Identification and characterization of a biliary atresia hepatic progenitor pool from an organoid-transcriptomic approach

Abstract

Biliary atresia (BA) is the commonest cause of paediatric liver transplantation (>50%). Kasai hepatoportoenterostomy (HPE) is a universal surgical treatment when a diagnosis is made early. Yet, the disease continues to progress, leading to a series of complications and need of liver transplantation in 60% of patients. Clinical research on BA investigated into the clinical variants of BA, age of performing HPE for better success, neonatal hepatobiliary biochemical profiles and the effect of using post-HPE adjuvant steroid therapy on clinical outcome. While clinical research shed insights on the clinical features, course and outcome of BA, basic research is critical to uncovering the underlying pathogenic mechanisms based on which new intervention can be proposed to improve the dismal outlook of BA. Rhesus rotavirus-induced BA mouse model, animal outbreak of BA, zebrafish-biliatresone model, induced pluripotent stem cells (iPSCs), genetics studies and pathological human tissue immunohistochemistry analyses were used to suggest hypotheses of BA pathogenesis, including viral, environmental toxin, developmental immaturity, genetic predisposition, immune dysregulation and fibrogenic ductular reaction. Discrepancies exist between human and animal models. Questions remain regarding how close animal models are recapitulating the actual human pathology. For example, multiple viruses and inconsistencies were identified on human tissue and BA epidemic iii has not been observed in humans. While iPSCs studies rely on genetic manipulation and access to human biliary material for cell culture studies is limited due to the lack of cell ability to proliferate, there is a gap between basic research and their reflection on human BA pathology. A key and interesting question to a paediatric surgeon and the patients' family is: Is the palliative biliary fibrosis outcome preventable at the time of Kasai HPE? This thesis details a novel approach-combining human liver biopsy-derived diseased organoids-in-the-dish and transcriptomic analyses-addressing the first step of this question. It identified and characterized a group of aberrant hepatic progenitors in paediatric BA livers at the time of Kasai HPE. The attempt made use of an optimized biliary organoid culture protocol and a series of next-generation sequencing (transcriptomics) analyses to identify an aberrant group of hepatocytes with progenitor properties that are non-existent in non-BA human livers. At the end, a long-overlooked perspective on the hepatocyte populations in the context of BA-a biliary disease, was revisited and suggested as a modern research direction. Such finding opens two key possibilities: first, human biliary cells from diseased tissue are not available in abundance to support further basic research into the pathogenic mechanisms; second, the same approach could be utilized on diseased tissues of different ethnicities, clinical background and disease stages in future to capture the distinctions among them.