(278c) Recapitulating Liver Fibrosis in Vitro By Recreating the Fibrotic Milieu

Liver damage as a consequence of chronic stress can lead to fibrosis causing restructuring of the liver microenvironment (LME). The increase in the liver stiffness is a strong indication of the progression of liver diseases. However, the effect of increased stiffness on different liver cells has not been established. Clinical and animal studies have contributed significantly to our understanding of liver fibrosis biology but there exists a need for in vitro models to investigate the essential biochemical mechanistic pathways that associated with liver fibrosis microenvironment. Such models need to incorporate the dynamic changes in LME including the change in liver stiffness. In our study, we investigated the effect of substrate stiffness on the functions of cultured primary hepatocytes and liver sinusoidal endothelial cells (LSECs) utilizing a polydimethyl siloxane (PDMS) based substrate with tunable stiffness. We employed a soft substrate to represent the healthy liver tissue stiffness and a stiff substrate to represent diseased liver tissue and compared the cellular properties with the cells cultured on tissue culture polystyrene surfaces (TCPS). We observed that hepatocytes cultured on the soft substrate displayed a consistently more differentiated phenotype for a prolonged duration as compared to the hepatocytes that were cultured on the stiff substrate and TCPS. We further demonstrated that hepatocyte cultured on soft substrate showed better maintenance of hepatocyte-specific functions of urea and albumin synthesis. Similarly, cytochrome P450 (CYP) activity on soft substrate displayed a multi-fold increase in CYP activity, as compared to TCPS. In LSECs, we observed a stiffness mediated dedifferentiation in the cells, validated by defenestration of cells and overexpression of vascular endothelial cell markers. Together, these results indicate that the substrate stiffness plays a significant role in modulating hepatic cell behavior. Establishing mechanosensitivity of hepatic cells in liver fibrosis could result in the development of anti-fibrosis therapeutic approaches that target the mechanotransduction pathways of the cells to achieve functional restoration.