Phenotype reversion in fetal human liver epithelial cells identifies the role of an intermediate meso-endodermal stage before hepatic maturation

Abstract

Understanding the biological potential of fetal stem/progenitor cells will help define mechanisms in liver development and homeostasis. We isolated epithelial fetal human liver cells and established phenotype-specific changes in gene expression during continuous culture conditions. Fetal human liver epithelial cells displayed stem cell properties with multilineage gene expression, extensive proliferation and generation of mesenchymal lineage cells, although the initial epithelial phenotype was rapidly supplanted by meso-endodermal phenotype in culture. This meso-endodermal phenotype was genetically regulated through cytokine signaling, including transforming growth factor β, bone morphogenetic protein, fibroblast growth factor and other signaling pathways. Reactivation of HNF3α (FOXA1) transcription factor, a driver of hepatic specification in the primitive endoderm, indicated that the meso-endodermal phenotype represented an earlier developmental stage of cells. We found that fetal liver epithelial cells formed mature hepatocytes in vivo, including after genetic manipulation using lentiviral vectors, offering convenient assays for analysis of further cell differentiation and fate. Taken together, these studies demonstrate plasticity in fetal liver epithelial stem cells, offer paradigms for defining mechanisms regulating lineage switching in stem cells, and provide potential avenues for regulating cell phenotypes for applications of stem cells, such as for cell therapy.

Evidence for a common progenitor of epithelial and mesenchymal components of the liver

“…25 Moreover, we believe that our results evoke those previously described in three elegant reports, [26][27][28] in which liver progenitor cells bearing an epithelial-mesenchymal habit were showed. However, in the characterization of all of those cells the evidence of a mesenchymal differentiation in vivo is lacking.…”

Spontaneous origin from human embryonic stem cells of liver cells displaying conjoint meso-endodermal phenotype with hepatic functions

“…2D). This was similar to the conjoint meso-endodermal properties in FH-Ep-P3 cells, as previously described (Inada et al, 2008b).…”

“…In studies of fetal liver development and of isolated fetal liver stem or progenitor cells, significant differences were noted in these cells compared with mature hepatocytes, including multilineage gene expression patterns in fetal cells (Inada et al, 2008a;Inada et al, 2008b). The discovery of an unexpected conjoint meso-endodermal phenotype in fetal liver stem cells, which was characterized by simultaneous expression of epithelial and mesenchymal properties, was in agreement with their ability to generate endodermal and mesodermal lineages, such as adipocytes, osteocytes or endothelial cells, as well as hepatocytes or biliary cells (Inada et al, 2008b).…”

“…The discovery of an unexpected conjoint meso-endodermal phenotype in fetal liver stem cells, which was characterized by simultaneous expression of epithelial and mesenchymal properties, was in agreement with their ability to generate endodermal and mesodermal lineages, such as adipocytes, osteocytes or endothelial cells, as well as hepatocytes or biliary cells (Inada et al, 2008b). During expansion under cell culture conditions, fetal liver stem cells were found to express mesenchymal genes such as vimentin or a-smooth muscle actin (SMA) better, without loss of expression of the epithelial genes E-cadherin, albumin (Alb), glucose-6-phosphatase (G-6-P), glycogen, cytokeratin (CK)-19, c-glutamyltranspeptidase (GGT), dipeptidyl peptidase IV (DPPIV), etc.…”

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

“…hFHPCs express the hepatic stem cells/progenitor cells marker AFP, ALB, CK19, epithelial cell adhesion molecule (EpCAM) (Schmelzer et al, 2006;Inada et al, 2008), Delta-like protein (Dlk) (Yanai et al, 2010), and sal-like protein 4 (Sall4) (Oikawa et al, 2009) (supplementary material Fig. S1A).…”