Liver damage as a consequence of injury or disease 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, there is a lack of understanding in the effect of increased stiffness on the liver cells in vivo. There is a critical need to engineer in vitro models that will mimic various stages of the diseased liver to serve as accurate models for studying disease mechanism of individual cell types. 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 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. Together, these results indicate that the substrate stiffness plays a significant role in modulating hepatocyte behavior. The intricate LME-hepatocytes signaling pathways through which healthy and diseased liver cells communicate within the LME will be further investigated in our future studies.