HSC-derived exosomal miR-122- 5p inhibits EMT and fibrosis of intrahepatic biliary epithelial cells to alleviate primary biliary cholangitis

Objective: The objective was to study the expression profile of serum exosomal ncRNAs in patients with PBC and to explore their potential molecular mechanisms. Methods: The expression profile of ncRNAs in serum exosomes of PBC patients was analyzed by RNA-sequence. Bioinformatics analysis predicted target genes and related signaling pathways. Spearson correlation analysis and receiver operating characteristic (ROC) curve were used to explore the relationship between ncRNAs and PBC clinical indicators and application value. Primary human intrahepatic biliary epithelial cells (HIBECs) were cultured in vitro and lipopolysaccharide (LPS) induced HIBECs inflammatory injury model. The expression of miR-122-5p is regulated by miR-122-5p mimics and inhibitors. Enzyme linked immunosorbent assay (Elisa) measured levels of inflammatory cytokines. HIBECs cycle and apoptosis were detected by flow cytometry and HIBECs activity was detected by CCK8. The biomarkers of epithelium-mesenchymal transition (EMT), fibrogenesis, and p38 mitogen-activated protein kinase (MAPK) pathway were detected by real time quantitative polymerase chain reaction (RT-qPCR) and Western blot (WB). Dual luciferase reporter assay to confirm the binding of miR-122-5p. Results: 8531 differential expressed (DE) lncRNAs and 263 DE miRNAs were identified in serum exosomes of PBC patients. RNA-sequence and verification results showed that miR-122-5p was increased in serum exosomes of PBC patients, and was positively correlated with liver injury and cholestasis indexes. Exosomal miR-122-5p combined with gp210 and sp100 could improve the sensitivity of PBC diagnosis. In vitro experiments showed that the high expression of miR-122-5p promoted the proliferation of HIBECs, and inhibited the apoptosis, EMT and fibrosis of HIBEC. TNFRSF19, p38, p-p38, and inflammatory cytokines interleukin (IL)-1β, IL-6, IL-8, IL-12, IL-17, tumor necrosis factor (TNF)-α, transforming growth factor (TGF)-β1, and interferon (IFN)-γ were down-regulated and the effect was largely reversed after blocking p38 MAPK pathway. In addition, TNFRSF19 has binding site with miR-122-5p, which is the target gene of miR-122-5p, and could regulate the level of apoptosis signal-regulated kinase 1 (ASK1). Then ASK1 further regulate the downstream p38 MAPK signaling pathway. Conclusion: MiR-122-5p regulates ASK1 by targeting TNFRSF19 and plays an important role in EMT and fibrosis of PBC through p38 MAPK signaling pathway. MiR-122-5p may be a new marker for the diagnosis of PBC and a potential therapeutic target.