Bioengineered 3D hPSC‐Cholangiocyte Ducts With Physiological Signals for Biliary Disease Modeling

The progression of intrahepatic biliary diseases remains poorly understood, underscoring the urgent need to develop physiologically relevant human intrahepatic cholangiocyte disease models. Current approaches lack the complexity and throughput to capture how diverse biliary microenvironmental signals shape cholangiocyte behavior. To address this gap, we created a fully epithelialized and perfusable three-dimensional (3D) bile duct from human pluripotent stem cell (hPSC)-derived cholangiocytes characterized by robust primary ciliation, apical-basal polarity and cystic fibrosis transmembrane conductance regulator (CFTR)-mediated chloride conductance. Our results revealed physiologically relevant fluid flow and biliary stroma cells as essential components for sustaining cholangiocyte epithelial barrier integrity and ciliation. From here, we interrogated a broad spectrum of biliary signals to study bile acid toxicity and cytokine-driven injury, offering an unprecedented view into intrahepatic cholangiocyte stress responses and potential pathological mechanisms. By integrating physiologically relevant signals, fidelity and functional resolution, this platform provides the foundation needed to precisely decode pathogenic drivers and accelerate therapeutic development for devastating cholangiopathies.