关于非生殖性成年细胞能够被重新编程而成为多能细胞、从而能够分化成任何细胞类型的发现,让人们看到了一些激动人心的可能性。重新编程的细胞(被称为“诱导多能干细胞”,即iPS细胞)在再生医学中应当有巨大潜力,但目前用来生成它们的方法大多数都涉及到病毒基因的使用,这有可能使所诱导的细胞发生异常。
两个小组在本期Nature上报告了一项合作研究工作,这项工作成功地在没有使用病毒载体的情况下使人体细胞产生了多能性。研究人员利用来自病毒的2A肽序列生成一种结合重新编程因子的“多顺反子载体”(multicistronic vector),该载体被piggyBac转位子载体送入细胞中,从而在人及小鼠成纤维细胞中都生成了稳定的iPS细胞。然后,与2A结合在一起的重新编程因子(在已经生成的iPS细胞系中是不需要的)被除去。(生物谷Bioon.com)
生物谷推荐原始出处:
Nature 458, 766-770 (9 April 2009) | doi:10.1038/nature07863
piggyBac transposition reprograms fibroblasts to induced pluripotent stem cells
Knut Woltjen1, Iacovos P. Michael1,2, Paria Mohseni1,2, Ridham Desai1,2, Maria Mileikovsky1, Riikka H?m?l?inen1, Rebecca Cowling1, Wei Wang3, Pentao Liu3, Marina Gertsenstein1, Keisuke Kaji4, Hoon-Ki Sung1 & Andras Nagy1,2
1 Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario M5G 1X5, Canada
2 Department of Molecular Genetics, University of Toronto, Toronto M5S 1A8, Canada
3 The Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridgeshire CB10 1SA, UK
4 MRC Centre for Regenerative Medicine, Institute for Stem Cell Research, University of Edinburgh, Edinburgh EH9 3JQ, UK
Transgenic expression of just four defined transcription factors (c-Myc, Klf4, Oct4 and Sox2) is sufficient to reprogram somatic cells to a pluripotent state1, 2, 3, 4. The resulting induced pluripotent stem (iPS) cells resemble embryonic stem cells in their properties and potential to differentiate into a spectrum of adult cell types. Current reprogramming strategies involve retroviral1, lentiviral5, adenoviral6 and plasmid7 transfection to deliver reprogramming factor transgenes. Although the latter two methods are transient and minimize the potential for insertion mutagenesis, they are currently limited by diminished reprogramming efficiencies. piggyBac (PB) transposition is host-factor independent, and has recently been demonstrated to be functional in various human and mouse cell lines8, 9, 10, 11. The PB transposon/transposase system requires only the inverted terminal repeats flanking a transgene and transient expression of the transposase enzyme to catalyse insertion or excision events12. Here we demonstrate successful and efficient reprogramming of murine and human embryonic fibroblasts using doxycycline-inducible transcription factors delivered by PB transposition13. Stable iPS cells thus generated express characteristic pluripotency markers and succeed in a series of rigorous differentiation assays. By taking advantage of the natural propensity of the PB system for seamless excision12, we show that the individual PB insertions can be removed from established iPS cell lines, providing an invaluable tool for discovery. In addition, we have demonstrated the traceless removal of reprogramming factors joined with viral 2A sequences14 delivered by a single transposon from murine iPS lines. We anticipate that the unique properties of this virus-independent simplification of iPS cell production will accelerate this field further towards full exploration of the reprogramming process and future cell-based therapies.