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

Abstract

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.

Advances in high throughput cell culture technologies for therapeutic screening and biological discovery applications

“…Recently, we demonstrated the implementation of such a strategy towards the creation and analysis of 92 heterogeneous microenvironments for the culture of hepatic stellate cells in the context of nonalcoholic steatohepatitis, a condition that is characterized by heterogeneity of cellular microenvironments. 220 …”