Inflammation-induced transgene expression in genetically engineered equine mesenchymal stem cells.
BACKGROUND
Osteoarthritis, a chronic and progressive degenerative joint disorder, ranks amongst the top five causes of disability. Given the high incidence, associated socioeconomic costs and the absence of effective disease-modifying therapies of osteoarthritis, cell-based treatments offer a promising new approach. Owing to their paracrine, differentiation and self-renewal abilities, mesenchymal stem cells (MSCs) have great potential for regenerative medicine, which might be further enhanced by targeted gene therapy. Hence, the development of systems allowing transgene expression, particularly when regulated by natural disease-dependent occuring substances, is of high interest.


METHODS
Bone marrow-isolated equine MSCs were stably transduced with an HIV-1 based lentiviral vector expressing the luciferase gene under control of an inducible nuclear factor κB (NFκB)-responsive promoter. Marker gene expression was analysed by determining luciferase activity in transduced cells stimulated with different concentrations of interleukin (IL)-1β or tumour necrosis factor (TNF)α.


RESULTS
A dose-dependent increase in luciferase expression was observed in transduced MSCs upon cytokine stimulation. The induction effect was more potent in cells treated with TNFα compared to those treated with IL-1β. Maximum transgene expression was obtained after 48 h of stimulation and the same time was necessary to return to baseline luciferase expression levels after withdrawal of the stimulus. Repeated cycles of induction allowed on-off modulation of transgene expression without becoming refractory to induction. The NFκB-responsive promoter retained its inducibility also in chondrogenically differentiated MSC/Luc cells.


CONCLUSIONS
The results of the present study demonstrate that on demand transgene expression from the NFκB-responsive promoter using naturally occurring inflammatory cytokines can be induced in undifferentiated and chondrogenically differentiated equine MSCs. Copyright © 2016 John Wiley & Sons, Ltd.
Institute of Anatomy, Histology and Embryology, University of Veterinary Medicine Vienna, Vienna, Austria.
Simone
S
Gabner
Institute of Anatomy, Histology and Embryology, University of Veterinary Medicine Vienna, Vienna, Austria.
Juraj
J
Hlavaty
Equine University Hospital, University of Veterinary Medicine Vienna, Vienna, Austria.
Karsten
K
Velde
Division of Medical Biotechnology, Paul-Ehrlich-Institut, Langen, Germany.
Matthias
M
Renner
Equine University Hospital, University of Veterinary Medicine Vienna, Vienna, Austria.
Florien
F
Jenner
Institute of Anatomy, Histology and Embryology, University of Veterinary Medicine Vienna, Vienna, Austria.
Monika
M
Egerbacher
D000818:Animals; D001854:Bone Marrow Cells / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000187:drug effects / Q000235:genetics; D016207:Cytokines / Q000494:pharmacology; D015870:Gene Expression; D005818:Genetic Engineering / Q000379:methods*; D006736:Horses; D006801:Humans; D007249:Inflammation / Q000235:genetics*; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D016328:NF-kappa B / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D019076:Transgenes / Q000235:genetics*
D016207:Cytokines; D016328:NF-kappa B; D008156:Luciferases
Arthritis; inducible promoter; mesenchymal stem cells; regulated transgene expression