Adenovirus-mediated gene transfer of human inducible nitric oxide synthase in porcine vein grafts inhibits intimal hyperplasia

Abstract

Objective: The aim of this study is to determine whether adenoviral inducible nitric oxide synthase (iNOS) gene transfer could inhibit intimai hyperplasia (IH) in porcine internal jugular veins interposed into the carotid artery circulation. Methods: Porcine internal jugular veins were transduced passively with 1 × 1011 particles of an adenoviral vector carrying either the human iNOS (AdiNOS) or β-galactosidase (AdlacZ) cDNA for 30 minutes and then interposed into the carotid artery circulation. Segments of each vein graft were maintained in an ex vivo organ culture to measure nitrite accumulation, a marker of nitric oxide synthesis. The grafts were analyzed immunohistochemically for the presence of neutrophils, macrophages, and leukocytes by staining for myeloperoxidase, EDI, and CD45, respectively, at 3 (n = 4) and 7 (n = 4) days. Morphometric analyses and cellular proliferation (Ki67 staining) were assessed at 3 (n = 4), 7 (n = 4), and 21 days (n = 8). Results: AdlacZ-treated vein grafts demonstrated high levels of β-galactosidase expression at 3 days with a gradual decline thereafter. Nitrite production from AdiNOS-treated vein grafts was approximately fivefold greater than AdlacZ-treated grafts (P = .00001). AdiNOS or AdlacZ treatment was associated with minimal graft inflammation. Cellular proliferation rates were significantly reduced in AdiNOS-treated grafts as compared with controls at both 3 (41%, P = .000004) and 7 days (32%, P = .0001) after bypass. This early antiproliferative effect was most pronounced at the distal anastomosis (65%, P = .0005). The iNOS gene transfer reduced the intimal/medial area ratio in vein grafts at 7 (36%, P = .009) and 21 days (30%, P = .007) versus controls. This inhibition of IH was again more prominent in the distal segments of the grafts (P = .01). Conclusion: Adenovirus-mediated iNOS gene transfer to porcine internal jugular vein grafts effectively reduced cellular proliferation and IH. Although iNOS gene transfer reduced IH throughout the entire vein graft, the most pronounced effect was measured at the distal anastomosis. These results suggest potential for iNOS-based genetic modification of vein grafts to prolong graft patency.