Engineering Culture Platforms to Study Human Liver Cancer and Fibrosis

The liver executes a plethora of functions such as drug metabolism, plasma protein secretion, and urea synthesis. Additionally, drug toxicity to the liver is a leading cause of drug attrition and liver cancer is the third leading cause of cancer-related deaths worldwide. Given differences between animals and humans in liver pathways, models of the human liver are essential for mechanistic studies and for drug screening. Unfortunately, human liver cells, both primary and cancer-derived, display low liver cell functions in conventional 2D monolayers. The micropatterned coculture (MPCC) platform, in which hepatocytes are organized onto collagen domains of empirically optimized dimensions and subsequently cocultured with fibroblasts, has been shown to induce high hepatocyte functions for several weeks. However, the current configuration of MPCCs lacks a key cell type of the liver, cholangiocytes (biliary epithelial cells), and it is not clear if MPCCs can be used for the development of a liver cancer screening system. This thesis aims to address these abovementioned limitations. Results obtained demonstrate that several liver cancer cell lines cultured in the MPCC platform maintain cell proliferation, higher liver cell functions, and can be effectively used to determine cytotoxicity of cancer drugs and detect cancer related gene expression. Furthermore, results show that while cholangiocytes cannot support primary human hepatocytes to the same extent as the fibroblasts, creation of a three-cell type model (hepatocytes, cholangiocytes, and fibroblasts) with higher ratio of fibroblasts and lower ratio of cholangiocytes allows high levels of functions in hepatocytes and retention of cholangiocyte morphology. Interestingly, three-cell type model also showed significantly inhibition of hepatic function and displayed increased microvascular steatosis which may suggest that the three-cell type model could be developed as a liver disease (e.g. fibrosis) model. In conclusion, the model systems developed are potentially useful for fundamental research in liver cancer and hepatocyte-cholangiocyte interactions, and for screening the cancer related pathways and efficacy and/or toxicity of drugs. In the future, the model systems can be integrated to study the effects of cholangiocytes in liver diseases or on liver cancer cells and to build models of both hepatocellular carcinoma and cholangiocarcinoma (liver cancer subtypes).