Reconstruction of the mouse extrahepatic biliary tree using primary human extrahepatic cholangiocyte organoids

The treatment of common bile duct (CBD) disorders, such as biliary atresia or ischemic strictures, is restricted by the lack of biliary tissue from healthy donors suitable for surgical reconstruction. Here we report a new method for the isolation and propagation of human cholangiocytes from the extrahepatic biliary tree in the form of extrahepatic cholangiocyte organoids (ECOs) for regenerative medicine applications. The resulting ECOs closely resemble primary cholangiocytes in terms of their transcriptomic profile and functional properties. We explore the regenerative potential of these organoids $\textit{in vivo}$ and demonstrate that ECOs self-organize into bile duct–like tubes expressing biliary markers following transplantation under the kidney capsule of immunocompromised mice. In addition, when seeded on biodegradable scaffolds, ECOs form tissue-like structures retaining biliary characteristics. The resulting bioengineered tissue can reconstruct the gallbladder wall and repair the biliary epithelium following transplantation into a mouse model of injury. Furthermore, bioengineered artificial ducts can replace the native CBD, with no evidence of cholestasis or occlusion of the lumen. In conclusion, ECOs can successfully reconstruct the biliary tree, providing proof of principle for organ regeneration using human primary cholangiocytes expanded $\textit{in vitro}$.This work was funded by ERC starting grant Relieve IMDs (281335; L.V., N.R.F.H.), the Cambridge Hospitals National Institute for Health Research Biomedical Research Centre (L.V., N.R.F.H., S. Sinha., F.S.), the Evelyn Trust (N.H.) and the EU FP7 grant TissuGEN (M.C.D.B.) and was supported in part by the Intramural Research Program of the NIH/NIAID (R.L.G., C.A.R.). F.S. has been supported by an Addenbrooke's Charitable Trust Clinical Research Training Fellowship and a joint MRC–Sparks Clinical Research Training Fellowship. (MR/L016761/1) A.W.J. and A.E.M. acknowledge support from EPSRC (EP/L504920/1) and an Engineering for Clinical Practice Grant from the Department of Engineering, University of Cambridge. J.B. was supported by a BHF Studentship (Grant FS/13/65/30441)

Reconstruction of the mouse extrahepatic biliary tree using primary human extrahepatic cholangiocyte organoids

The treatment of common bile duct (CBD) disorders, such as biliary atresia or ischemic strictures, is restricted by the lack of biliary tissue from healthy donors suitable for surgical reconstruction. Here we report a new method for the isolation and propagation of human cholangiocytes from the extrahepatic biliary tree in the form of extrahepatic cholangiocyte organoids (ECOs) for regenerative medicine applications. The resulting ECOs closely resemble primary cholangiocytes in terms of their transcriptomic profile and functional properties. We explore the regenerative potential of these organoids $\textit{in vivo}$ and demonstrate that ECOs self-organize into bile duct–like tubes expressing biliary markers following transplantation under the kidney capsule of immunocompromised mice. In addition, when seeded on biodegradable scaffolds, ECOs form tissue-like structures retaining biliary characteristics. The resulting bioengineered tissue can reconstruct the gallbladder wall and repair the biliary epithelium following transplantation into a mouse model of injury. Furthermore, bioengineered artificial ducts can replace the native CBD, with no evidence of cholestasis or occlusion of the lumen. In conclusion, ECOs can successfully reconstruct the biliary tree, providing proof of principle for organ regeneration using human primary cholangiocytes expanded $\textit{in vitro}$