Cholangiocyte organoids from human bile retain a local phenotype and can repopulate bile ducts in vitro

摘要

The well‐established 3D organoid culture method enabled efficient expansion of cholangiocyte‐like cells from intrahepatic (IHBD) and extrahepatic bile duct (EHBD) tissue biopsies. The extensive expansion capacity of these organoids enables various applications, from cholangiocyte disease modelling to bile duct tissue engineering. Recent research demonstrated the feasibility of culturing cholangiocyte organoids from bile, which was minimal‐invasive collected via endoscopic retrograde pancreaticography (ERCP). However, a detailed analysis of these bile cholangiocyte organoids (BCOs) and the cellular region of origin was not yet demonstrated. In this study, we characterize BCOs and mirror them to the already established organoids initiated from IHBD‐ and EHBD‐tissue. We demonstrate successful organoid‐initiation from extrahepatic bile collected from gallbladder after resection and by ERCP or percutaneous transhepatic cholangiopathy from a variety of patients. BCOs initiated from these three sources of bile all show features similar to in vivo cholangiocytes. The regional‐specific characteristics of the BCOs are reflected by the exclusive expression of regional common bile duct genes ( HOXB2 and HOXB3 ) by ERCP‐derived BCOs and gallbladder‐derived BCOs expressing gallbladder‐specific genes. Moreover, BCOs have limited hepatocyte‐fate differentiation potential compared to intrahepatic cholangiocyte organoids. These results indicate that organoid‐initiating cells in bile are likely of local (extrahepatic) origin and are not of intrahepatic origin. Regarding the functionality of organoid initiating cells in bile, we demonstrate that BCOs efficiently repopulate decellularized EHBD scaffolds and restore the monolayer of cholangiocyte‐like cells in vitro. Bile samples obtained through minimally invasive procedures provide a safe and effective alternative source of cholangiocyte organoids. The shedding of (organoid‐initiating) cholangiocytes in bile provides a convenient source of organoids for regenerative medicine. Lay summary : Cholangiocyte organoids provide a powerful tool for characterizing bile duct epithelium and expanding cholangiocytes for tissue engineering purposes. However, organoid initiation typically requires invasive tissue‐biopsies obtained via surgical procedures associated with complications. To overcome this limitation, organoid cultures were initiated from minimal invasive bile‐samples obtained during routine clinical procedures. Characterization revealed that these bile‐cholangiocyte organoids (BCOs) originate from extrahepatic biliary tissue and are capable of repopulating human extrahepatic bile duct scaffolds. Upon repopulation of scaffolds in vitro, the cells obtain a transcriptomic profile more closely resembling primary cholangiocytes. By using bile as novel source for extrahepatic cholangiocyte organoids, bile duct tissue engineering as well as personalized disease modelling become feasible.