Characterization of mesendoderm: a diverging point of the definitive endoderm and mesoderm in embryonic stem cell differentiation culture

Abstract

Bipotent mesendoderm that can give rise to both endoderm and mesoderm is an established entity from C. elegans to zebrafish. Although previous studies in mouse embryo indicated the presence of bi-potent mesendoderm cells in the organizer region, characterization of mesendoderm and its differentiation processes are still unclear. As bi-potent mesendoderm is implicated as the major precursor of definitive endoderm, its identification is also essential for exploring the differentiation of definitive endoderm. In this study, we have established embryonic stem (ES) cell lines that carry GFP gene in the goosecoid (Gsc) gene locus and have investigated the differentiation course of mesendodermal cells using Gsc expression as a marker. Our results show that mesendoderm is represented as a Gsc-GFP+E-cadherin(ECD)+PDGFRα(αR)+population and is selectively induced from ES cells under defined conditions containing either activin or nodal. Subsequently, it diverges to Gsc+ECD+αR- and Gsc+ECD-αR+ intermediates that eventually differentiate into definitive endoderm and mesodermal lineages,respectively. The presence of mesendodermal cells in nascent Gsc+ECD+αR+ population was also confirmed by single cell analysis. Finally, we show that the defined culture condition and surface markers developed in this study are applicable for obtaining pure mesendodermal cells and their immediate progenies from genetically unmanipulated ES cells.

In vitro derivation of functional insulin-producing cells from human embryonic stem cells

“…The individual effect of activin A and RA on endoderm and pancreas development has also been demonstrated by several other groups [18][19][20][21][22][23][24]. It has been shown that mouse and human ES cells can be efficiently induced into Sox17-positive definitive endoderm by activin A treatment [18][19][20]. RA has been reported to be critical for early pancreas formation during zebrafish and mouse embryonic development [21][22][23][24].…”

“…Results from our previous study [11], as well as the work of others [18][19][20], showed that high concentrations of activin A can lead to efficient endoderm differentiation. We tested with various concentrations of activin A, and found that activin A at a concentration as RT-PCR analyses revealed that activin A treatment at 50 ng/ml resulted in an increased expression of endoderm genes such as sox17 and hnf4α ( Figure 2B).…”

Roles of TGF-β family signaling in stem cell renewal and differentiation

“…Recently, reporter mouse ES cell lines in which reporter cDNAs (e.g. GFP) are targeted to specific marker genes have been engineered to model the regional specification of primitive streak in vitro [30].…”

“…Activation of Nodal pathways by addition of activin induced a primitive streak population that expresses Foxa2 and Goosecoid, and subsequent formation of definitive endoderm or mesoderm depending on the concentrations of activin. Clonal analysis revealed that individual Goosecoid-positive cells have the potential to develop into both mesoderm and endoderm, suggesting that they may represent mesendoderm progenitors [30]. In contrast to Nodal/activin signals, BMP signals alone induce the formation of posterior primitive streak population.…”

Developmental potential of Gcn5−/− embryonic stem cells in vivo and in vitro

“…Expression of a third mesodermal marker, Goosecoid (Gsc; Fig. 6C; Tada et al, 2005;Naito et al, 2006) was unchanged in Gcn5 Ϫ/Ϫ EBs. Together these results indicate that some aspects of mesoderm development are altered in the absence of Gcn5.…”