VASCULAR GENE-TRANSFER OF THE HUMAN INDUCIBLE NITRIC-OXIDE SYNTHASE -CHARACTERIZATION OF ACTIVITY AND EFFECTS ON MYOINTIMAL HYPERPLASIA

Background: Nitric oxide (NO) has been shown to decrease myointimal hy perplasia in injured blood vessels. We hypothesize inducible NO syntha se (iNOS) gene transfer even at low efficiency will provide adequate l ocal NO production to achieve this goal. Materials and Methods: A retr oviral vector containing the human iNOS cDNA (DFGiNOS) was used to tra nsfer the iNOS gene into vascular cells and isolated blood vessels to answer the following questions: can vascular endothelial and smooth mu scle cells support iNOS activity and will low efficiency iNOS gene tra nsfer suppress myointimal hyperplasia in injured porcine arteries? Res ults: DFGiNOS-infected sheep pulmonary artery endothelial cells (SPAEC ) expressed significant iNOS mRNA and protein, releasing nitrite level s of 155.0 +/- 10.7 nmol/mg protein/24 h vs. 5.5 +/- 1.1 by control ce lls. Transduced rat smooth muscle cells (RSMC) also expressed abundant iNOS mRNA and protein, but, in contrast to SPAEC, NO synthesis was de pendent on exogenous tetrahydrobiopterin (BH4) (291.8 +/- 10.4 nmol ni trite/mg protein/24 hr with BH4, 37.7 +/- 2.6 without BH4). Only porci ne arteries infected with DFGiNOS following balloon injury exhibited a 3-fold increase in total NO synthesis and a 15-fold increase in cGMP levels over control vessels in a BH4 dependent fashion, despite only a 1% gene transfer efficiency. Transfer of iNOS completely prevented th e 53% increase in myointimal thickness induced by balloon catheter inj ury; the administration of a NOS inhibitor reversed this effect. Concl usions: These in vitro findings suggest that vascular iNOS gene transf er may be feasible. Furthermore, a low gene transfer efficiency may be sufficient to inhibit myointimal hyperplasia following arterial ballo on injury, although a source of BH4 may be required.