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Nat Protoc. 2019 Jun;14(6):1884-1925. doi: 10.1038/s41596-019-0168-0. Epub 2019 May 20.

Isolation and propagation of primary human cholangiocyte organoids for the generation of bioengineered biliary tissue.

Author information

1
Wellcome Trust-Medical Research Council Stem Cell Institute, Cambridge Stem Cell Institute, Anne McLaren Laboratory, Department of Surgery, University of Cambridge, Cambridge, UK.
2
Department of Surgery, University of Cambridge, Cambridge, UK.
3
Department of Engineering, University of Cambridge, Cambridge, UK.
4
Department of Medical Biotechnology and Translational Medicine (BIOMETRA), Università degli Studi di Milano, Milan, Italy.
5
Norwegian PSC Research Center, Department of Transplantation Medicine, Division of Surgery, Inflammatory Diseases and Transplantation, Oslo University Hospital, Rikshospitalet, Oslo, Norway.
6
Research Institute of Internal Medicine, Division of Surgery, Inflammatory Diseases and Transplantation, Oslo University Hospital, Rikshospitalet, Oslo, Norway.
7
Department of Medicine III, University Hospital Aachen, Aachen, Germany.
8
Section for Gastroenterology, Department of Transplantation Medicine, Division of Surgery, Inflammatory Diseases and Transplantation, Oslo University Hospital Rikshospitalet, Oslo, Norway.
9
Institute of Clinical Medicine, Faculty of Medicine, University of Oslo, Oslo, Norway.
10
Wellcome Trust-Medical Research Council Stem Cell Institute, Cambridge Stem Cell Institute, Anne McLaren Laboratory, Department of Surgery, University of Cambridge, Cambridge, UK. lv225@cam.ac.uk.
11
Department of Surgery, University of Cambridge, Cambridge, UK. lv225@cam.ac.uk.
12
Wellcome Trust-Medical Research Council Stem Cell Institute, Cambridge Stem Cell Institute, Anne McLaren Laboratory, Department of Surgery, University of Cambridge, Cambridge, UK. fs347@cam.ac.uk.
13
Department of Surgery, University of Cambridge, Cambridge, UK. fs347@cam.ac.uk.
14
Department of Hepatology, Cambridge University Hospitals NHS Foundation Trust, Cambridge, UK. fs347@cam.ac.uk.
15
Department of Medicine, University of Cambridge, Cambridge, UK. fs347@cam.ac.uk.

Abstract

Pediatric liver transplantation is often required as a consequence of biliary disorders because of the lack of alternative treatments for repairing or replacing damaged bile ducts. To address the lack of availability of pediatric livers suitable for transplantation, we developed a protocol for generating bioengineered biliary tissue suitable for biliary reconstruction. Our platform allows the derivation of cholangiocyte organoids (COs) expressing key biliary markers and retaining functions of primary extra- or intrahepatic duct cholangiocytes within 2 weeks of isolation. COs are subsequently seeded on polyglycolic acid (PGA) scaffolds or densified collagen constructs for 4 weeks to generate bioengineered tissue retaining biliary characteristics. Expertise in organoid culture and tissue engineering is desirable for optimal results. COs correspond to mature functional cholangiocytes, differentiating our method from alternative organoid systems currently available that propagate adult stem cells. Consequently, COs provide a unique platform for studies in biliary physiology and pathophysiology, and the resulting bioengineered tissue has broad applications for regenerative medicine and cholangiopathies.

PMID:
31110298
DOI:
10.1038/s41596-019-0168-0
[Indexed for MEDLINE]

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