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Stem Cells Transl Med. 2012 Dec;1(12):886-97. doi: 10.5966/sctm.2012-0086. Epub 2012 Nov 29.

Efficient lentiviral transduction of human mesenchymal stem cells that preserves proliferation and differentiation capabilities.

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1
Department of Biomedical Engineering, Case Western Reserve University, Cleveland, OH, USA. pxl57@case.edu

Abstract

Long-term lentiviral transduction of human mesenchymal stem cells (hMSCs) greatly enhances the usefulness of these cells. However, such transduction currently requires the use of polybrene, which severely inhibits hMSC proliferation. In contrast, protamine sulfate at 100 μg/ml doubled transduction efficiencies without affecting proliferation or differentiation potential. Expression levels improved 2.2-fold with the addition of a woodchuck hepatitis post-transcriptional regulatory element. Further improvements in transduction efficiencies could be obtained by a modest increase in viral concentrations through increased viral titers or decreased transduction volumes without changing multiplicity of infection, by transducing over multiple days, or by culturing the cells in fibroblast growth factor-2. Centrifugation improved expression but had no effect on efficiency. Transgene expression was stable over 6 weeks in vitro and in vivo. Donor-to-donor and intradonor variability were observed in primary passage through passage 2 cultures, but not at passage 3. These results provide a better optimized approach for expanded use of hMSCs through genetic manipulation.

PMID:
23283550
PMCID:
PMC3659678
DOI:
10.5966/sctm.2012-0086
[Indexed for MEDLINE]
Free PMC Article
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