Regulation of endodermal differentiation of human embryonic stem cells through integrin-ECM interactions.
Regulation of endodermal differentiation of human embryonic stem cells through integrin-ECM interactions.
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DOI：
10.1038/cdd.2012.138
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发表时间：
2013-03
影响因子：
12.4
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中科院分区：
文献类型：
--
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在发育过程中，许多细胞反应受整合素受体和细胞外基质蛋白（ECMPs）之间的相互作用调节。尽管最近大多数关于人胚胎干细胞（hESC）分化的研究都集中在生长因子（如FGF、TGFβ和WNT）的作用上，但对ECM P-整合素信号传导在该过程中的作用知之甚少。此外，目前指导hESC分化为各种谱系的策略是低效的，并且尚未在体外产生功能成熟的细胞。这表明，额外的因素，如ECMPs，需要的hESC的有效分化。使用高通量多因子细胞阵列技术，我们研究了数百种ECMP组合和浓度对几种hPSC系分化为定形内胚层（DE）的影响，定形内胚层是一种早期胚胎细胞群，注定会产生内脏器官，如肺、肝、胰腺、胃和肠。从这个屏幕上，我们确定纤连蛋白（FN）和玻连蛋白（VTN）的ECMP组件，促进DE分化。整合素表达分析显示，向DE分化导致FN结合整合素α5（ITGA 5）和VN结合整合素αV（ITGAV）增加。条件性短发夹RNA介导的ITGA 5和ITGAV的敲低破坏了hESC向DE的分化。最后，ITGA 5和ITGAV的基于荧光的细胞分选显著富集了具有与DE相关的基因表达特征的细胞，表明这些细胞表面蛋白允许从hESC分离和富集DE。这些数据提供的证据表明，FN和VTN通过与ITGA 5和ITGAV的相互作用促进hESC的内胚层分化，并且ECM-整联蛋白相互作用是hESC分化为功能成熟细胞所必需的。
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.
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