Biliary organoids uncover delayed epithelial development and barrier function in biliary atresia

• Published in: Hepatology (Baltimore, Md.) Vol. 75; no. 1; pp. 89 - 103
• Main Authors: Amarachintha, Surya P., Mourya, Reena, Ayabe, Hiroaki, Yang, Li, Luo, Zhenhua, Li, Xiaofeng, Thanekar, Unmesha, Shivakumar, Pranavkumar, Bezerra, Jorge A.
• Format: Journal Article
• Language: English
• Published: United States Wiley Subscription Services, Inc 01-01-2022
• Subjects: Actin; Adolescent; Aquaporin 1; Bile Ducts - cytology; Bile Ducts - growth & development; Bile Ducts - pathology; Biliary atresia; Biliary Atresia - complications; Biliary Atresia - pathology; Biopsy; Case-Control Studies; Catenin; Cells, Cultured; Child; Child, Preschool; Cholestasis; Cholestasis - etiology; Cholestasis - pathology; Cilia; Cystic fibrosis; Cytokeratin; Cytology; Epithelial Cells - cytology; Epithelial Cells - pathology; Etiology; Ezrin; Fibroblast growth factor 2; Healthy Volunteers; Hepatocyte nuclear factor 1; Hepatology; Homeobox; Humans; Infant; Infant, Newborn; Inflammation; Liver diseases; Localization; Neonates; Organoids - pathology; Permeability; Polarity; Primary Cell Culture; Rhodamine; Somatostatin; Tight Junctions - pathology; Verapamil
• ISSN: 0270-9139; 1527-3350
• DOI: 10.1002/hep.32107

Background and Aims
Biliary atresia is a severe inflammatory and fibrosing cholangiopathy of neonates of unknown etiology. The onset of cholestasis at birth implies a prenatal onset of liver dysfunction. Our aim was to investigate the mechanisms linked to abnormal cholangiocyte development.
Approach and Results
We generated biliary organoids from liver biopsies of infants with biliary atresia and normal and diseased controls. Organoids emerged from biliary atresia livers and controls and grew as lumen‐containing spheres with an epithelial lining of cytokeratin‐19posalbuminnegSOX17neg cholangiocyte‐like cells. Spheres had similar gross morphology in all three groups and expressed cholangiocyte‐enriched genes. In biliary atresia, cholangiocyte‐like cells lacked a basal positioning of the nucleus, expressed fewer developmental and functional markers, and displayed misorientation of cilia. They aberrantly expressed F‐actin, β‐catenin, and Ezrin, had low signals for the tight junction protein zonula occludens‐1 (ZO‐1), and displayed increased permeability as evidenced by a higher Rhodamine‐123 (R123) signal inside organoids after verapamil treatment. Biliary atresia organoids had decreased expression of genes related to EGF signaling and FGF2 signaling. When treated with EGF+FGF2, biliary atresia organoids expressed differentiation (cytokeratin 7 and hepatocyte nuclear factor 1 homeobox B) and functional (somatostatin receptor 2, cystic fibrosis transmembrane conductance regulator [CFTR], aquaporin 1) markers, restored polarity with improved localization of F‐actin, β‐catenin and ZO‐1, increased CFTR function, and decreased uptake of R123.
Conclusions
Organoids from biliary atresia are viable and have evidence of halted epithelial development. The induction of developmental markers, improved cell‐cell junction, and decreased epithelial permeability by EGF and FGF2 identifies potential strategies to promote epithelial maturation and function.