Functional hepatobiliary organoids recapitulate liver development and reveal essential drivers of hepatobiliary cell fate determination.

During liver development, hepatocytes, and cholangiocytes are concurrently differentiated from common liver progenitor cells and are assembled into hepatobiliary architecture to perform proper hepatic function. However, the generation of functional hepatobiliary architecture from hepatocytes in vitro is still challenging, and the exact molecular drivers of hepatobiliary cell lineage determination is largely unknown. In this study, functional hepatobiliary organoids (HBOs) are generated from hepatocytes. These HBOs contain a bile duct network surrounded by mature hepatocytes and stably maintain hepatic characteristics and function in vitro and upon transplantation in vivo. Morphological transition and expression profile of hepatocyte-derived organoids recapitulate the process of liver development. Gene regulation landscape of hepatocyte-derived organoids reveal that Tead4 and Ddit3 promote the cell fate commitment of liver progenitors to functional cholangiocytes and hepatocytes, respectively. Liver cell fate determination is reversed by inhibiting Tead4 or increasing Ddit3 expression both in vitro and upon transplantation in vivo. Collectively, hepatocyte-derived HBOs reveal the essential transcription drivers of liver hepatobiliary cell lineage determination and represent powerful models for liver development and regeneration.