Abstract
Human induced pluripotent stem cells (hiPSCs1,2,3) are useful in disease modeling and drug discovery, and they promise to provide a new generation of cell-based therapeutics. To date there has been no systematic evaluation of the most widely used techniques for generating integration-free hiPSCs. Here we compare Sendai-viral (SeV)4, episomal (Epi)5 and mRNA transfection mRNA6 methods using a number of criteria. All methods generated high-quality hiPSCs, but significant differences existed in aneuploidy rates, reprogramming efficiency, reliability and workload. We discuss the advantages and shortcomings of each approach, and present and review the results of a survey of a large number of human reprogramming laboratories on their independent experiences and preferences. Our analysis provides a valuable resource to inform the use of specific reprogramming methods for different laboratories and different applications, including clinical translation.
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Acknowledgements
We would like to thank B. Hamilton (Stemgent) for miRNA reprogramming agents and protocols, G. Mostoslavsky (Boston University) for human STEMCCA lentiviral plasmid constructs, M. Armant (Boston Children's Hospital) for MRC5 and hCD34+ cells, G. MacLean (Boston Children's Hospital) for advice on episomal blood reprogramming and S. D'Souza (Icahn School of Medicine at Mount Sinai) for a list of contacts for human cell reprogramming laboratories. This work was supported in part by grants R01HL75737, U01HL107440, UO1-HL100001, U01HL87402 and U01HL100408 (National Heart, Lung, and Blood Institute (NHLBI) Progenitor Cell Biology Consortium), R24DK092760 (the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)), PO1NS066888 (the National Institute of Neurological Disorders and Stroke (NINDS)), by the SMA Foundation, the Jerome Le Jeune Foundation, an EMBO postdoctoral fellowship (E.B.) and the Harvard Stem Cell Institute.
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T.M.S., L.D., C.A.C., A.M., L.L.R., A.P.F., L.I.Z. and G.Q.D. contributed to the conception and design of the study; T.M.S., L.D. and G.Q.D. wrote the manuscript; T.R.B., S.E., K.C., A.C., A.D., A.E., K.F., M.G., D.G., J.M., P.M., M.G. and B.B. performed reprogramming and cell culture experiments; A.D., N.J., X.L. and A.P.F. performed epigenetics analyses, T.M.S., K.F., D.G., P.M., L.D., T.R.B. and S.E. performed PCR assays; M.S.L., E.B., A.B.C.C. and D.D. provided cell samples and reprogramming data; A.M.T. and A.M. performed the scorecard analysis; T.M.S. performed statistical analyses; T.M.S. and L.D. performed the survey.
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Schlaeger, T., Daheron, L., Brickler, T. et al. A comparison of non-integrating reprogramming methods. Nat Biotechnol 33, 58–63 (2015). https://doi.org/10.1038/nbt.3070
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DOI: https://doi.org/10.1038/nbt.3070
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