Prevention of Arterial Thrombosis by Adenovirus-Mediated Transfer of Cyclooxygenase Gene

Abstract

Background Prostacyclin is an important vasoprotective molecule. It inhibits platelet aggregation, monocyte interaction with endothelium, and smooth muscle cell lipid accumulation. Vascular cyclooxygenase-1 (COX-1) is the rate-limiting step in prostacyclin synthesis. The objective of this study was to determine whether adenovirus-mediated transfer of COX-1 could restore COX-1 activity, augment prostacyclin synthesis, and prevent thrombus formation in a porcine carotid angioplasty model. Methods and Results Human COX-1 cDNA driven by a cytomegalovirus promoter was constructed into a replication-defective adenovirus 5 vector by homologous recombination. Recombinant adenovirus without a foreign gene (Ad-RR) and buffer were included as controls. Recombinant Ad-LacZ was used for marking the transfected cells in vivo. In the in vitro experiments, cultured human endothelial cells (ECs) and porcine arterial smooth muscle cells (SMCs) were incubated with Ad-COX-1 for 2 hours and 6-keto-PGF 1α level and the transgene expression were determined 72 hours after infection. In the in vivo experiments, recombinant adenoviruses were directly instilled into angioplasty-injured porcine carotid arteries for 30 minutes. Cyclic flow changes were monitored for 10 days and thrombus formation was examined histologically thereafter. Transgene expression and prostaglandin I 2 (PGI 2 ) synthesis by the injured arteries were determined. Cultured ECs infected with Ad-COX-1 produced a fivefold to eightfold increase in PGI 2 , and the transgene expression in cultured porcine SMCs was demonstrated by Northern analysis. Direct administration of Ad-COX-1 at a dose of 3×10 10 pfu completely inhibited carotid cyclic flow changes and thrombus formation accompanied by a fourfold increase in PGI 2 synthesis by the injured arteries 10 days after infection, whereas Ad-COX-1 at a lower dose, 5×10 9 pfu, had no antithrombotic effects when compared with Ad-RR vector and buffer controls. Conclusions Adenovirus-mediated transfer of COX-1 to angioplasty-injured carotid arteries was efficacious in augmenting PGI 2 synthesis and was associated with an inhibition of thrombosis when a relatively high titer of adenovirus was instilled.