UC San Diego

Engineered liver tissue has the potential to support patients with hepatic dysfunction as well as provide an in vitro model of the liver for pharmaceutical drug development. Such applications mandate the use of primary hepatocytes- the parenchymal cell of the liver; however, hepatocytes rapidly lose viability and phenotypic functions in vitro. Co-cultivation of hepatocytes with nonparenchymal cells (NPC) has been reported to prevent this deterioration. Several aspects of this so-called 'co- culture effect' remain incompletely understood: the molecular mechanisms by which NPC stabilize liver-specific functions, dynamics of interaction (i.e. whether continuous NPC support is required to maintain hepatic functions), and whether this model system can provide a useful tool for drug development. In this dissertation, gene expression profiling, electroactive surface chemistry, and microfabrication tools are used to answer these questions. First, a functional genomic approach utilizing gene expression profiling was developed to identify and validate molecular mediators that modulate liver-specific functions in co-cultures. Second, the role of one candidate, T-cadherin, was characterized in detail. T- cadherin was found to upregulate hepatocyte functions in vitro upon presentation in both cellular (CHO cells) and acellular (immobilized protein) contexts. Third, micropatterned electroactive substrates were used to selectively release fibroblasts at various time points from co-culture to probe the dynamics of cell-cell interaction. Results indicated that continuous fibroblast stimulation was required to maintain hepatic functions. Finally, soft lithography tools were utilized to develop miniaturized, multi-well co-culture models of the human and rat liver that were stable for several weeks. Utility of microscale tissues for evaluating species-specific drug metabolism, drug-drug interactions, and susceptibility to a panel of hepatotoxins was demonstrated. These studies will have an impact on developing functional models of liver tissue for use in fundamental hepatology, cell-based therapies for liver disease and pharmaceutical drug development. Furthermore, the methods developed here can be generalized to other tissues where cell-cell interactions modulate cellular fates