Molecular mechanisms regulating the establishment of hepatocyte polarity during human hepatic progenitor cell differentiation into a functional hepatocyte-like phenotype

Abstract

Summary The correct functioning of hepatocytes requires the establishment and maintenance of hepatocyte polarity. However, the mechanisms regulating the generation of hepatocyte polarity are not completely understood. The differentiation of human fetal hepatic progenitor cells (hFHPCs) into functional hepatocytes provides a powerful in vitro model system for studying the molecular mechanisms governing hepatocyte development. In this study, we used a two-stage differentiation protocol to generate functional polarized hepatocyte-like cells (HLCs) from hFHPCs. Global gene expression profiling was performed on triplicate samples of hFHPCs, immature-HLCs and mature-HLCs. When the differential gene expression was compared based on the differentiation stage, a number of genes were identified that might be essential for establishing and maintaining hepatocyte polarity. These genes include those that encode actin filament-binding protein, protein tyrosine kinase activity molecules, and components of signaling pathways, such as PTK7, PARD3, PRKCI and CDC42. Based on known and predicted protein-protein interactions, the candidate genes were assigned to networks and clustered into functional categories. The expression of several of these genes was confirmed using real-time RT-PCR. By inactivating genes using small interfering RNA, we demonstrated that PTK7 and PARD3 promote hepatic polarity formation and affect F-actin organization. These results provide unique insight into the complex process of polarization during hepatocyte differentiation, indicating key genes and signaling molecules governing hepatocyte differentiation.