MicroRNA-9a-5p inhibits mucosal barrier injury in inflammatory bowel disease.

Abstract

Our previous research found that FOXO1 aggravates the mucosal barrier injury in inflammatory bowel disease (IBD) by regulating TLR4/MD2 signaling. In this study, we further reveal the mechanism of action whereby miRNA-9a-5p inhibits the mucosal barrier injury after regulating FOXO1. An IBD model was established in C57BL/6N mice using dextran sulfate sodium (DSS). The effects of endogenous miRNA-9a-5p were mimicked/antagonized by intraperitoneally injecting miRNA-9a-5p agomir and antagomir. Body weights of mice were monitored and the disease activity indexscores were assessed. H&E staining was performed to examine pathological changes, while immunohistochemical (IHC) staining was conducted to measure the expressions of TJ proteins (ZO-1, Occludin), as well as FOXO1 and TLR4. The mucosal permeability was assessed by FITC-D, the tissue inflammatory cytokines were detected by enzyme linked immunosorbent assay, and the expressions of ZO-1 and Occludin were measured through Western blot analysis. Caco-2 cells were cultured in vitro to establish a monolayer model of the mucosal barrier. TNF-α was used to induce the cell damage, while agomir and antagomir were transfected to mimic/antagonize the miRNA-9a-5p action, followed by determination of barrier permeability. There was a targeted regulatory relationship between MiRNA-9a-5p and FOXO1. MiRNA-9a-5p could suppress the FOXO1 expression, thereby downregulating the TLR4 signaling activation, inhibiting the mucosal barrier injury, and elevating the expressions of TJ proteins. We also found in Caco-2 cells that miRNA-9a-5p could protect cells from inflammatory injury and reduce permeability. In rescue experiments, the effect of agomir was found inhibited by the overexpression of FOXO1 in agomir-treated cells. This study found that miRNA-9a-5p could inhibit the TLR4 signaling activation by targeting FOXO1, thereby exerting a protective effect on the mucosal barrier injury in IBD.