Living liver biopsies: 3D organoid cell culture models of liver disease for clinical medicine

Collins S1,2,3, Bryant K1,2,3 and Shackel N1,2,3

  1. Ingham Institute for Applied Medical Research.
  2. UNSW Sydney.
  3. South Western Sydney Local Health District.

Liver disease and cancer represent a significant disease burden worldwide. In vitro liver models have gained significant insights into the basic science of liver disease. A 3D organoid cell culture is an in vitro model defined as a collection of cells with several cell types, that develop from stem cells or organ progenitors which self-organise through cell sorting and spatially restricted lineage commitment, similar to organogenesis in vivo. This self-renewing primary cell culture has tissue-specific architecture, intercellular heterogeneity, mechanical and biochemical cues, cell-cell signalling and can be passaged into large data sets or frozen down for long term storage. We hypothesise that liver disease including hepatocellular carcinoma can be modelled ex vivo with 3D liver organoid cell cultures. The aim of this study is to establish and characterise mouse liver organoids for future use as a model for investigating liver injury. Mouse LGR5+ stem cells were isolated from dissociated liver tissue using fluorescence-activated cell sorting and viability assessed. LGR5+ cells were cultured in vitro under growth factor-defined conditions that included noggin and R-spondin-1 and were found to expand and self-organised into 3D structures which had the ability to differentiate into functionally mature hepatocytes. Mouse liver organoids were characterised for mature hepatocyte function using phase-contrast microscopy, hematoxylin and eosin staining, and immunofluorescence analysis. This research will establish methodology for culturing liver organoids, which can be used as an ex vivo assay of liver injury increase our understanding of the relationship between liver injury and the progression of fibrosis, cirrhosis and liver cancer.