Downregulation of microRNA‐9 reduces inflammatory response and fibroblast proliferation in mice with idiopathic pulmonary fibrosis through the ANO1‐mediated TGF‐β–Smad3 pathway

Abstract

Idiopathic pulmonary fibrosis (IPF) is a progressive interstitial lung disease with increasing occurrence, high death rates and unfavorable treatment regimens. In the current study, we identified the expression of microRNA-9 (miR-9) and anoctamin-1 (ANO1) in IPF mouse models induced by bleomycin, and their effects on inflammation and fibroblast proliferation through the transforming growth factor-β (TGF-β)-Smad3 pathway. To verify the targeting relationship between miR-9 and ANO1, we used bioinformatics prediction and conducted a dual-luciferase reporter gene assay. The underlying regulatory mechanisms of miR-9 and the target gene ANO1 were investigated mainly with the treatment of miR-9 mimic, miR-9 inhibitor, or siRNA against ANO1 in fibroblasts isolated from IPF mice. Enzyme-linked immunosorbent assay was performed to investigate the effect of miR-9 or ANO1 on inflammatory factors. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and flow cytometry were used to detect fibroblast proliferation and apoptosis. Reverse transcription quantitative polymerase chain reaction and western blot analysis were applied to measure the expression of the TGF-β-Smad3 pathway-related genes. The determination of luciferase activity suggested that miR-9 targets ANO1. Upregulation of miR-9 or silencing of ANO1 intensified inflammation in IPF, promoted proliferation and inhibited apoptotic ability of lung fibroblasts. MiR-9 negatively modulated ANO1, and thus activated the TGF-β-Smad3 pathway. These findings suggest that miR-9 can indirectly activate the TGF-β-Smad3 pathway by inhibiting the expression of ANO1, thereby aggravating inflammation, promotes proliferation and suppressing apoptosis of lung fibroblasts in mice models of IPF.

Potential function and molecular mechanism of circRNAs involved in idiopathic pulmonary fibrosis

“…is involved in the pathogenesis of IPF [30,31], and increased expression of IGFBP5 and ITGB1 plays a vital role in the development of IPF [32,33]. Our present results suggested that up-regulated hsa_circ_0004099 might promote the development of IPF by decreasing the expression of miR-4633 or miR-9, and further increasing the expression of IGFBP5 or ITGB1, respectively.…”

Detection of microRNA expression levels based on microarray analysis for classification of idiopathic pulmonary fibrosis

“…Additionally, miR-101 attenuates IPF by inhibiting SMAD3, thus reducing transforming growth factor (TGF)-β expression and inhibiting fibroblast proliferation and activation ( 8 ). Other previous studies have shown that the downregulation of miR-9 targets anoctamin-1, which results in decreased expression levels of TGF-β and SMAD3, slowed progression of IPF and reduced fibroblast proliferation in bleomycin-induced mice ( 9 , 10 ).…”

Tumor genotype, location, and malignant potential shape the immunogenicity of primary untreated gastrointestinal stromal tumors

“…Recent evidence also implicates this molecule in chemokine signaling (41). In particular, anoctamin-1 has been shown to suppress the release of proinflammatory cytokines, thus hindering the innate immune response (66,67). Accordingly, preliminary data suggest that ANO1 silencing in GIST cells alters the expression of genes involved in immune-related pathways.…”