Wnt/-catenin and NFB signaling synergize to trigger growth-factor-free regeneration of adult primary human hepatocytes

Background & Aims: The liver has a remarkable capacity to regenerate, which is sustained by the ability of hepatocytes to act as facultative stem cells that, while normally quiescent, re-enter the cell cycle after injury. Growth factor signaling is indispensable in rodents, whereas Wnt/beta-catenin is not required for effective tissue repair. However, the molecular networks that control human liver regeneration remain unclear.Methods: Organotypic 3D spheroid cultures of primary human or murine hepatocytes were used to identify the signaling network underlying cell cycle re-entry. Furthermore, we performed chemogenomic screening of a library enriched for epigenetic regulators and modulators of immune function to determine the importance of epigenomic control for human hepatocyte regeneration.Results: Our results showed that, unlike in rodents, activation of Wnt/beta-catenin signaling is the major mitogenic cue for adult primary human hepatocytes. Furthermore, we identified TGF beta inhibition and inflammatory signaling via NF-kappa B as essential steps for the quiescent-to-regenerative switch that allows Wnt/beta-catenin-induced proliferation of human cells. In contrast, growth factors, but not Wnt/beta-catenin signaling, triggered hyperplasia in murine hepatocytes. High-throughput screening in a human model confirmed the relevance of NF kappa B and revealed the critical roles of polycomb repressive complex 2 (PRC2), as well as of the bromodomain families I, II, and IV.Conclusions: This study revealed a network of NF kappa B, TGF beta, and Wnt/beta-catenin that controls human hepatocyte regeneration in the absence of exogenous growth factors, identified novel regulators of hepatocyte proliferation, and highlighted the potential of organotypic culture systems for chemogenomic interrogation of complex physiological processes.