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Human iPSC-derived hepatocyte system models cholestasis with tight junction protein 2 deficiency

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Chao Zheng Li, Hiromi Ogawa, Soon Seng Ng, Xindi Chen, Eriko Kishimoto, Kokoro Sakabe, Aiko Fukami, Yueh Chiang Hu, Christopher N. Mayhew, Jennifer Hellmann, Alexander Miethke, Nahrin L. Tasnova, Samuel J.I. Blackford, Zu Ming Tang, Adam M. Syanda, Liang Ma, Fang Xiao, Melissa Sambrotta, Oliver Tavabie, Filipa Soares & 6 more Oliver Baker, Davide Danovi, Hisamitsu Hayashi, Richard J. Thompson, S. Tamir Rashid, Akihiro Asai

Original languageEnglish
Article number100446
JournalJHEP Reports
Volume4
Issue number4
Early online date1 Feb 2022
DOIs
E-pub ahead of print1 Feb 2022
PublishedApr 2022

Bibliographical note

Funding Information: AA was supported by NIH grant P30 DK078392 , Pilot & Feasibility Award (Sanger Sequencing, Research Pathology, Live Microscopy at the University of Cincinnati, Confocal Imaging, Transgenic Animal and Genome Editing, and Pluripotent Stem Cell Facility cores) of the Digestive Diseases Research Core Center in Cincinnati , the AASLD Foundation (Pinnacle Research Award), North American Society of Gastroenterology Hepatology and Nutrition (NASPGHAN) Foundation (George Ferry Young Investigator Award), Cincinnati Children’s Research Foundation (Procter Scholar Award), STR was supported by Medical Research Council (MRC) Clinician Scientist Fellowship Award ( MR/L006537/1 ), UK. We thank PerkinElmer (UK) for supporting ZMT within the framework of the PE OneSource scheme and the Kings Medical Research Trust for supporting CZL with a studentship award. Publisher Copyright: © 2022 The Author(s)

King's Authors

Abstract

Background & Aims: The truncating mutations in tight junction protein 2 (TJP2) cause progressive cholestasis, liver failure, and hepatocyte carcinogenesis. Due to the lack of effective model systems, there are no targeted medications for the liver pathology with TJP2 deficiency. We leveraged the technologies of patient-specific induced pluripotent stem cells (iPSC) and CRISPR genome-editing, and we aim to establish a disease model which recapitulates phenotypes of patients with TJP2 deficiency. Methods: We differentiated iPSC to hepatocyte-like cells (iHep) on the Transwell membrane in a polarized monolayer. Immunofluorescent staining of polarity markers was detected by a confocal microscope. The epithelial barrier function and bile acid transport of bile canaliculi were quantified between the two chambers of Transwell. The morphology of bile canaliculi was measured in iHep cultured in the Matrigel sandwich system using a fluorescent probe and live-confocal imaging. Results: The iHep differentiated from iPSC with TJP2 mutations exhibited intracellular inclusions of disrupted apical membrane structures, distorted canalicular networks, altered distribution of apical and basolateral markers/transporters. The directional bile acid transport of bile canaliculi was compromised in the mutant hepatocytes, resembling the disease phenotypes observed in the liver of patients. Conclusions: Our iPSC-derived in vitro hepatocyte system revealed canalicular membrane disruption in TJP2 deficient hepatocytes and demonstrated the ability to model cholestatic disease with TJP2 deficiency to serve as a platform for further pathophysiologic study and drug discovery. Lay summary: We investigated a genetic liver disease, progressive familial intrahepatic cholestasis (PFIC), which causes severe liver disease in newborns and infants due to a lack of gene called TJP2. By using cutting-edge stem cell technology and genome editing methods, we established a novel disease modeling system in cell culture experiments. Our experiments demonstrated that the lack of TJP2 induced abnormal cell polarity and disrupted bile acid transport. These findings will lead to the subsequent investigation to further understand disease mechanisms and develop an effective treatment.

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