Dataset Information

Human extrahepatic and intrahepatic cholangiocyte organoids show region-specific differentiation potential and model cystic fibrosis-related bile duct disease.

ABSTRACT: The development, homeostasis, and repair of intrahepatic and extrahepatic bile ducts are thought to involve distinct mechanisms including proliferation and maturation of cholangiocyte and progenitor cells. This study aimed to characterize human extrahepatic cholangiocyte organoids (ECO) using canonical Wnt-stimulated culture medium previously developed for intrahepatic cholangiocyte organoids (ICO). Paired ECO and ICO were derived from common bile duct and liver tissue, respectively. Characterization showed both organoid types were highly similar, though some differences in size and gene expression were observed. Both ECO and ICO have cholangiocyte fate differentiation capacity. However, unlike ICO, ECO lack the potential for differentiation towards a hepatocyte-like fate. Importantly, ECO derived from a cystic fibrosis patient showed no CFTR channel activity but normal chloride channel and MDR1 transporter activity. In conclusion, this study shows that ECO and ICO have distinct lineage fate and that ECO provide a competent model to study extrahepatic bile duct diseases like cystic fibrosis.

SUBMITTER: Verstegen MMA

PROVIDER: S-EPMC7736890 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Human extrahepatic and intrahepatic cholangiocyte organoids show region-specific differentiation potential and model cystic fibrosis-related bile duct disease.

Verstegen Monique M A MMA Roos Floris J M FJM Burka Ksenia K Gehart Helmuth H Jager Myrthe M de Wolf Maaike M Bijvelds Marcel J C MJC de Jonge Hugo R HR Ardisasmita Arif I AI van Huizen Nick A NA Roest Henk P HP de Jonge Jeroen J Koch Michael M Pampaloni Francesco F Fuchs Sabine A SA Schene Imre F IF Luider Theo M TM van der Doef Hubert P J HPJ Bodewes Frank A J A FAJA de Kleine Ruben H J RHJ Spee Bart B Kremers Gert-Jan GJ Clevers Hans H IJzermans Jan N M JNM Cuppen Edwin E van der Laan Luc J W LJW

Scientific reports 20201214 1

The development, homeostasis, and repair of intrahepatic and extrahepatic bile ducts are thought to involve distinct mechanisms including proliferation and maturation of cholangiocyte and progenitor cells. This study aimed to characterize human extrahepatic cholangiocyte organoids (ECO) using canonical Wnt-stimulated culture medium previously developed for intrahepatic cholangiocyte organoids (ICO). Paired ECO and ICO were derived from common bile duct and liver tissue, respectively. Characteriz ...[more]

PMID: 33318612

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Project description:Cholangiopathies, which affect extrahepatic bile ducts (EHBDs), include biliary atresia, primary sclerosing cholangitis, and cholangiocarcinoma. They have no effective therapeutic options. Tools to study EHBD are very limited. Our purpose was to develop an organ-specific, versatile, adult stem cell-derived, preclinical cholangiocyte model that can be easily generated from wild type and genetically engineered mice. Thus, we report on the novel technique of developing an EHBD organoid (EHBDO) culture system from adult mouse EHBDs. The model is cost-efficient, able to be readily analyzed, and has multiple downstream applications. Specifically, we describe the methodology of mouse EHBD isolation and single cell dissociation, organoid culture initiation, propagation, and long-term maintenance and storage. This manuscript also describes EHBDO processing for immunohistochemistry, fluorescent microscopy, and mRNA abundance quantitation by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR). This protocol has significant advantages in addition to producing EHBD-specific organoids. The use of a conditioned medium from L-WRN cells significantly reduces the cost of this model. The use of mouse EHBDs provides almost unlimited tissue for culture generation, unlike human tissue. Generated mouse EHBDOs contain a pure population of epithelial cells with markers of endodermal progenitor and differentiated biliary cells. Cultured organoids maintain homogenous morphology through multiple passages and can be recovered after a long-term storage period in liquid nitrogen. The model allows for the study of biliary progenitor cell proliferation, can be manipulated pharmacologically, and may be generated from genetically engineered mice. Future studies are needed to optimize culture conditions in order to increase plating efficiency, evaluate functional cell maturity, and direct cell differentiation. Development of co-culture models and a more biologically neutral extracellular matrix are also desirable.

Dataset Information

Human extrahepatic and intrahepatic cholangiocyte organoids show region-specific differentiation potential and model cystic fibrosis-related bile duct disease.

Publications

Human extrahepatic and intrahepatic cholangiocyte organoids show region-specific differentiation potential and model cystic fibrosis-related bile duct disease.

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