The Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai 200241, China.
Biomaterials. 2012 Jul;33(20):5047-55. doi: 10.1016/j.biomaterials.2012.03.061. Epub 2012 Apr 9.
Generation of induced pluripotent stem cells (iPSCs) holds great promise to regenerative medicine. However, before this technology can be applied for clinical purpose, the issues of iPSC efficiency and safety need to be addressed. In this study, we have compared a simple TAT- and 11 arginine (R)-protein transduction domain (PTD) for somatic cell reprogramming and explored the optimal conditions for the PTD to transduce reprogramming factors (RFs). We show that all recombinant TAT- and 11R-fused RFs are transcriptionally active as they activate their corresponding reporter genes in reporter assays. The TAT-RFs are in general transcriptionally more active than the corresponding 11R-RFs, but less active than the corresponding retroviral transduced RFs. Furthermore, each of TAT-RFs can substitute for their corresponding retrovirus in reprogramming. Finally, using five TAT-RFs together with an HDAC inhibitor, we can generate iPSC-like colonies from human fibroblast cells with high efficiency approximately 2 weeks after the first protein transduction. These colonies exhibit unique features of pluripotent stem cells including the morphology and the expression of pluripotency-associated markers. This characterization of recombinant RFs in reprogramming should facilitate the generation of clinically useful and genetic material-free human iPSCs.
诱导多能干细胞（iPSCs）的产生为再生医学带来了巨大的希望。然而，在这项技术能够应用于临床之前，需要解决 iPSC 的效率和安全性问题。在这项研究中，我们比较了一种简单的 TAT-和 11 个精氨酸（R）-蛋白转导结构域（PTD）用于体细胞重编程，并探索了 PTD 转导重编程因子（RFs）的最佳条件。我们表明，所有重组的 TAT-和 11R-融合的 RFs 在报告基因检测中均具有转录活性，因为它们可以激活相应的报告基因。TAT-RFs 的转录活性通常高于相应的 11R-RFs，但低于相应的逆转录病毒转导的 RFs。此外，TAT-RFs 中的每一个都可以替代其相应的逆转录病毒在重编程中使用。最后，我们使用五种 TAT-RFs 与组蛋白去乙酰化酶抑制剂一起，能够从人成纤维细胞中高效地产生 iPSC 样集落，大约在第一次蛋白转导后 2 周。这些集落表现出多能干细胞的独特特征，包括形态和多能性相关标志物的表达。这种对重编程中重组 RFs 的特性描述应该有助于产生临床上有用且不含遗传物质的人 iPSCs。
