An Efficient Nonviral Method to Generate Integration-Free Human-Induced Pluripotent Stem Cells from Cord Blood and Peripheral Blood Cells
An Efficient Nonviral Method to Generate Integration-Free Human-Induced Pluripotent Stem Cells from Cord Blood and Peripheral Blood Cells
复制标题
DOI：
10.1002/stem.1293
复制
发表时间：
2013-03-01
期刊：
影响因子：
5.2
通讯作者：
Yamanaka, Shinya
中科院分区：
文献类型：
--
作者：
Okita, Keisuke;Yamakawa, Tatsuya;Yamanaka, Shinya
诱导多能干细胞(IPSCs)的产生为使用患者特有的体细胞提供了机会，而体细胞是疾病建模和药物发现的宝贵来源。为了促进涉及这些细胞的研究，重要的是从更容易接触到的、侵入性更小的组织中制造出IPSCs，比如血液。我们最近报道了使用编码Oct3/4、SOX2、KLF4、L-MYC、LIN28和TP53的shRNA的质粒组合，从成人成纤维细胞高效地产生人IPSCs。我们在此报告了一种改进的方案，能够使用这些质粒载体从CD34(+)脐带血细胞和从健康捐赠者的外周血中高效地诱导IPSC。原混合质粒组能诱导出IPSCs，但效率较低。载体外体扩增的关键因子EBNA1的加入大大提高了IPSC的诱导效率，尤其是在从α-βT细胞进行诱导的情况下。这一改进使7名年龄从20多岁到60多岁的健康献血者建立了血液来源的IPSCs。这种诱导方法将有助于患者特异性无整合IPSCs的派生，也适用于未来临床级IPSCs的产生。干细胞2013；31：458-466
The generation of induced pluripotent stem cells (iPSCs) provides the opportunity to use patient-specific somatic cells, which are a valuable source for disease modeling and drug discovery. To promote research involving these cells, it is important to make iPSCs from easily accessible and less invasive tissues, like blood. We have recently reported the efficient generation of human iPSCs from adult fibroblasts using a combination of plasmids encoding OCT3/4, SOX2, KLF4, L-MYC, LIN28, and shRNA for TP53. We herein report a modified protocol enabling efficient iPSC induction from CD34(+) cord blood cells and from peripheral blood isolated from healthy donors using these plasmid vectors. The original plasmid mixture could induce iPSCs; however, the efficiency was low. The addition of EBNA1, an essential factor for episomal amplification of the vectors, by an extra plasmid greatly increased the efficiency of iPSC induction, especially when the induction was performed from alpha beta T cells. This improvement enabled the establishment of blood-derived iPSCs from seven healthy donors ranging in age from their 20s to their 60s. This induction method will be useful for the derivation of patient-specific integration-free iPSCs and would also be applicable to the generation of clinical-grade iPSCs in the future. STEM CELLS 2013;31:458-466