Combinatorial Microgels for 3D ECM Screening and Heterogeneous Microenvironmental Culture of Primary Human Hepatic Stellate Cells

Non-alcoholic fatty liver disease affects 30% of the United States population and its progression can lead to non-alcoholic steatohepatitis (NASH), which can result in cirrhosis and hepatocellular carcinoma. NASH is characterized by a highly heterogeneous liver microenvironment created by the fibrotic activity of hepatic stellate cells (HSCs). While HSCs have been widely studied in 2D, further advancements in physiologically-relevant 3D culture platforms for the in vitro modeling of these heterogeneous environments are needed. In this study, we have demonstrated the use of stiffness-variable, ECM protein-conjugated polyethylene glycol microgels as 3D cell culture scaffolds to modulate HSC activation. We further employed these microgels as a high throughput ECM screening system to identify HSC matrix remodeling and metabolic activities in distinct heterogeneous microenvironmental conditions. In particular, 6 kPa fibronectin microgels were shown to significantly increase HSC matrix remodeling and metabolic activities in single or multiple component microenvironments. Overall, heterogeneous microenvironments consisting of multiple distinct ECM microgels promoted a decrease in HSC matrix remodeling and metabolic activities compared to homogeneous microenvironments. We envision this ECM screening platform being adapted to a broad number of cell types to aid the identification of ECM microenvironments that best recapitulate the desired phenotype, differentiation, or drug efficacy.