BACKGROUND: Nitric oxide, synthesized from L-arginine by nitric oxide synthase (NOS), is a vasodilator and inhibits vascular smooth muscle cell (SMC) proliferation and migration. The effects of local NOS gene transfer on restenosis after experimental balloon angioplasty were investigated. METHODS AND RESULTS: Left anterior descending coronary artery angioplasty was performed in 25 pigs. Animals received an intramural injection of adenovirus (1.5 x 10(9) pfu) carrying either the NOS cDNA (AdCMVceNOS) or no cDNA (AdRR5) via the Infiltrator. Local gene transfer efficiency and bioactivity of recombinant protein were assessed after 4 days. Indices of restenosis were evaluated by computerized planimetry on coronary artery sections prepared 28 days after angioplasty. Adenoviral vectors permitted efficient gene delivery to medial SMCs and adventitial cells of coronary arteries. Vascular cGMP levels were depressed after angioplasty from 1.30+/-0.42 to 0.33+/-0.20 pmol/mg protein (P<0.05) but were restored after constitutive endothelial (ce) NOS gene transfer to 1.82+/-0.98 pmol/mg (P<0.05 versus injured group and P=NS versus control). The ratio of the neointimal area to the internal elastic lamina fracture length, maximal neointimal thickness, and percent stenosis were all reduced in AdCMVceNOS- versus AdRR5-transduced pigs (0.59+/-0.14 versus 0.80+/-0.19 mm, P=0.02; 0.75+/-0.21 versus 1.04+/-0.25 mm, P=0.019; and 53+/-15% versus 75+/-11%, P=0.006, respectively). Lumen area was significantly larger (0.70+/-0.35 mm2 in AdCMVceNOS versus 0.32+/-0.18 mm2 in AdRR5, P=0.007). CONCLUSIONS: Percutaneous adenovirus-mediated NOS gene transfer resulted in efficient local overexpression of functional NOS after angioplasty in coronary arteries. Restored NO production in injured coronary arteries significantly reduced luminal narrowing, most likely through a combined effect on neointima formation and on vessel remodeling after angioplasty.