The Effect of an Admix BMPs on Human Osteoprogenitor Activity

Objectives: The ability to generate new bone has tremendous therapeutic implications. Biomimetic scaffolds generated from appropriate matrices with judiciously selected osteoinductive factors which promote human osteoprogenitor differentiation and osteogenesis offers a promising approach in the regeneration of skeletal tissue. BMPs are key osteoinductive factors pivotal in the recruitment, commitment and differentiation of mesenchymal cells. However, the ideal combination of BMPs for bone formation remains unclear. The aims of this study were to examine the efficacy of 'retentate', an osteoinductive extract of Saos-2 cells, on human bone marrow stromal cells (HBMSCs). Methods: HBMSCs were cultured in presence or absence of 'retentate' in vitro. Cell attachment and patterning were examined by CFU-F formation. Cell growth on 3-D PLGA scaffolds were assayed using live/dead fluorescent markers, confocal microscope and SEM. Cell proliferation and differentiation were examined by DNA content and alkaline phosphatase specific activity. In vivo, cell-growth factor-scaffold constructs were subcutaneously implanted in MF1nu/nu mice for six weeks. Bone formation was confirmed by histology. Results: Saos-2 extracted osteoinductive factors significantly stimulated alkaline phosphatase CFU-F formation and alkaline phosphatase specific activity by 30-40% in both basal and osteogenic conditions. The concentrations of 'retentate' required to induce differentiation and mineralisation were approximately 100-fold lower than concentrations of rhBMP's required to achieve similar effects. Osteoinductive factors present in Saos-2 cell extracts promoted adhesion, expansion and differentiation of human osteoprogenitor cells on 3-D scaffolds leading to bone matrix formation in vitro. Examination of in vivo activity of 'retentate' on PLGA porous scaffold in subcutaneously implant model indicated formation of bone as evidenced by alcian blue/Sirius red staining as well as Type I collagen immunohistochemistry. Conclusion: The generation of 3-D biomimetic structures of PLGA incorporating osteoinductive factors from Saos-2 cells indicates their potential for de novo bone formation that exploits osteoinductive factors and cell-matrix interactions.