（CFU）测定来分析 M.tb 在 THP-1 巨噬细胞中的存活情况。通过双荧光素酶报告基因测定和 RNA 免疫沉淀 (RIP) 测定验证分子间靶相互作用。

 结果

TB 患者和 M.tb 感染的 THP-1 巨噬细胞的血清样本中 Circ_0001490 表达下调。 Circ_0001490 过表达抑制 M.tb 存活并促进 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，并且 circ_0001490 过表达诱导的 M.tb 感染的 THP-1 巨噬细胞的保护作用在很大程度上被 miR-579-3p 的过表达所推翻。 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 沉默很大程度上颠覆了 M.tb 感染的 THP-1 巨噬细胞中 miR-579-3p 敲低诱导的效应。 Circ_0001490 作为 miR-579-3p 海绵上调 THP-1 巨噬细胞中的 FSTL1。

 结论

总之，我们的结果表明，circ_0001490部分通过调节miR-579-3p/FSTL1轴抑制M.tb存活并促进M.tb感染的THP-1巨噬细胞的活力和炎症反应。