INDUCED PLURIPOTENT STEM CELL DIFFERENTIATION

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 目的：探讨人诱导多能干细胞的分化潜能 细胞 进展： 滋养外胚层谱系的形成代表了哺乳动物中最早的事件之一。 哺乳动物胚胎分化多能干细胞来源于人类和 动物胚胎在其形成细胞的能力方面表现出一系列分化潜能， 滋养细胞谱系自发或定向分化。这些 观察结果可能反映了导致这种情况的异质条件和事件 胚胎干细胞的永生化。诱导多能干 细胞（iPSC）提供了一个机会，以进一步阐明对 滋养层的形成。人iPSC系IMR 90 -1、IMR 90 - 2和IMR 90 - 4分别在小鼠和小鼠中表达。 将包皮-1、包皮-2和包皮-3置于体外培养条件下，显示出诱导 人胚胎干细胞滋养层分化：BMP-4和胚状体处理 形成，然后在二维或三维基质胶上贴壁培养 环境.滋养层细胞分化通过分泌人 绒毛膜促性腺激素（hCG），并通过组织学和免疫组织化学分析 从iPSC形成的胚状体。在所有的细胞系中，用BMP-4处理而不用BMP-4处理， 激活素、TGF-β、FGF 4或斑马鱼FGF 2导致了剂量相关的增加， 在处理的六天内hCG分泌到培养基中。同样地， 在Matrigel包被的二维表面上贴壁培养胚状体 支持在长达4周的培养期间hCG表达的一致起始。 有些出乎意料的是，iPSC衍生的胚状体不太一致地上调， 当置于三维基质胶外植体中时hCG的分泌。这是在 免疫组化法检测hCG分泌的困难， 这些三维胚状体培养物，虽然滋养层分化， 表面和生长细胞的细胞角蛋白染色表明。我们的结论是 iPSC对BMP-4处理具有与人胚胎来源 多能干细胞在滋养层分化方面，并且进一步 在三维细胞外基质环境中评估分化， needed. 出版物： 没有。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Objective: To define the differentiation potential of human induced pluripotent stem cells. PROGRESS: The formation of the trophectoderm lineage represents one of the earliest events in mammalian embryonic differentiation. Pluripotent stem cells derived from human and animal embryos exhibit a range of differentiation potential in their ability to form the trophoblast lineage upon spontaneous or directed differentiation. These observations may reflect the heterogeneous conditions and events that lead to the immortalization of embryonic cells to embryonic stem cells. Induced pluripotent stem cells (iPSC) provide an opportunity to shed further light on the requirements for trophoblast formation in human embryos. The human iPSC lines IMR90-1, -2 and Foreskin-1, -2 and -3 were subjected to in vitro culture conditions shown to induce trophoblast differentiation in human ESC: treatment with BMP-4, and embryoid body formation followed by adherent culture on 2-dimensional or 3-dimensional Matrigel environments. Trophoblast differentiation was monitored by secretion of human chorionic gonadotropin (hCG), and by histological and immunohistochemical analysis of embryoid bodies formed from iPSC. In all cell lines, treatment with BMP-4 but not activin, TGF-beta, FGF4 or zebrafish FGF2 resulted in a dose-related increase in the secretion of hCG into the culture medium within six days of treatment. Likewise, adherent culture of embryoid bodies on Matrigel-coated 2-dimensional surfaces supported consistent initiation of hCG expression during up to 4 weeks of culture. Somewhat unexpectedly, iPSC-derived embryoid bodies less consistently upregulated the secretion of hCG when placed into 3-dimensional Matrigel explants. This was in agreement with difficulty in detecting hCG secretion by immunohistochemistry in these 3-D embryoid body cultures, although trophoblast differentiation was suggested by cytokeratin staining of surface and outgrowth cells. We conclude that iPSC have similar responses to BMP-4 treatment as human embryo-derived pluripotent stem cells with regard to trophoblast differentiation, and that further evaluation of differentiation in 3-dimensional extracellular matrix environments is needed. PUBLICATIONS: None.