Biliary YB-1/GLI2 axis facilitates ductular reaction and promotes hepatic stellate cell activation via SPP1/Integrin αvβ1 signaling during liver fibrogenesis

Background & Aims: Emerging evidence suggests that ductular reactive cells (DRCs)-mediated ductular reaction (DR) accelerates the activation of hepatic stellate cells (HSCs) and contributes to liver fibrogenesis. Previous studies implicated Y-box binding protein 1 (YB-1) in promoting DRC expansion. This study aims to investigate the mechanisms underlying YB-1-mediated DR and its role in HSC activation. Approach & Results: YB-1 was highly expressed in DRCs in human injured livers. CK19 CreERT mice were crossed with YB-1 flox/flox mice to generate DRC-specific YB-1 knockout mice. DRC-specific YB-1 deletion attenuated DR and liver injury induced by 3,5-methoxycarbonyl-1,4-dihydrocollidine (DDC) feeding and carbon tetrachloride (CCl 4 ) treatment. Transcriptomic analyses, along with chromatin immunoprecipitation and luciferase assays, revealed that YB-1 transcriptionally regulated GLI2 and promoted DRC proliferation. Pharmacological inhibition of GLI2 significantly attenuated DR and liver fibrosis in DDC and CCl 4 mouse models. Transwell co-culture assay indicated that YB-1/GLI2 axis in DRCs drived HSC activation. Liquid chromatography-mass spectrometry combined with bioinformatic analyses identified secreted phosphoprotein 1 (SPP1) as the key molecule linking YB-1/GLI2-mediated DR to HSC activation. SPP1 was highly expressed in human injured livers and interacted with integrins. DRC-specific YB-1 knockout decreased the co-localization of SPP1 and integrin αvβ1 receptors in mouse fibrotic livers. Blocking integrin αvβ1 receptors in HSCs suppressed their activation, which was induced by DRC-derived SPP1. Conclusions: YB-1/GLI2 axis promotes DRC proliferation and SPP1 secretion, which facilitates HSC activation through integrin αvβ1 receptors. This study highlights the YB-1/GLI2/SPP1 signaling pathway as a potential target for therapeutic intervention in liver fibrosis.