Avian models of atherosclerosis helped pioneer the study of vascular biology, and offer economic and technical advantages over mammalian models. As an initial step towards investigating important molecular pathways involved in avian atherogenesis and restenosis, we developed a recombinant adenovirus (Ad) which expresses the reporter gene beta-galactosidase (beta-gal), and applied it to cultured chicken vascular smooth muscle cells (SMCs) and a rooster model of acute vascular injury. In cultured chicken SMCs, recombinant gene expression increased as a function of multiplicity of infection (MOI) and incubation time. Maximal expression occurred at an MOI of 10(4) plaque-forming units (pfu)/cell with approximately 50% of quiescent and non-quiescent chicken SMCs expressing beta-gal. Human aorta SMCs had two- to four-fold increased beta-gal expression compared with chicken SMCs at all MOI and incubation times. In vivo instillation of recombinant Ad into uninjured rooster femoral artery segments revealed low efficiency endothelial cell expression of the reporter gene. In contrast, recombinant Ad infection of rooster femoral artery segments 3-21 days after balloon injury revealed up to 60% of luminal surface beta-gal expression, confined predominantly to the neointimal layer. Peak reporter gene expression efficiencies occurred in arterial segments infected 3 days after balloon injury. Uninfected and control Ad infected arteries had no detectable beta-gal expression. Rooster neointimal cells targeted by the recombinant Ad were identified as alpha-smooth muscle actin containing cells by immunohistochemistry. We conclude that Ad-mediated gene transfer is efficient and selective for the neointima in the rooster acute vascular injury model, and offers the potential to efficiently introduce exogenous genes that may impact on the injury response. This model of acute vascular injury may also be manipulated into more established avian models of atherosclerosis, permitting the investigation of acute injury progression to chronic injury.