Myllys, Maiju Karoliina2021-11-242021-11-242021http://hdl.handle.net/2003/4057310.17877/DE290R-22442Non-alcoholic fatty liver disease (NAFLD) is the most common chronic liver disease in Western countries with an increasing prevalence of approximately 25 %. NAFLD comprises several stages, starting as benign steatosis and progressing to non-alcoholic steatohepatitis (NASH), and in some cases to liver cirrhosis and hepatocellular carcinoma (HCC). Although several emerging therapies are currently in clinical trials, so far there are no approved drugs for treatment of NASH. The overarching goal of this thesis was to investigate the mechanisms of NAFLD stage transitions, and to establish preventive measures for the progression from benign steatosis to NASH. First, a mouse model of NAFLD progression was established by long-term feeding of male C57Bl/6N mice with western-style diet (WD) up to 54 weeks. The disease progression was evaluated time-dependently by biochemical, histopathological, and immunohistochemical analyses as well as by intravital two-photon-based imaging. This analyses revealed six stages in NAFLD progression: (1) benign steatosis, (2) macrophage crown-like structure formation, (3) macropinocytosis of bile, (4) ductular reaction, (5) dedifferentiation and functional shutdown, and (6) tumor nodule formation. The novel finding of this thesis was the identification of stage 3, where a retrograde vesicular uptake of bile from bile canaliculi to hepatocytes led to toxic accumulation of bile acids in the liver tissue, providing a link between NAFLD and cholestasis. The phenomenon was further identified as macropinocytosis by treating the mice with a macropinocytosis-specific inhibitor imipramine. As a result, a single application of imipramine efficiently blocked macropinocytosis in WD-fed mice. Interestingly, a long-term application of imipramine for 8 weeks decreased the bile acid concentrations in the liver tissue and led to significant NAFLD amelioration. Moreover, bile macropinocytosis was also found relevant in human NAFLD patients as detected by the presence of fragments of bile canaliculi within steatotic hepatocytes. In conclusion, an NAFLD mouse model recapitulating the different stages of human NAFLD progression to NASH and eventually to HCC was successfully established. Moreover, a novel mechanism possibly driving NALFD progression was identified as macropinocytosis of bile from bile canaliculi back to hepatocytes.enNAFLDNASHCholestasisImipramineObeticholic acid540570doctoral thesisFettleberkrankheit