Bile duct regeneration: Characterization of human bile duct derived organoids

K. Burka, M.M.A. Verstegen, H.P. Roest, M. de Wolf, M.J.C. Bijvelds, H. Gehart, J. de Jonge, H.R. de Jonge, J.N.M. IJzermans, L.J.W. van der Laan

Thursday 15 march 2018

11:30 - 11:40h at Van Weelde Zaal

Categories: Basic / translational research, Session (parallel)

Parallel session: Parallel session 2: Basic / translational research

Integrity of the biliary tree is imperative for liver function. During and after liver transplantation the biliary tree is often damaged by ischemia and may result in graft loss. Evidence suggests that the biliary tree harbors stem cells which contribute to bile duct homeostasis and repair during disease and after transplantation. The aim of this study is to expand and characterize biliary stem cells using 3-dimensional cultures of human bile duct organoids.

Human extra-hepatic bile ducts (n=32) were collected from donor liver grafts or explant patient livers at time of liver transplantation. Biliary organoid cultures were initiated using similar conditions as described for human liver biopsies (Huch et al., 2015, Cell) and propagated by weekly passaging for over 6 months. RNA expression analysis (gene array and q-PCR) and immunohistochemistry was performed. Transporter channel function was measured using Ussing chamber technology and Forskolin Induced Swelling assays. In addition, the hepatocyte differentiation potential of biliary stem cells was studied.

Organoids were efficiently grown from the common bile duct for many passages (>6 months) and compared to liver parenchyma-derived organoids. As expected, bile duct organoids stain positive for biliary cell markers CK19, EpCAM and MUC1. gene expression analysis showed that bile duct organoids are positive for adult stem cell markers LGR5 and Sox9 but also have some district gene expression profile compared to conventional liver organoids. Bile duct organoids are able to differentiate towards cholangiocyte cell phenotype. However, bile duct organoids showed less hepatic differentiation capacity compared to liver organoids. Further results suggest the presence of functional transport channels in the biliary organoids, including CFTR, as they were responsive to forskolin, vasoactive intestinal peptide and bicarbonate.

This study demonstrates the presence of LGR5-positive stem/progenitor cells in human extrahepatic bile duct which can be expanded long-term as organoids in cultures. In the future, these biliary organoids can potentially be used for bile duct regeneration in damaged grafts prior to liver transplantation.