Mesenchymal stem cell-derived extracellular vesicles attenuate periductal fibrosis by inhibiting Th17 differentiation in human liver multilineage organoids and Mdr2-/- mice.

Primary sclerosing cholangitis (PSC) pathogenesis involves immune dysregulation, genetic factors, and bile duct pathology; however, a comprehensive pathogenesis model and effective therapeutic strategies remain limited. Here, we develop a novel human liver multilineage organoid (Mulorg) model combined with Mdr2^-/- mice to investigate the pro-fibrotic role of T helper 17 cells (Th17) and the therapeutic potential of mesenchymal stem cell-derived extracellular vesicles (EV^MSC) for PSC, particularly periductal fibrosis. EV^MSC alleviates interleukin-17A (IL-17A)-induced fibrotic Mulorgs (FibHOs) and mitigates periductal fibrosis in Mdr2^-/- mice by inhibiting Th17 differentiation, decreasing Th17 numbers, and lowering intrahepatic IL-17A levels. Functional assays, miRNA array, and CUT & Tag analyses reveal that EVs-derived hsa-miR-7977 targets NFKBIZ, repressing IκBζ translation to reduce IL-17A and its downstream targets involved in Th17 differentiation, IL-17 signaling, and bile secretion pathways. Moreover, miR-7977-enriched EV^MSC efficiently reduces IL-17A⁺ cell percentages in fibrotic areas and improves periductal fibrosis in Mdr2^-/- mice. Co-culture of FibHOs with Th17 found miR-7977 inhibits Th17 migration to the periductal fibrosis area, with distinct morphological differences observed between patient- and healthy-derived FibHOs. These findings demonstrate that EV-derived miR-7977 mitigates the periductal fibrosis microenvironment by inhibiting Th17 differentiation and migration, the former by targeting NFKBIZ, regulating IL-17A and IκBζ-targeted gene expression. This study clarifies Th17's role in the PSC fibrotic microenvironment, underscores the modeling contributions of Mulorgs, and highlights EV-derived miR-7977's potential to ameliorate Th17-related periductal fibrosis, offering insights and novel therapeutic avenues for PSC.