title: Development and characterization of human-induced pluripotent stem cell-derived cholangiocytes.
abstract: Cholangiocytes are the target of a heterogeneous group of liver diseases known as the cholangiopathies. An evolving understanding of the mechanisms driving biliary development provides the theoretical underpinnings for rational development of induced pluripotent stem cell (iPSC)-derived cholangiocytes (iDCs). Therefore, the aims of this study were to develop an approach to generate iDCs and to fully characterize the cells in vitro and in vivo. Human iPSC lines were generated by forced expression of the Yamanaka pluripotency factors. We then pursued a stepwise differentiation strategy toward iDCs, using precise temporal exposure to key biliary morphogens, and we characterized the cells, using a variety of morphologic, molecular, cell biologic, functional, and in vivo approaches. Morphology shows a stepwise phenotypic change toward an epithelial monolayer. Molecular analysis during differentiation shows appropriate enrichment in markers of iPSC, definitive endoderm, hepatic specification, hepatic progenitors, and ultimately cholangiocytes. Immunostaining, western blotting, and flow cytometry demonstrate enrichment of multiple functionally relevant biliary proteins. RNA sequencing reveals that the transcriptome moves progressively toward that of human cholangiocytes. iDCs generate intracellular calcium signaling in response to ATP, form intact primary cilia, and self-assemble into duct-like structures in three-dimensional culture. In vivo, the cells engraft within mouse liver, following retrograde intrabiliary infusion. In summary, we have developed a novel approach to generate mature cholangiocytes from iPSCs. In addition to providing a model of biliary differentiation, iDCs represent a platform for in vitro disease modeling, pharmacologic testing, and individualized, cell-based, regenerative therapies for the cholangiopathies.
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Thiago M
authors: TM
authors: De Assuncao
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Yan
authors: Y
authors: Sun
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Nidhi
authors: N
authors: Jalan-Sakrikar
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Mary C
authors: MC
authors: Drinane
authors: Center for Basic Research in Digestive Diseases, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Bing Q
authors: BQ
authors: Huang
authors: Division of Biomedical Statistics and Informatics, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Ying
authors: Y
authors: Li
authors: Division of Biomedical Statistics and Informatics, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Jaime I
authors: JI
authors: Davila
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Ruisi
authors: R
authors: Wang
authors: Center for Basic Research in Digestive Diseases, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Steven P
authors: SP
authors: O'Hara
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA [3] Center for Cell Signaling in Gastroenterology, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Gwen A
authors: GA
authors: Lomberk
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA [3] Center for Cell Signaling in Gastroenterology, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Raul A
authors: RA
authors: Urrutia
authors: Department of Molecular Medicine, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Yasuhiro
authors: Y
authors: Ikeda
authors: 1] Division of Gastroenterology and Hepatology, Mayo Clinic and Foundation, Rochester, MN, USA [2] Gastroenterology Research Unit, Mayo Clinic and Foundation, Rochester, MN, USA [3] Center for Cell Signaling in Gastroenterology, Mayo Clinic and Foundation, Rochester, MN, USA.
authors: Robert C
authors: RC
authors: Huebert
mesh_terms: D000818:Animals; D001652:Bile Ducts / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism; D020013:Calcium Signaling; D002454:Cell Differentiation; D060846:Cell Engineering; D002460:Cell Line; D004847:Epithelial Cells / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000737:chemistry / Q000166:cytology* / Q000378:metabolism; D008099:Liver / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D051379:Mice; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D015415:Biomarkers