Laboratory of Genetics, National Institute on Aging, National Institutes of Health, Baltimore, Maryland, USA;
Laboratory of Genetics, National Institute on Aging, National Institutes of Health, Baltimore, Maryland, USA; Centre for Eye Research Australia, Department of Ophthalmology, University of Melbourne, Australia;
Stem Cells Transl Med. 2014 Jul;3(7):787-91. doi: 10.5966/sctm.2013-0036. Epub 2014 Jun 5.
Keratinocytes represent an easily accessible cell source for derivation of human induced pluripotent stem (hiPS) cells, reportedly achieving higher reprogramming efficiency than fibroblasts. However, most studies utilized a retroviral or lentiviral method for reprogramming of keratinocytes, which introduces undesirable transgene integrations into the host genome. Moreover, current protocols of generating integration-free hiPS cells from keratinocytes are mostly inefficient. In this paper, we describe a more efficient, simple-to-use, and cost-effective method for generating integration-free hiPS cells from keratinocytes. Our improved method using lipid-mediated transfection achieved a reprogramming efficiency of ∼0.14% on average. Keratinocyte-derived hiPS cells showed no integration of episomal vectors, expressed stem cell-specific markers and possessed potentials to differentiate into all three germ layers by in vitro embryoid body formation as well as in vivo teratoma formation. To our knowledge, this represents the most efficient method to generate integration-free hiPS cells from keratinocytes.
角质形成细胞是一种易于获得的细胞来源，可用于衍生人类诱导多能干细胞（hiPS 细胞），据报道其重编程效率高于成纤维细胞。然而，大多数研究利用逆转录病毒或慢病毒方法对角质形成细胞进行重编程，这会将不期望的转基因整合到宿主基因组中。此外，目前从角质形成细胞生成无整合 hiPS 细胞的方案大多效率不高。在本文中，我们描述了一种更有效、使用更简便且更经济高效的方法，用于从角质形成细胞生成无整合 hiPS 细胞。我们使用脂质转染的改进方法，平均重编程效率约为 0.14%。角质形成细胞衍生的 hiPS 细胞没有整合附加型载体，表达干细胞特异性标志物，并具有通过体外胚胎体形成以及体内畸胎瘤形成分化为三个胚层的潜能。据我们所知，这是从角质形成细胞生成无整合 hiPS 细胞的最有效方法。
