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

D. A. Brafman, C. Phung, N. Kumar, K. Willert

Research output: Contribution to journalArticlepeer-review

76 Scopus citations

Abstract

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 5 (ITGA5) and VTN-binding integrin V (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.

Original languageEnglish (US)
Pages (from-to)369-381
Number of pages13
JournalCell Death and Differentiation
Volume20
Issue number3
DOIs
StatePublished - 2013
Externally publishedYes

Keywords

  • arrayed cellular microenvironments
  • endoderm development
  • extracellular matrix proteins
  • human embryonic stem cells
  • integrin signaling

ASJC Scopus subject areas

  • Molecular Biology
  • Cell Biology

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