Development of a primary hepatocyte-based preclinical spheroid model system for the analysis of Phase I and Phase II enzyme activity in vitro

Hepatic spheroids formed by the spontaneous self-aggregation of hepatocytes maintain better liver function than two-dimensional (2D) monolayer cultures. Although structural polarity and intercellular adhesion can be maintained in hepatic spheroids, cell-extracellular matrix (ECM) interactions cannot be reproduced in spheroid-based models. In this study, we leveraged customizable ECM component and fabricated hepatic by injecting a cell suspension supplemented with primary mouse hepatocytes and Matrigel into a high-density methylcellulose (MC) solution. Amongst other findings, gene expression analysis of hepatic spheroid aggregates containing Matrigel revealed elevated levels of drug-metabolizing enzymes (Phase I and II) and transporters (compared to spheroids without Matrigel). Furthermore, the induction of drug metabolism enzymes and activity in hepatic spheroids containing Matrigel also increased compared to those without Matrigel. Lastly, we demonstrated that hepatic spheroids containing Matrigel can maintain liver function by increasing HNF4α through activation of Hippo signaling. Overall, our results indicate that high-performance hepatic spheroids containing Matrigel can maintain liver function through cell-cell and cell-ECM interactions and can be used to increase the efficiency of drug screening.