%0 Journal Article
%T Reprogramming of Human Huntington Fibroblasts Using mRNA
%A Antje Arnold
%A Yahaira M. Naaldijk
%A Claire Fabian
%A Henry Wirth
%A Hans Binder
%A Guido Nikkhah
%A Lyle Armstrong
%A Alexandra Stolzing
%J ISRN Cell Biology
%D 2012
%R 10.5402/2012/124878
%X The derivation of induced pluripotent stem cells (iPS) from human cell sources using transduction based on viral vectors has been reported by several laboratories. Viral vector-induced integration is a potential cause of genetic modification. We have derived iPS cells from human foreskin, adult Huntington fibroblasts, and adult skin fibroblasts of healthy donors using a nonviral and nonintegrating procedure based on mRNA transfer. In vitro transcribed mRNA for 5 factors, oct-4, nanog, klf-4, c-myc, sox-2 as well as for one new factor, hTERT, was used to induce pluripotency. Reprogramming was analyzed by qPCR analysis of pluripotency gene expression, differentiation, gene expression array, and teratoma assays. iPS cells were shown to express pluripotency markers and were able to differentiate towards ecto-, endo-, and mesodermal lineages. This method may represent a safer technology for reprogramming and derivation of iPS cells. Cells produced by this method can more easily be transferred into the clinical setting. 1. Introduction The feasibility of reprogramming somatic cells to induced pluripotent stem cells (iPS) [1¨C4] has led to the possibility of developing disease-specific iPS cells for improved disease modeling in vitro [5¨C7] and potential use in clinical applications [8, 9]. Since the initial generation of iPS cells from mouse embryonic fibroblast (MEF) cells [1], there have been numerous refinements of the method. The potential therapeutic application of initial iPS cell lines was hampered by the fact that applied methods of iPS cell derivation modified the host genome through the integration of DNA sequences [3, 10¨C15]. Kim and colleagues [16] showed that it is possible to reprogram human foreskin fibroblasts through exposure to membrane-permeable recombinant proteins of the pluripotency factors Oct-4, Sox-2, Klf-4, and c-Myc. The factors were fused to a 9-arginine sequence to establish the ability of cell penetration. HEK 293 cells were transfected with plasmids for producing the described proteins. The whole HEK 293 cell extract was used for reprogramming. The method was refined by Zhou et al. [17] for MEF cells using recombinant cell-penetrating proteins. It has been shown that the modified mRNA-mediated delivery of reprogramming factors based on nucleofection is an efficient and nontoxic alternative approach to cell modification [18] which has recently facilitated the derivation of iPS cell lines [19¨C21]. Here, we investigate in vitro transcribed mRNA transfection as a method for producing iPS cells that does not bear any risk with respect
%U http://www.hindawi.com/journals/isrn.cell.biology/2012/124878/