Division of Reproductive Endocrinology and Infertility, Department of Obstetrics and Gynecology, College of Medicine and the Hospital, National Taiwan University, Taipei, Taiwan.
Stem Cell Rev Rep. 2011 Sep;7(3):722-35. doi: 10.1007/s12015-011-9233-y.
The derivation of induced pluripotent stem cells (iPSCs) requires not only efficient reprogramming methods, but also reliable markers for identification and purification of iPSCs. Here, we demonstrate that surface markers, epithelial cells adhesion molecule (EpCAM) and epithelial cadherin (E-cadherin) can be used for efficient identification and/or isolation of reprogrammed mouse iPSCs. By viral transduction of Oct4, Sox2, Klf4 and n- or c-Myc into mouse embryonic fibroblasts, we observed that the conventional mouse embryonic stem cell (mESC) markers, alkaline phosphatase (AP) and stage-specific embryonic antigen 1 (SSEA1), were expressed in incompletely reprogrammed cells that did not express all the exogenous reprogramming factors or failed to acquire pluripotent status even though exogenous reprogramming factors were expressed. EpCAM and E-cadherin, however, remained inactivated in these cells. Expression of EpCAM and E-cadherin correlated with the activation of Nanog and endogenous Oct4, and was only seen in the successfully reprogrammed iPSCs. Furthermore, purification of EpCAM-expressing cells at late reprogramming stage by FACS enriched the Nanog-expressing cell population suggesting the feasibility of selecting successful reprogrammed mouse iPSCs by EpCAM expression. We have thus identified new surface markers that can efficiently identify successfully reprogrammed iPSCs and provide an effective means for iPSC isolation.
诱导多能干细胞（iPSCs）的衍生不仅需要高效的重编程方法，还需要可靠的标志物来鉴定和纯化 iPSCs。在这里，我们证明表面标志物上皮细胞黏附分子（EpCAM）和上皮钙黏蛋白（E-cadherin）可用于有效鉴定和/或分离重编程的小鼠 iPSCs。通过病毒转导将 Oct4、Sox2、Klf4 和 n- 或 c-Myc 导入小鼠胚胎成纤维细胞，我们观察到常规的小鼠胚胎干细胞（mESC）标志物碱性磷酸酶（AP）和阶段特异性胚胎抗原 1（SSEA1）在不完全重编程的细胞中表达，这些细胞不表达所有的外源性重编程因子，或者即使表达了外源性重编程因子，也未能获得多能状态。然而，EpCAM 和 E-cadherin 在这些细胞中仍然失活。EpCAM 和 E-cadherin 的表达与 Nanog 和内源性 Oct4 的激活相关，仅在成功重编程的 iPSCs 中可见。此外，通过 FACS 在晚期重编程阶段纯化表达 EpCAM 的细胞富集了表达 Nanog 的细胞群体，这表明通过 EpCAM 表达选择成功重编程的小鼠 iPSCs 是可行的。因此，我们鉴定了新的表面标志物，可有效鉴定成功重编程的 iPSCs，并为 iPSC 分离提供有效手段。
