Spencer Helen, Keramari Maria, Ward Christopher M
Faculty of Medical and Human Sciences, Centre for Molecular Medicine, The University of Manchester, Manchester, UK.
Methods Mol Biol. 2011;690:81-94. doi: 10.1007/978-1-60761-962-8_5.
Embryonic stem cells (ESCs) are pluripotent cells derived from preimplantation embryos and can be maintained in an undifferentiated state over prolonged periods in vitro. In addition, ESCs can be induced to differentiate into cells representative of the three primary germ layers. As such, ESCs are a useful system for studying early developmental events in vitro and have the potential to provide a ubiquitous supply of somatic cells for use in regenerative medicine. However, significant differences in the expression pattern of various cell surface markers between murine and human ESCs, e.g. the SSEA series, necessitate the use of separate markers for determining the undifferentiated state of these cells. We have recently shown that an E- to N-cadherin switch occurs during spontaneous differentiation of both murine and human ESCs. Here we describe the use of E-cadherin and N-cadherin proteins and transcript expression for assessing the proportion of undifferentiated and spontaneously differentiated cells within ESC populations. In summary, loss of cell surface E-cadherin and/or gain of N-cadherin protein expression provides a useful nondestructive assay for the determination of the proportion of spontaneously differentiated cells within an ESC population. In addition, presence of N-cadherin transcripts in an ESC population is indicative of spontaneous differentiation of a proportion of the cells.
胚胎干细胞（ESCs）是源自植入前胚胎的多能细胞，能够在体外长期维持未分化状态。此外，胚胎干细胞可被诱导分化为代表三个主要胚层的细胞。因此，胚胎干细胞是体外研究早期发育事件的有用系统，并且有潜力为再生医学提供大量的体细胞。然而，小鼠和人类胚胎干细胞之间各种细胞表面标志物（如SSEA系列）的表达模式存在显著差异，因此需要使用不同的标志物来确定这些细胞的未分化状态。我们最近发现，在小鼠和人类胚胎干细胞的自发分化过程中会发生E-钙黏蛋白到N-钙黏蛋白的转换。在此，我们描述了使用E-钙黏蛋白和N-钙黏蛋白的蛋白质及转录本表达来评估胚胎干细胞群体中未分化细胞和自发分化细胞的比例。总之，细胞表面E-钙黏蛋白的缺失和/或N-钙黏蛋白蛋白表达的增加为测定胚胎干细胞群体中自发分化细胞的比例提供了一种有用的非破坏性检测方法。此外，胚胎干细胞群体中存在N-钙黏蛋白转录本表明一部分细胞发生了自发分化。
