Development of an in vitro model of healthy and diseased hepatic tissue using 3D matrices

In vitro liver models are an important tool in hepatic tissue engineering as well as in drug and chemical toxicity testing. Recent studies have shown the importance of the several factors to preserve a differentiated hepatotypic phenotype, such as extracellular matrix (ECM) and its adhesive factors, the three dimensional architecture, the enrichment of the cell culture medium (e.g. growth factors, hormones, vitamins), the high cell density and the presence of other hepatic cell types. The aim of this thesis is to develop new in vitro models of healthy and fibrotic human hepatic tissue for drug toxicity study. In particular the attention is focused on studying the influence of the hepatic fibrosis in the modulation of drug toxicity. The research program is divided in two main sections: the development of protocols for the decellularization of pig liver in order to obtain matrices to be used as substrates for cell culture (activity performed during the first year) and the development of hepatic in vitro models (second and third year). The first part of this project was also useful in determining the main properties of the hepatic extracellular matrix, which were then used for the selection of the scaffolding materials used in further in vitro models. In this latter section, the study was focused on the selection of ‘an optimal’ hepatocyte cellular model and scaffolding material(s): different hepatocyte sources were grown on several engineered constructs evaluating differences in cell function and morphology. The results obtained allowed to select a ‘healthy’ and ‘diseased’ hepatic in vitro models, able to preserve the hepatic phenotype of cultured cells in a 3D environment. These in vitro models can be further used for drug testing.