Abstract
BackgroundIn this study, we have examined local non-viral gene delivery, transfection, and therapeutic efficacy of endothelial nitric oxide synthase (eNOS) encoding plasmid DNA administered using coated stents in a rabbit iliac artery restenosis model.MethodsLipopolyplexes (LPPs) with eNOS expressing plasmid DNA were immobilized on stainless steel stents using poly(D,L-lactide-co-glycolide) (PLGA) and type B gelatin coatings. The gene-eluting stents were implanted bilaterally in the denuded iliac arteries and eNOS transfection and therapeutic efficacy were examined 14 days after implantation.ResultsThe results show that non-viral lipopolyplex-coated stents can efficiently tranfect eNOS locally in the arterial lumen assessed by PCR and ELISA. Human eNOS ELISA levels were significantly raised 24 hours after transfection compared to controls (125 pg eNOS compared to <50 pg for all controls including naked DNA). Local eNOS production suppressed smooth muscle cell proliferation and promoted re-endothelialization of the artery showing a significant reduction in restenosis of 1.75 neointima/media ratio for stents with lipoplexes encoding eNOS compared with 2.3 neointima/media ratio for stents with lipoplexes encosing an empty vector.ConclusionsThese results support the hypothesis that a potent non-viral gene vector encoding for eNOS coated onto a stent can inhibit restenosis through inhibition of smooth muscle cell growth and promotion of a healthy endothelium.
Highlights
Over 24 million individuals are diagnosed with a form of heart disease every year and cardiovascular disease and its complications continue to be associated with the highest rate of mortality in the United States [1]
Stent Coatings and In Vitro Characterization Plasmid DNA expressing endothelial nitric oxide synthase [14] was cloned in E. coli, amplified, and purified at Elim Biopharmaceuticals (Hayward, CA). pVAX-1 was purchased from
LPPs were prepared according to an optimized method, published previously [16], using endothelial nitric oxide synthase (eNOS) expressing plasmid complexed with poly(beta-amino ester) (PbAE) and encapsulated in cationic phospholipid, 1,2-dioleoyl3-trimethylammonium propane (DOTAP)
Summary
Over 24 million individuals are diagnosed with a form of heart disease every year and cardiovascular disease and its complications continue to be associated with the highest rate of mortality in the United States [1]. As the plaque size increases it can become unstable and rupture, triggering a clotting cascade that can occlude the vessel, obstructing blood flow. The obstruction of blood flow in the coronary artery leads to myocardial ischemia and infarction, and can be fatal. Removal of the plaque deposit is commonly performed by catheter-based non-invasive techniques, such as percutaneous transluminal coronary angioplasty, atherectomy and stent implantation procedures. All of these procedures can lead to a complication called restenosis, where the vessel diameter decreases due to proliferation of smooth muscle cells or neointimal hyperplasia in the lumen of the artery [3,4]. We have examined local non-viral gene delivery, transfection, and therapeutic efficacy of endothelial nitric oxide synthase (eNOS) encoding plasmid DNA administered using coated stents in a rabbit iliac artery restenosis model
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