Pressure-mediated oligonucleotide transfection of rat and human cardiovascular tissues

The application of gene therapy to human disease is currently restricted by the relatively low efficiency and potential hazards of methods of oligonuc leotide or gene delivery. Antisense or transcription factor decoy oligonucl eotides have been shown to be effective at altering gene expression in cell culture experiments, but their ill vivo application is limited by the effi ciency of cellular delivery, the intracellular stability of the compounds, and their duration of activity. We report herein the development of a highl y efficient method for naked oligodeoxynucleotide (ODN) transfection into c ardiovascular tissues by using controlled, nondistending pressure without t he use of viral vectors, lipid formulations, or exposure to other adjunctiv e, potentially hazardous substances. In this study, we have documented the ability of ex vivo, pressure-mediated transfection to achieve nuclear local ization of fluorescent (FITC)-labeled ODN in approximately 90% and 50% of c ells in intact human saphenous vein and rat myocardium, respectively. We ha ve further documented that pressure mediated delivery of antisense ODN can functionally inhibit target gene expression in both of these tissues in a s equence-specific manner at the mRNA and protein levels. This oligonucleotid e transfection system may represent a safe means of achieving the intraoper ative genetic engineering of failure-resistant human bypass grafts and may provide an avenue for the genetic manipultation of cardiac allograft reject ion, allograft vasculopathy, or other transplant diseases.