The use of designed, solid free form scaffolds for gene therapy directed bone tissue engineering

摘要

Varied clinical problems have created a need to repair and regenerate bone. Tissue engineering is an alternative to conventional therapies that may allow the generation of functional tissue. The hypothesis of this work is that combining solid freeform fabricated scaffolds and localized gene therapy will result in the formation of biologically and mechanically functional bone throughout the scaffold. Initial experiments focused on the use of fibrin and collagen hydrogels to suspend and deliver adenoviral vectors. Results showed that the hydrogels maintained or extended the activity of adenoviral vectors in vitro. In vivo, gels containing adenovirus expressing bone morphogenetic protein-7 led to ectopic bone formation. In vivo experiments were undertaken to optimize the combination of scaffolds and gene therapy, focusing on the type of gene therapy (in vivo or ex vivo), the carrier (hydrogel or polymer), and the scaffold pore size. In vivo implantation triggered the generation of bone with marrow space and vasculature. The variables tested had various effects on bone generation: gene therapy method had the largest effect, with ex vivo gene therapy generating significantly more bone than in vivo gene therapy; carrier type also had a significant effect, with scaffolds seeded with fibrin gel showing more bone generation; pore size did not have a significant effect. The results demonstrated the conditions best suited to maximize the formation of bone and in the final phase of the thesis these conditions were combined with topology optimized scaffolds. These scaffolds were implanted in vivo and then evaluated mechanically and histologically. Over extended implantation times, large amounts of bone formed. Though the bone volume was relatively constant over time, it was increasingly localized on the scaffold contours as time increased. The compressive modulus of the bone increased as the modulus of the degrading scaffold declined. Thus, the overall modulus of the implant was nearly constant, an excellent indicator that such an implant could continuously function in a load bearing site. These results show designed solid freeform fabricated scaffolds combined with optimized gene therapy can be used to achieve rapid osteogenesis and maintain appropriate mechanical properties, thereby meeting the essential requirements for a bone tissue engineering implant.