Canine endothelial progenitor cell-lined hybrid vascular graft with nonthrombogenic potential

Objective: We sought to fabricate a compliant engineered vascular graft (inner diameter of approximately 4.5 mm and length of 6 cm) lined with endothelial progenitor cells derived from circulating peripheral canine blood and to verify its nonthrombogenicity potential in vivo. Methods: Autologous circulating endothelial progenitor cells derived from the peripheral veins of 6 adult mongrel dogs were isolated by using a density gradient method. The cells were proliferated in vitro in EGM-2 culture medium, prelined on the luminal surface of in situ-formed collagen type I meshes as an extracellular matrix, and wrapped with a segmented polyurethane thin film with multiple micropores as a compliant scaffold. After canine carotid arteries were bilaterally implanted with these grafts for I and 3 months, microscopic observation, histologic staining, and immunochemical staining were performed to evaluate morphogenesis. Results: After 33.3 +/- 10.5 days of culture in vitro, 4.2 +/- 1.2 x 10(6) endothelial progenitor cells were obtained. Eleven of the 12 engineered vascular grafts were patent. The grafts possessed smooth, glistening, and ivory-colored luminal surfaces at the predetermined observation period up to 3 months. The intimal layer was covered with confluent, cobblestone-like monolayered cells that were positively stained with factor VIIIB-related antigen. The thickness of the neoarterial walls was approximately 300 mum at 3 months after implantation. A few smooth muscle cells were observed in the medial tissue, and fibroblasts dominated the adventitial tissue. Conclusion: Circulating endothelial progenitor cells could be a substitute source of endothelial cells for endothelialization on small-diameter-vessel prostheses to ensure nonthrombogenicity.