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Nat Commun. 2018 Feb 21;9(1):745. doi: 10.1038/s41467-018-03190-3.

High-efficiency RNA-based reprogramming of human primary fibroblasts.

Author information

1
Department of Dermatology, University of Colorado School of Medicine, Anschutz Medical Campus, 12801 East 17th Avenue, Aurora, CO, 80045, USA.
2
Charles C. Gates Center for Regenerative Medicine, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO, 80045, USA.
3
Department of Biochemistry and Molecular Genetics, University of Colorado School of Medicine, Anschutz Medical Campus, 12801 East 17th Avenue, Aurora, CO, 80045, USA.
4
Department of Pediatrics, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO, 80045, USA.
5
Department of Immunology and Microbiology, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO, 80045, USA.
6
Department of Dermatology, University Medical Center, Groningen AB21 Hanzeplein 1, 9713, GZ Groningen, The Netherlands.
7
Department of Dermatology, University of Colorado School of Medicine, Anschutz Medical Campus, 12801 East 17th Avenue, Aurora, CO, 80045, USA. dennis.roop@ucdenver.edu.
8
Charles C. Gates Center for Regenerative Medicine, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO, 80045, USA. dennis.roop@ucdenver.edu.
9
Department of Dermatology, University of Colorado School of Medicine, Anschutz Medical Campus, 12801 East 17th Avenue, Aurora, CO, 80045, USA. ganna.bilousova@ucdenver.edu.
10
Charles C. Gates Center for Regenerative Medicine, University of Colorado School of Medicine, Anschutz Medical Campus, 12800 East 19th Avenue, Aurora, CO, 80045, USA. ganna.bilousova@ucdenver.edu.
11
Linda Crnic Institute for Down Syndrome, University of Colorado School of Medicine, Anschutz Medical Campus, 12700 East 19th Avenue, Aurora, CO, 80045, USA. ganna.bilousova@ucdenver.edu.

Abstract

Induced pluripotent stem cells (iPSCs) hold great promise for regenerative medicine; however, their potential clinical application is hampered by the low efficiency of somatic cell reprogramming. Here, we show that the synergistic activity of synthetic modified mRNAs encoding reprogramming factors and miRNA-367/302s delivered as mature miRNA mimics greatly enhances the reprogramming of human primary fibroblasts into iPSCs. This synergistic activity is dependent upon an optimal RNA transfection regimen and culturing conditions tailored specifically to human primary fibroblasts. As a result, we can now generate up to 4,019 iPSC colonies from only 500 starting human primary neonatal fibroblasts and reprogram up to 90.7% of individually plated cells, producing multiple sister colonies. This methodology consistently generates clinically relevant, integration-free iPSCs from a variety of human patient's fibroblasts under feeder-free conditions and can be applicable for the clinical translation of iPSCs and studying the biology of reprogramming.

PMID:
29467427
PMCID:
PMC5821705
DOI:
10.1038/s41467-018-03190-3
[Indexed for MEDLINE]
Free PMC Article

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