Using microRNAs to enhance the generation of induced pluripotent stem cells.

Program for RNA Biology, Sanford-Burnham Medical Research Institute, La Jolla, California, USA.
Current protocols in stem cell biology 03/2012; Chapter 4:Unit 4A.4. DOI: 10.1002/9780470151808.sc04a04s20
Source: PubMed

ABSTRACT Somatic cells reprogrammed to acquire an ES-like state are termed iPS cells. In this unit, a protocol to use microRNAs as enhancers to increase the reprogramming efficiency is described. Mouse embryonic fibroblasts (MEFs) are isolated from E13.5 mouse embryos and seeded for reprogramming by defined factors. microRNA mimics are transfected into MEFs at two time points during this process to enhance the overall reprogramming efficiency. Two standard protocols for characterization of these miR-iPSCs, embryoid body formation and teratoma formation, are also provided. By using this method, the investigators can obtain a significantly higher number of bona-fide iPSC colonies and miR-iPSCs can be derived at a faster rate than with non-treated cells.

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    • "Work from Ali et al. clearly demonstrates that the post-mitotic photoreceptor precursor cell is the optimal cell type for efficient rod photoreceptor cell replacement (Lakowski et al., 2011; Pearson et al., 2012). A variety of different protocols, utilizing both two-and three-dimensional culture systems, have succeeded in deriving photoreceptor precursor cells from less differentiated precursors (Osakada et al., 2008; Hirami et al., 2009; Meyer et al., 2009; Osakada et al., 2009; Lamba et al., 2010; Meyer et al., 2011; Tucker et al., 2011; Nakano et al., 2012; Phillips et al., 2012; Sasai et al., 2012; Homma et al., 2013; Mekala et al., 2013; Tucker et al., 2013). Although cultured three-dimensional eyecups will undoubtedly have many applications in developmental biology (Nakano et al., 2012; Sasai et al., 2012), two-dimensional systems have the advantage of easier identification and isolation of specific cell types for post-differentiation subculture. "
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    eLife Sciences 08/2013; 2:e00824. DOI:10.7554/eLife.00824 · 8.52 Impact Factor
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    ABSTRACT: There are an estimated 7,000 "orphan diseases," but treatments are currently available for only about 5% of them. Recent progress in the advanced platforms of gene therapy, stem cell therapy, gene modification, and gene correction offers possibilities for new therapies and cures for rare diseases. Many rare diseases are genetic in origin, and gene therapy is being successfully applied to treat them. Human stem cell therapy, apart from bone marrow transplants, is still experimental. Genetic modification of stem cells can make stem cell-based products more effective. Autologous induced pluripotent stem (iPS) cells, when combined with new classes of artificial nucleases, have great potential in the ex vivo repair of specific mutated DNA sequences (zinc-finger proteins and transactivator-like effector nucleases). Patient-specific iPS cells can be corrected and transplanted back into the patient. Stem cells secrete paracrine factors that could become new therapeutic tools in the treatment of orphan diseases. Gene therapy and stem cell therapy with DNA repair are promising approaches to the treatment of rare, intractable diseases.
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