Regulation of endodermal differentiation of human embryonic stem cells through integrin-ECM interactions

Department of Cellular and Molecular Medicine, Stem Cell Program, University of California, La Jolla, CA, USA.
Cell death and differentiation (Impact Factor: 8.18). 11/2012; 20(3). DOI: 10.1038/cdd.2012.138
Source: PubMed


Many cellular responses during development are regulated by interactions between integrin receptors and extracellular matrix proteins (ECMPs). Although the majority of recent studies in human embryonic stem cell (hESC) differentiation have focused on the role of growth factors, such as FGF, TGFβ, and WNT, relatively little is known about the role of ECMP-integrin signaling in this process. Moreover, current strategies to direct hESC differentiation into various lineages are inefficient and have yet to produce functionally mature cells in vitro. This suggests that additional factors, such as ECMPs, are required for the efficient differentiation of hESCs. Using a high-throughput multifactorial cellular array technology, we investigated the effect of hundreds of ECMP combinations and concentrations on differentiation of several hPSC lines to definitive endoderm (DE), an early embryonic cell population fated to give rise to internal organs such as the lung, liver, pancreas, stomach, and intestine. From this screen we identified fibronectin (FN) and vitronectin (VTN) as ECMP components that promoted DE differentiation. Analysis of integrin expression revealed that differentiation toward DE led to an increase in FN-binding integrin α5 (ITGA5) and VTN-binding integrin αV (ITGAV). Conditional short hairpin RNA-mediated knockdown of ITGA5 and ITGAV disrupted hESC differentiation toward DE. Finally, fluorescence-based cell sorting for ITGA5 and ITGAV significantly enriched cells with gene expression signatures associated with DE, demonstrating that these cell surface proteins permit isolation and enrichment of DE from hESCs. These data provide evidence that FN and VTN promote endoderm differentiation of hESCs through interaction with ITGA5 and ITGAV, and that ECMP-integrin interactions are required for hESC differentiation into functionally mature cells.Cell Death and Differentiation advance online publication, 16 November 2012; doi:10.1038/cdd.2012.138.

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    • "Induction of DE formation is the first and critical step of development in endodermal organs (Noguchi, 2009; Brafman et al., 2013; Hoveizi et al., 2014a). This germ layer gives rise to the intestinal epithelium and various other cell types, such as pancreas, hepatocytes, derived from the gut (D'Amour et al., 2005; Kim et al., 2010; Brafman et al., 2013; Jiang et al., 2013). The importance of the Activin A, Nodal, Wnt3a and TGFb pathway in DE induction has been demonstrated (Hosoya et al., 2012). "
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    • "Figure 2 Growth models of hPSCs under various growth conditions. (A) Growth models of hPSC colonies in hESC medium that contains growth factors FGF2 and TGFβ on Matrigel-based extracellular matrices with deduced anatomical structures and molecular complexes: Laminin (LM), fibronectin (FN), and vitronectin (VNT) mediate their effect via the α6β1, α5β1, and αVβ5 integrins respectively (Braam et al., 2008; Brafman et al., 2012). Different zones of out-growing hPSC colonies have differential epithelial-to-mesenchymal transition (EMT) states, in which the cells in the center of the colonies possess more epithelial characteristics and cells at the periphery develop mesenchymal-like phenotypes. "
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