Synergistic Effects of rhBMP-2/rhbFGF on Bone Marrow Stromal Cells

Objective: Bone morphogenetic proteins-2 (BMP-2) or basic fibroblast growth factor (bFGF) have been found to be the most promising osteoinductive substances and is expected to be utilized clinically for oral implants. However, it is not clear that the optimal ratios of BMP-2 / bFGF to improve bone formation. In this study, we examined the effect of rhBMP-2/rhbFGF on proliferation and differentiation of bone marrow stromal cells (BMSCs) in beagle dog. Methods: The experiments were completed at Lab of Bioengineering, Ninth People's Hospital Shanghai Jiaotong University Medical College. Under general anesthesia, 2ml bone marrow was obtained from ilium marrow of young beagle dogs. The BMSCs were cultured in delbecco's modified eagle medium (DMEM). When the cells were confluent, they were liberated by trypsin/EDTA. The 3rd generated BMSCs were seeded treated with rhBMP-2 (0, 25ng/ml, 50ng/ml, 100ng/ml, 200ng/ml),rhbFGF (0, 25ng/ml, 50ng/ml, 100ng/ml, 200ng/ml) and rhBMP-2/rhbFGF (1:1, 2:1, 4:1, 8:1) respectively. Control group was BMSCs treated with osteogenic medium only. The proliferation and differentiation of BMSCs were observed through MTT and ALP for 6d, ALP and Von Kossa staining on the 6th day. The data among the groups were evaluated by ANOVA and p-values less than 0.05 were considered significant. Results: (1)Measurement of proliferation: The concentration of 100ng/ml and 200ng/ml rhBMP-2 (P<0.01), 50ng/ml rhbFGF (P<0.05), rhBMP-2/rhbFGF (2:1) could significantly promote BMSCs proliferation. (2)Measurement of differentiation: 25ng/ml rhBMP-2 (P<0.01), rhBMP-2/bFGF (1:1, 2:1) promote BMSCs differentiation, and rhbFGF has no effect on differentiation. (3)ALP and Von Kossa staining: The ALP staining proofed the brown positive express; Von Kossa staining proofed the positive express of calcific node. Conclusion: rhBMP-2/rhbFGF (2:1) exerts synergistic effect to promote BMSCs proliferation and differentiation. The combination of rhBMP-2 and rhbFGF would be useful for oral implant.