title: Efficient lentiviral transduction of human mesenchymal stem cells that preserves proliferation and differentiation capabilities.
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.
authors.affiliation: Department of Biomedical Engineering, Case Western Reserve University, Cleveland, OH, USA. pxl57@case.edu
authors.forename: Paul
authors.initials: P
authors.lastname: Lin
authors.forename: Yuan
authors.initials: Y
authors.lastname: Lin
authors.forename: Donald P
authors.initials: DP
authors.lastname: Lennon
authors.forename: Diego
authors.initials: D
authors.lastname: Correa
authors.forename: Mark
authors.initials: M
authors.lastname: Schluchter
authors.forename: Arnold I
authors.initials: AI
authors.lastname: Caplan
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D049109:Cell Proliferation; D002478:Cells, Cultured; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D006494:Heparin Antagonists / Q000494:pharmacology; D017919:Hepatitis B Virus, Woodchuck / Q000235:genetics; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D045164:Mesenchymal Stem Cell Transplantation / Q000379:methods*; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D016513:Mice, SCID; D011479:Protamines / Q000494:pharmacology; D050436:Regulatory Elements, Transcriptional / Q000235:genetics; D014161:Transduction, Genetic / Q000379:methods*; D019076:Transgenes / Q000235:genetics
chemical_list: D006494:Heparin Antagonists; D011479:Protamines; D016222:Fibroblast Growth Factor 2