Generation of Induced Pluripotent Stem Cells from Human Peripheral T Cells Using Sendai Virus in Feeder-free Conditions.
Generation of Induced Pluripotent Stem Cells from Human Peripheral T Cells Using Sendai Virus in Feeder-free Conditions.
复制标题
DOI：
10.3791/53225
复制
发表时间：
2015-11-11
期刊：
影响因子：
--
通讯作者：
Fukuda K
中科院分区：
文献类型：
--
作者：
Kishino Y;Seki T;Yuasa S;Fujita J;Fukuda K
最近，iPSC作为再生疗法的新细胞来源引起了人们的关注。虽然产生iPSC的最初方法依赖于通过侵入性活检和转基因的逆转录病毒基因组插入获得的真皮成纤维细胞，但已经做出了许多努力来避免这些缺点。人外周T细胞是产生iPSC的独特细胞来源。来源于T细胞的iPSC含有T细胞受体（TCR）基因的重排，并且是抗原特异性T细胞的来源。此外，基因组中的T细胞受体重排具有标记单个细胞系并区分移植细胞和供体细胞的潜力。为了iPSC的安全临床应用，重要的是将新产生的iPSC暴露于有害试剂的风险最小化。尽管胎牛血清和饲养细胞对于多能干细胞培养是必不可少的，但优选将它们从培养系统中去除以降低不可预测的致病性的风险。为了解决这个问题，我们已经建立了一种使用仙台病毒从人外周T细胞产生iPSC的方案，以降低将iPSC暴露于未定义病原体的风险。虽然处理仙台病毒需要具有适当生物安全水平的设备，但仙台病毒在没有基因组插入的情况下感染活化的T细胞，但效率很高。在该方案中，我们证明了在无饲养条件下使用活化T细胞培养物和仙台病毒的组合从人外周T细胞产生iPSC。
Recently, iPSCs have attracted attention as a new source of cells for regenerative therapies. Although the initial method for generating iPSCs relied on dermal fibroblasts obtained by invasive biopsy and retroviral genomic insertion of transgenes, there have been many efforts to avoid these disadvantages. Human peripheral T cells are a unique cell source for generating iPSCs. iPSCs derived from T cells contain rearrangements of the T cell receptor (TCR) genes and are a source of antigen-specific T cells. Additionally, T cell receptor rearrangement in the genome has the potential to label individual cell lines and distinguish between transplanted and donor cells. For safe clinical application of iPSCs, it is important to minimize the risk of exposing newly generated iPSCs to harmful agents. Although fetal bovine serum and feeder cells have been essential for pluripotent stem cell culture, it is preferable to remove them from the culture system to reduce the risk of unpredictable pathogenicity. To address this, we have established a protocol for generating iPSCs from human peripheral T cells using Sendai virus to reduce the risk of exposing iPSCs to undefined pathogens. Although handling Sendai virus requires equipment with the appropriate biosafety level, Sendai virus infects activated T cells without genome insertion, yet with high efficiency. In this protocol, we demonstrate the generation of iPSCs from human peripheral T cells in feeder-free conditions using a combination of activated T cell culture and Sendai virus.