Long-Term Expansion of Murine Primary Hepatocyte Organoids

The in vitro maintenance and expansion of primary hepatocytes provide immense opportunities for disease modeling and other biological, viral, and toxicological studies, as well as for applications in regenerative medicine such as cell transplantation for the treatment of metabolic liver diseases. Here, we describe a protocol for the isolation and in vitro culture of primary hepatocytes in a three-dimensional extracellular matrix gel. The inflammatory cytokine tumor necrosis factor alpha (TNFα) is crucial for the long-term expansion of mouse hepatocyte organoids by mimicking the regenerative response in vitro. Long-term cultured hepatocyte organoids express high levels of hepatocyte markers upon differentiation.