Global transcriptional analysis of human functional hepatocytes Long-term maintained by small-molecule cocktails by RNA-seq

Functional maintenance of terminally differentiated cells outside the in vivo microenvironment has proved challenging. Current strategies that manipulate cell-cell or cell-matrix connections are difficult to constitute complex regulatory networks for cell function maintenance. Small molecules are easily combined for flexible spatiotemporal modulations, theoretically favorable for synergetic regulation of cell-innate signaling pathways to maintain cell function in vitro. Here, we developed small-molecule cocktails enabling robust maintenance of primary human hepatocytes (PHHs) longer than four weeks, with gene expression profiles, resembling those of freshly isolated PHHs; and prolong-cultured PHHs, for the first time, could maintain drug-metabolizing activities of enzymes accounting for over 80% of drug-oxidation and support hepatitis B virus infection in vitro for over one month. Importantly, this cocktail also promotes functional maturation of human pluripotent stem cell-derived hepatocytes. Our study demonstrates that this chemical approach effectively maintains terminally differentiated hepatocytes in vitro, which could be extended to various cell types.