Effective, safe nonviral gene transfer to preserve the chondrogenic differentiation potential of human mesenchymal stem cells.
BACKGROUND
Genetic modification of mesenchymal stem cells (MSCs) comprises a promising tool to generate cell- and gene-based platforms for regenerative approaches of articular cartilage repair. In the present study, we systematically screened a panel of 15 nonviral compounds for their ability to promote safe, efficient and durable gene expression in human bone marrow-derived MSCs (hMSCS) without impeding their commitment towards chondrogenic differentiation.


METHODS
Primary hMSCs were transfected with plasmid vectors carrying sequences for the Photinus pyralis luciferase Escherichia coli β-galactosidase, or human insulin-like growth factor I via 15 nonviral formulations. Transgene expression and transfection efficiencies were monitored for each component in parallel with the effects on cell viability and cytotoxicity. Upon optimization, the most promising reagent was then evaluated for a possible influence on the chondrogenic potential of hMSCs.


RESULTS
Among all formulations tested, GeneJammer® gave the best results for transgene expression and transfection efficacy (25-14% from days 2-21 in monolayer cultures and 35% in 21-day aggregate cultures), allowing for high levels of viability (92-94%) and modest cytotoxicity (< 12%). Most notably, the application of this reagent did not affect the potential of the cells for chondrogenic differentiation when maintained in long-term (21 days) three-dimensional (aggregate) cultures.


CONCLUSIONS
The data indicate that safe, efficient transgene expression can be achieved in hMSCs over time using the nonviral GeneJammer® compound, showing promise for future therapeutic settings aiming to treat human articular cartilage disorders.
Center of Experimental Orthopaedics, Saarland University Medical Center, Saarland University, Homburg/Saar, Germany.
Sebastian
S
Elsler
Sarah
S
Schetting
Gertrud
G
Schmitt
Dieter
D
Kohn
Henning
H
Madry
Magali
M
Cucchiarini
D002358:Cartilage, Articular / Q000378:metabolism; D002449:Cell Aggregation / Q000187:drug effects / Q000235:genetics; D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics*; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000187:drug effects*; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000187:drug effects / Q000502:physiology*; D010957:Plasmids; D011073:Polyamines / Q000494:pharmacology*; D023822:Tissue Engineering / Q000379:methods*; D014162:Transfection / Q000379:methods*; D019076:Transgenes / Q000235:genetics
D011073:Polyamines; C522645:genejammer