Nicht-virale Gentherapie verbessert Regeneration im ischämischen Lappenmodell der Ratte Non-viral gene therapy for regeneration in an ischemic flap model

Backround: Protein delivery from transfected cells can induce expression of tissue inductive factors to stimulate the cellular processes required for regeneration. We established a cell-based, non viral gene-transfer method using fibroblasts to temporarily produce bFGF and VEGF 165 , as a form of pharmacological local preconditioning before tissue ischemia occurs. Material and Methods: The eukaryotic expression vectors harboring VEGF and bFGF cDNAs were transfected into rat primary skin fibroblasts mediated by Amaxa Nucleofector and optimized by our own laboratory protocol. To determine an improvement in ischemically challenged tissue, a genetically modified cellspool was injected into the target tissue 1 week before inducing an ischemic flap model. Gene expression and protein production in vivo and in vitro were measured by real time PCR and immunoassay (BioPlex) respectively. Clinical outcome was demonstrated by planimetric measurements. Results: Temporary protein expression of bFGF and VEGF 165 in the target tissue of the ischemic flap model increased compared to controls after injection of genetically modified cells. A highly significant improvement of tissue survival was observed after the transfected cell administration. A reduction in flap necrosis by one-third or more was detected after two weeks if transfected cells were applied 1 week before ischemia. Conclusion: In our work we showed that temporary expression of bFGF and VEGF 165 induces therapeutically relevant effects in the rat flap model of ischemia. Our standardized high efficiency non viral bFGF and VEGF 165 transfection technology is now used in preclinical research.