Abstract
An alternative source of livers for transplantation in patients with (genetic) liver diseases and liver failure is needed because liver donors are scarce. HPC-derived hepatocyte-like cells could be one of the options. Because dogs and humans share liver-pathologies and disease-pathways, the dog is considered the best model for human liver
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disease. We can use dogs to test safety and efficacy of a new treatment before considering to apply it in human clinics. We developed the long-term culture system of adult canine hepatic progenitor cells in three-dimensions, called organoids. Canine liver organoids showed characteristics of stem cells, cholangiocytes and hepatocytes, mimicking in vivo liver progenitor cell phenotype, and had potential to differentiate towards functional hepatocytes. Organoids derived from COMMD1-deficient dogs presented a copper excretion defect similar to the in vivo situation in dogs with copper storage disease. Upon transduction with a COMMD1 construct, successful restoration of copper excretion was achieved. Patient-derived liver organoids are a powerful tool to perform disease modeling and demonstrate the feasibility to perform gene correction in liver organoids, a key prerequisite for autologous transplantation for gene-therapy. Upon liver injury either hepatocytes themselves or progenitor cells proliferate to regenerate the liver. However, in many virus-induced liver diseases and cancers the hepatocytes harbor mutations in the tumor suppressor genes, Rb and p53. Therefore the regenerative response may differ from regeneration under normal situations. We analyzed the cooperative role of Rb and p53 in accordance with the 3, 5-diethoxycarbonyl-1, 4-dihydrocollidine (DDC) diet, a xenobiotic inducing hepatobiliary injury and oval cell proliferation. DDC is metabolized by Cyp3a. Deletion of both Rb and p53 deregulates Cyp3a activity causing less porphyrin accumulation, therefore mild oval cell proliferation is induced. In addition, Rb/p53 deficient livers experience less biliary injury, but instead more hepatocellular injury under DDC exposure. Finally, inactivation of Rb/p53 shortens life span and accelerates HCC development after DDC exposure. Rb and p53 are not only important cell cycle regulators, but also influence xenobiotic metabolism of cells. Liver resection and radiofrequency ablation (RFA) are the treatment of choice for liver cancers. Yet, cancer recurrence rate after resection is high. We focused on identifying the mechanism of liver tumor formation after surgical interventions. Partial hepatectomy and RFA were performed in mice with p53 and Rb-specific deletion in the livers. Cancer developed at the necrotic injury site, and cancer initiation was associated with migration, expansion, and transformation of bile duct cells within the injury site. This resulted in the formation of undifferentiated carcinomas. A necrotic environment induced by surgery combined with loss of tumor suppressor genes causes liver cancer and epithelial-mesenchymal transition at the surgery site. Bile duct cells or HPCs are the cell-of-origin of liver tumors in this model. Polyploidization is a normal physiological event for livers. Atypical E2Fs, especially E2F8, are essential for hepatocyte binucleation and polyploidization. E2F8 inactivation resulted in up-regulation of E2F-target genes promoting DNA replication, DNA repair, mitosis and cytokinesis. Prevention of polyploidization through inactivation of atypical E2Fs did not have any impact on liver differentiation, zonation, metabolism and regeneration.
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