A subset of hepatic fibroblasts marked by Gli1 expression generates portal fibrosis and promotes ductular reaction

BACKGROUND & AIMS: Gli1+ cells were previously described as myofibroblast precursors. We aimed to delineate the identity of Gli1+ cells and their contribution to wound healing in liver. METHODS: Gli1+ cells were analyzed in normal mouse liver by single-cell RNA sequencing (n = 3). Genetic cell fate tracing and depletion of Gli1+ cells were achieved in male mice, undergoing cholestatic injury induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) or bile duct ligation (BDL), and metabolic dysfunction-associated steatohepatitis (MASH). Gli1 expression was examined by fluorescence in-situ hybridization in human liver (n = 16). Cell interactions were assessed using organoids. RESULTS: In normal liver, Gli1 demarcated a periductal subpopulation (11.8%) of portal fibroblasts, which transcriptome pertains to extracellular matrix and structure organization. In liver injury, Gli1+ fibroblasts accumulated with fibrosis as multilayers around bile ducts and discontinuously along the inner border of the ductular reaction, predominantly in biliary models and less markedly in MASH. In humans, periductal Gli1+ cells were present in normal liver and accumulated in primary sclerosing cholangitis and, to a lesser extent, in MASH, notably alongside the ductular reaction in late-stage diseases. Co-culture of cholangiocyte organoids with Gli1+ fibroblasts showed bidirectional interactions fostering proliferation of both cell types. Genetic depletion of Gli1+ cells (n ≥4/group) caused a marked reduction of fibrosis, assessed by Sirius Red area, in DDC-fed and BDL mice after 1 week (p