Distinct Roles of Matrigel Enabled the Production of Expandable Hepatoblast and Polarized Hepatocyte Organoids from Human Embryonic Stem Cells under 3-Dimensional Suspension Conditions.

The liver is essential for a range of metabolic functions, and recent advances in organoid technology have enabled the development of various liver organoids. However, the challenges of replicating the in vivo microenvironment, particularly the extracellular matrix, remain important in liver organoid culture. In this study, we explored the roles of Matrigel, a commonly used extracellular matrix component, in the expansion, polarization, and functional maturation of liver organoids derived from human embryonic stem cells (hESCs) under 3-dimensional suspension conditions. Using multiple assays, we demonstrated that a low concentration of Matrigel supported the efficient expansion of hESC-derived hepatoblast organoids by regulating reactive oxygen species (ROS)-autophagy homeostasis through the inhibition of ROS-AMPK-mTOR-mediated excessive autophagy. Moreover, Matrigel induced the polarization of mature hepatocyte organoids differentiated from hESC-derived expandable hepatoblast organoids via activation of the FAK-ERK-AMPK pathway under 3-dimensional suspension conditions. Our findings underscore the importance of integrin signaling in hepatocyte organoid culture and provide insights for the development of artificial, synthetic, bioactive hydrogels for the large-scale production of hepatocytes for clinical applications.