Objective: Nitric oxide regulates vascular tone, inhibits platelet aggregation, and inhibits leukocyte adhesion, all of which are important modulators of ischemia-reperfusion injury. This study aimed to determine the effects of endothelial constitutive nitric oxide synthase gene transfer on ischemia-reperfusion injury in a rat lung transplant model. Methods: In group I, donor animals were injected intravenously with 5 x 109 pfu of adenovirus- encoding endothelial constitutive nitric oxide synthase. Groups II and III served as controls, whereby donor animals were injected with either 5 x 109 pfu of adenovirus encoding β-galactosidase or saline solution, respectively. Twenty-four hours after injection, left lungs were harvested and preserved for 18 hours at 4°C, then implanted into isogeneic recipients, which were put to death 24 hours later. Recombinant endothelial constitutive nitric oxide synthase gene expression was evaluated by Western blotting and immunohistochemistry. Lung grafts were assessed by measuring arterial oxygenation, myeloperoxidase activity, and wet/dry weight ratios. Results: Western blotting confirmed the overexpression of endothelial constitutive nitric oxide synthase in lungs so transfected compared with controls. Twenty- four hours after reperfusion, mean arterial oxygenation was significantly improved in group I compared with group H and HI controls (189.4 ± 47.1 mm Hg vs 71.7 ± 8.9 mm Hg and 67.8 ± 12.2 mm Hg, P = .02, P = .01, respectively). Myeloperoxidase activity, a reflection of tissue neutrophil sequestration, was also significantly reduced in group I compared with groups H and III (0.136 ± 0.038 ΔOD/mg/min vs 0.587 ± 0.077 and 0.489 ± 0.126 ΔOD/mg/min, P = .001, P = .01, respectively). Conclusion: Adenovirus- mediated gene transfer with endothelial constitutive nitric oxide synthase ameliorates ischemia-reperfusion injury as manifested by significantly improved oxygenation and decreased neutrophil sequestration in transplanted lung isografts. Endothelial constitutive nitric oxide synthase gene transfer may reduce acute lung dysfunction after lung transplantation.