Modelling fatty liver disease with mouse liver-derived multicellular spheroids

Chronic liver disease can lead to liver fibrosis and ultimately cirrhosis, which is a significant health burden and a major cause of death worldwide. Reliable in vitro models are lacking and thus mono-cultures of cell lines are still used to study liver disease and evaluate candidate anti-fibrotic drugs. We established functional multicellular liver spheroid (MCLS) cultures using primary mouse hepatocytes, hepatic stellate cells, liver sinusoidal endo-thelial cells and Kupffer cells. Cell-aggregation and spheroid formation was enhanced with 96-well U-bottom plates generating over +/- 700 spheroids from one mouse. Extensive characterization showed that MCLS cultures contain functional hepatocytes, quiescent stellate cells, fenestrated sinusoidal endothelium and responsive Kupffer cells that can be maintained for 17 days. MCLS cultures display a fibrotic response upon chronic exposure to acetaminophen, and present steatosis and fibrosis when challenged with free fatty acid and lipopolysaccha-rides, reminiscent of non-alcoholic fatty liver disease (NAFLD) stages. Treatment of MCLS cultures with potential anti-NAFLD drugs such as Elafibranor, Lanifibranor, Pioglitazone and Obeticholic acid shows that all can inhibit steatosis, but only Elafibranor and especially Lanifibranor inhibit fibrosis. Therefore, primary mouse MCLS cultures can be used to model acute and chronic liver disease and are suitable for the assessment of anti-NAFLD drugs.

Automated summary

Chronic liver disease can lead to liver fibrosis and cirrhosis. However, reliable in vitro models for studying liver disease and evaluating drugs are lacking. In this study, functional multicellular liver spheroid (MCLS) cultures were established using different types of liver cells. The MCLS cultures exhibited key characteristics of liver tissue and responded to chronic exposure to different substances. Furthermore, the MCLS cultures were used to evaluate potential anti-NAFLD drugs. The results showed that certain drugs could effectively inhibit steatosis and fibrosis. In conclusion, primary mouse MCLS cultures can be used as a model for acute and chronic liver disease, as well as for drug assessment.