Liver Organoids From Hepatocytes of Healthy Humans and Non-alcoholic Fatty Liver Disease (NAFLD) Patients Display Multilineage Architecture and can be Used to Develop an In Vitro Model of Steatohepatitis.

Background/aim: Non-alcoholic fatty liver disease (NAFLD) is a global health concern with limited treatment options. The paucity of predictive i n v itro models in preclinical settings seems to be one of the limitations of identifying effective medicines. We therefore aimed to develop an i n v itro model that can display the key hallmarks of NAFLD, such as steatosis, inflammation, and fibrosis.

Methods: An in vitro model of steatohepatitis was developed using organoids prepared from hepatocytes of healthy individuals from a commercial source (HLOs) and the liver tissues collected from needle biopsies of NAFLD patients (HLO_NAFLD) using defined culture conditions. HLOs were treated with palmitic acid for 72 h to develop an i n v itro model of steatohepatitis, while HLO_NAFLD served as a natural model of steatohepatitis. Metformin and saroglitazar were used to validate the liver organoid model of steatohepatitis. Saroglitazar was also evaluated in the high-fat, high-fructose (HF-HF) diet-induced model of NAFLD using C57BL/6 mice to validate the findings from the i n v itro model.

Results: HLOs and HLO_NAFLD exhibited bipotent properties, showing the expression of markers of hepatocytes, ductal cells, and also stem cells. Furthermore, they demonstrated the expression of nonparenchymal cell markers such as stellate cells (CD166) and Kupffer-like cells (CD68 and EMR1). The steatohepatitis models developed using these organoids displayed markers associated with steatosis, inflammation and fibrosis, which were decreased by metformin and saroglitazar.

Conclusion: The in vitro models developed in our lab employing HLOs and HLO_NAFLD display all three key hallmarks of NAFLD: steatosis, inflammation, and fibrosis without the necessity for coculture with other nonparenchymal cells. The implementation of the HLO_NAFLD-based model is also expected to provide a more realistic assessment of test substances to develop therapeutics for NAFLD.