Collagen I-rich microenvironments influence cholangiocyte specification and tubulogenesis in stem cell-derived models of liver development

Cholangiocyte dysfunction accounts for a substantial proportion of liver transplantations, yet progress towards regenerative therapies is limited by an incomplete understanding of the developmental cues that regulate human cholangiocyte specification and morphogenesis. Here, we established a defined culture system to investigate how collagen I influences hepatobiliary differentiation from human induced pluripotent stem cells (hiPSCs). Increased collagen I presentation was associated with a bias towards cholangiocyte specification, reduced hepatocyte identity and the formation of branched duct-like tubular structures. Similar effects were observed in primary human foetal hepatoblast organoids, indicating that collagen I-dependent lineage bias is conserved across independent model systems. Mechanistically, collagen I presentation was associated with activation of integrin α2β1- and YAP-associated signalling, while inhibition of collagen-integrin interactions impaired both cholangiocyte specification and tubulogenesis. Together, our findings support a role for collagen I-rich extracellular environments in human hepatobiliary differentiation and provide a defined experimental system for investigating extracellular matrix-dependent mechanisms of bile duct development in vitro.