Implanted myoblast survival is dependent on the degree of vascularization in a novel delayed implantation/prevascularization tissue engineering model

In in vivo tissue engineering, many implanted cells die because of hypoxic conditions immediately postimplantation. The aim of this study was to determine whether delayed myoblast implantation, at day 4 or 7, improves myoblast survival compared with implantation at day 0 in an in vivo arterio-venous loop (AB loop) chamber model. In adult inbred Sprague-Dawley rats, an AB loop was inserted into a plastic chamber (day 0). In Group I, day 0, two million DiI-labeled (neonatal inbred) myoblasts were implanted around the AB loop. In Groups II and III, day 0, the AB loop was created and inserted into a novel delayed cell seeding chamber, and 4 (Group II) or 7 days (Group III) later the delay chamber was seeded with 2 million DiI-labeled myoblasts. Constructs were harvested 7-day postmyoblast implantation, for morphometric determination of DiI= DAPI-positive myoblasts=mm2, and percent vascular volume on Griffonia simplicifolia lectin (endothelial cell marker)–labeled tissue sections. Control (nonmyoblast seeded) and experimental (myoblast seeded) constructs demonstrated similar capillary and tissue growth patterns. DiI=DAPI-labeled myoblasts=mm2 appeared in similar numbers in constructs implanted at days 0 and 4, but increased markedly in day-7 implanted constructs. The percent vascular volume increased significantly ( p¼0.03) over time. A positive correlation existed between myoblast survival and construct vascularity ( p¼0.017). In conclusion, delaying myoblast implantation to 7-day postconstruct assembly, when new capillary growth is well established, significantly correlates with increased myoblast survival and indicates that cell seeding in regenerative procedures should always occur into an established vascular bed.