Tissue Engineering for Bone Regeneration Using Osteoblasts in Collagen Scaffolds

Objectives: Regeneration of osseous defects using a tissue engineering approach provides a novel means of treatment utilizing cell biology, materials science and molecular biology. In this study we tested the concept that artificial matrices, seeded with cells of osteogenic potential, may be implanted into sites where osseous damage has occurred and lead to significant osseous regeneration. Methods: Explant cultures of cells from human alveolar bone, gingiva and periodontal ligament were established. When seeded into a 3-dimensional collagen-based scaffold, the bone-derived cells maintained their osteoblast phenotype as monitored by gene (real time PCR) and protein (immunohistochemistry) expression of bone related proteins including bone sialoprotein, osteocalcin, osteopontin, bone morphogenetic protein 2 &4, and alkaline phosphatase. These in vitro-developed matrices, consisting of cultured osteoblasts and newly synthesized extracellular matrix within the collagen scaffold, were transplanted into critical-size bone defects sites in skulls of immunodeficient (SCID) mice. Wound healing was followed for up to 4 weeks. Results: The bone density in the defect areas filled with the osteoblast-derived matrix was significantly higher compared with the defects filled with either collagen scaffold alone or collagen scaffold impregnated with human gingival fibroblasts. New bone formation was identified in all of the sites treated with the osteoblast-derived matrix, while no obvious new bone formation was noted in the control groups. In situ hybridization for the human-specific alu gene sequence indicated that the newly formed bone tissue resulted from the activity of the grafted human cells. Conclusions: The results indicate that cells derived from human alveolar bone can be incorporated into bioengineered scaffolds and synthesize a matrix which can be implanted to induce new bone formation. This study was supported by a grant from the National Health & Medical Research Council of Australia.