Suppression of NANOG Induces Efficient Differentiation of Human Embryonic Stem Cells to Pancreatic Endoderm
Suppression of NANOG Induces Efficient Differentiation of Human Embryonic Stem Cells to Pancreatic Endoderm
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DOI：
10.1097/mpa.0b013e31822362e4
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发表时间：
2012-01-01
期刊：
影响因子：
2.9
通讯作者：
Sidhu, Kuldip S.
中科院分区：
文献类型：
--
作者：
Lie, Khun-Hong;Tuch, Bernard E.;Sidhu, Kuldip S.
目的：利用人胚胎干细胞(HESCs)作为替代β细胞来源的一个挑战是建立有效地引导其分化为有功能的β细胞的方法。本研究的目的是评估抑制NANOG基因在hESCs分化中的作用，以提高内胚层来源的胰腺细胞的生成效率。方法：利用基于质粒的siRNA方法在hESC Emvy细胞系中产生稳定抑制NANOG的同源细胞群。根据基于本体的体外选择方案进行胰腺分化，然后移植到免疫缺陷小鼠体内成熟。结果：我们观察到决定性内胚层基因的上调，扩大了NANOG对决定性内胚层分化的阻断作用。基于本体的分化方案导致胰腺上皮发育所必需的标志物表达增加。这些细胞移植后可见均一的胰腺外分泌样形态，淀粉酶染色呈阳性。结论：抑制NANOG的作用表现出向内胚层和胰腺前体细胞的有效分化。研究内分泌形成所需的因素，结合体内长期培养，可以进一步提高内分泌-外分泌细胞命运的比例。
Objective: A challenge in using human embryonic stem cells (hESCs) as the source of surrogate beta cells is the establishment of methods that could effectively direct their differentiation into functional beta cells. The aim of this study was to assess the effect of NANOG gene suppression in differentiating hESCs as a mean of increasing the efficiency with which endoderm-derived pancreatic cells could be generated.Methods: A homogenous cell population with stable suppression of NANOG was generated in hESC ENVY line using plasmid-based siRNA approach. Pancreatic differentiation was undertaken according to the ontology-based in vitro selection protocol and followed by transplantation into immunodeficiency mice to mature in vivo.Results: We observed up-regulation of definitive endoderm genes, which expand the role of NANOG in blocking definitive endoderm differentiation. The ontology-based differentiation protocol resulted in increased expression of markers essential for pancreatic epithelium development. Transplantation of these cells further revealed a homogenous pancreatic exocrine-like morphology that stained positively for amylase.Conclusions: The suppression of NANOG displayed an effective differentiation toward endoderm and pancreatic progenitors. Investigation of the factors required for endocrine formation combined with a prolonged in vivo culturing could be further used to increase the ratio of endocrine-exocrine cells fate.