The search for vessel substitutes to replace small-/medium-caliber vessels is an ongoing concern for vascular surgeons. Engineered vessels were designed for use as arterial equivalents and assessed in an in vivo model in dog. Three study groups were established: clinical expanded polytetrafluoroethylene (ePTFE; control, n = 24), ePTFE seeding with endothelial cells (EC graft, n = 12), and ePTFE with a fibroblast matrix seeded with EC (FM+EC graft, n = 12). Grafts were subjected to a custom-designed femoral ex vivo circuit and implanted in the carotid artery for 60 days. The viability of the prosthetic grafts was evaluated. The ex vivo circuit revealed that the presence of a fibroblast matrix induced over double the cell retention compared to EC grafts. A significant reduction in platelet adhesion in EC grafts was observed. After their in vivo implantation, the engineered vessels were more efficient at avoiding occlusion than the prosthetic grafts. The FM+EC grafts induced more endothelialization than those seeded with ECs alone. Intimal hyperplasia response was reduced in the EC substitutes. Significant differences in apoptotic cells emerged between the EC and control ePTFE grafts. In conclusion, engineered vessels showed improved initial patency over ePTFE grafts. The EC graft was best at combating restenosis, a good indicator of the long-term efficiency of the graft.