The major goal of this project is to improve the performance of intravascular prosthetic devices by seeding these devices with genetically altered endothelial cells. Failure of intravascular prostheses due to thrombosis and neointimal hyperplasia is a significant clinical problem for which current therapeutic approaches are largely ineffective. During the past year we continued our work with seeding of prosthetic vascular grafts and intravascular stents. Following successful in vitro cell retention studies, we implanted seeded stents into the femoral arteries of living sheep. 10 days after implantation, patency and flow dynamics of stented segments were analyzed by color Doppler. Stents were removed and subjected either to scanning electron microscopy or (along with adherent tissue) to DNA extraction to determine retention of the seeded cells. Stent coverage with endothelium was approximately 50%. Seeded cell retention was established by PCR analysis of extracted DNA. A similar autologous implantation system has been developed for seeded vascular grafts. Three prosthetic graft materials were compared for their ability both to be seeded with transduced endothelial cells and to retain these cells under flow conditions in vitro. Sheep endothelial cells transduced with the human t-PA gene were seeded onto 3 different prosthetic graft materials. Knitted Dacron grafts permitted the highest levels of recombinant t-PA expression; this level is approximately 1 log above the rate of t-PA secretion thought to be present from a normal vessel segment. We have begun implantation of the seeded grafts into sheep carotid arteries and have found significant retention of viable transduced cells 2 hours after implantation (the only time point studied so far). Following optimization of cell retention at 2 hours we will examine cell retention at later time points.
