IL-22 supports long-term expansion of mouse and human hepatocytes

Abstract Background & Aims Hepatocytes have wide applications. Although liver exhibits a remarkable regenerative capacity, expanding hepatocytes in large quantities in vitro without compromising their functions remains a challenging task, presumably due to the absence of requisite environmental signals. Identifying factors enabling long-term expansion of hepatocytes in vitro is essential to advance liver research. Methods Mouse models of hepatectomy and hepatocyte transplantation were used to identify cytokines involved in liver regeneration. Mouse and human hepatocytes were cultured and passaged in IL22 medium to assess their expansion ability. Genetic lineage tracing was applied to track the origin of the expanding cells. Functions of the expanded hepatocytes were evaluated via cell transplantation into Fah-/- mice. The mechanism of IL22-mediated proliferation was investigated via transcriptomic analysis and lineage-specific gene deletion. Results We discovered that IL22 supports long-term expansion (>30 passages, with a theoretical expansion of ∼1030 times within ∼150 days) of mouse hepatocytes in vitro by dedifferentiating hepatocytes into hepatocyte progenitor cells (HPCs), which maintain the capacity of differentiation into mature hepatocytes. Stat3 pathway is critically involved. Two key transcription factors (TFs) downstream of Stat3 pathway, Bhlha15 and Arntl2, govern IL22-induced hepatic dedifferentiation. Expression of these two TFs directly initiates the dedifferentiation of hepatocytes into HPCs, which could be expanded in long-term without the supplement of IL22. IL22 also supports human hepatocyte growth in simple culture condition. Within 30 days, human hepatocytes can be expanded for more than 10,000-fold by dedifferentiation into HPCs, which maintain the capacity of maturation. Conclusions IL22 enables large-scale expansion of both mouse and human hepatocytes in vitro by activating the Stat3-Bhlha15/Arntl2 axis. Our discoveries may support future utilization of IL22 in liver/hepatocyte-related clinical applications. Impact and implications Hepatocytes have wide applications in drug development, disease modeling, and cell therapy. However, expanding hepatocytes in large quantities in vitro without compromising their functions remains a significant challenge. We demonstrate that IL22, a unique cytokine primarily mediating localized epithelial repair and cytoprotection without inducing systemic immune activation, enables large-scale expansion of both mice and human hepatocytes in vitro by activating the Stat3-Bhlha15/Arntl2 axis. Our findings not only bridge the gap between liver regeneration in vivo and hepatocyte growth in vitro, but also support the future utilization of IL22 in liver/hepatocyte-related clinical applications.