Novel Model of Bile Duct Paucity Demonstrates the Critical Role of Yap1 in Biliary Morphogenesis in Development and Regeneration

Abstract

BackgroundYes‐associated protein 1 (Yap1) is a critical regulator of liver embryonic development, regeneration and tumorigenesis. Previous studies have shown that Yap1 regulates bile duct formation and hepatocyte transdifferentiation to cholangiocytes, but the mechanisms by which Yap1 regulates biliary morphogenesis as well as the functions of Yap1 in early embryonic liver development remain unknown.MethodsTo elucidate the role of Yap1 in early liver development, we developed a novel mouse model (Yap1 KO) in which Yap1 is deleted from the foregut endoderm progenitors by E12.5 using the Foxa3 promoter to drive Cre recombinase expression. Here, we comprehensively characterize the Yap1 KO mice through histology, whole organ tissue clearing and confocal microscopy and additional functional techniques.ResultsYap1 KO mice lack Yap1 in the liver from early development. Strong nuclear expression of Yap1 that was seen in hepatoblasts in WT livers at E14.5 was clearly absent in the KO mice. Although Yap1 KO mice were viable and have fully formed liver and digestive organs, they exhibited jaundice and stunted growth. Histological analysis identified a failure of intrahepatic bile duct formation. Yap1 deletion did not block Notch activation as shown by the appearance of Hes1‐positive and Sox9‐positive ductal plate cells in both WT and KO mice. Rather, Yap1 deletion prevented the morphological assembly of a two‐cell layer with a lumen from an asymmetrical single‐layered ductal plate. Surprisingly, KO mice survive long‐term with normalization of serum liver enzymes despite persistent cholestatic injury, suggesting hepatocyte adaptation to injury. While a recent study of Notch‐deficient livers (a model of Alagille syndrome) described complete de novo hepatocyte‐derived generation of bile ducts by 4 months of age, Yap1‐deficient livers fail to regenerate bile ducts by 8 months of age. Liver tissue clearing and 3D whole liver immunofluorescence imaging showed severely abbreviated biliary tree in KO mice relative to WT. Notably, we observe the appearance of a ductular reaction in the hilar region of the liver consisting of Yap1‐positive ducts surrounded by fibrosis and inflammation. Although all hepatocytes remain Yap1‐negative, the extrahepatic biliary tree remains unaffected by the Cre and is Yap1‐positive, suggesting a potential source of Yap1‐positive ductular cells.ConclusionsOur results demonstrate that Yap1 is dispensable for early development of the liver and foregut but is critical for biliary morphogenesis. We further show absence of hepatocyte‐to‐cholangiocyte transdifferentiation in Yap1 KO. Thus, Yap1 is essential for biliary morphogenesis regardless of the cellular origin, whether from hepatoblasts in development or from hepatocytes in liver regeneration.Support or Funding InformationThis work was supported by NIH grants 1R01DK62277, 1R01DK100287, 1R01DK116993 and Endowed Chair for Experimental Pathology to S.P.M, 1P30DK120531‐01 to the Pittsburgh Liver Research Center, and T32EB0010216 to LM.