Modulation effect of rhBMP2 on osteogenic activity in human periodontal ligament fibroblasts and osteoblast cell lines

Abstract

Background: Bone morphogenic proteins (BMPs) are known to promote osteogenesis, and clinical trials are currently under way to evaluate the potential of BMPs to promote fracture-healing and bone regeneration. The aim of this study was to investigate the biological effects of rhBMP2 on primary cultured cells isolated from periodontal ligament, gingival tissue, cancellous bone of human specimens and bone marrow aspirate before clinical trial of rhBMP2 in periodontal tissue engineering. Methods: We analyzed effect of different concentration of rhBMP2 on osteogenic activity in periodontal ligament fibroblasts (PDLF), gingival fibroblasts (GF), bone-derived cells (HBDC) and bone marrow-derived cells (HBMDC). Cytochemical staining and assay of alkaline phosphatase activity were performed in different cell types. Effect of rhBMP2 stimulation on in vitro mineralization was studied by Alizarin red S stain. Differential modulation of core biding factor 1 (Cbfa1), osteoprotegerin (OPG), and receptor of NF-κB (RANKL) mRNA expression by rhBMP2 were examined by semiquantitative RT-PCR. Results: rhBMP2 was found to stimulate an increase of alkaline phosphatase activity and cytochemical staining which was dose dependent (0-300ng/ml) in PDLF and HBDC. The study mentioned above was not in GF cells. HBMDC showed dramatically increased cytochemical staining at relative low dose of rhBMP2 (50ng/ml). rhBMP2 (300ng/ml) significantly enhanced in vitro mineralization in HBDC and HBMDC, but only had mild effect on PDLF cells in confluent 20-days culture. PDLF cells and HBDC showed the expression of Cbfa1, OPG, but not RANKL at mRNA level, and Cbfa1 mRNA level was up-regulated upon rhBMP2 stimulation. Conclusions: The data of this study support the concept that PDLF cells have phenotypes of osteoblast linage. The fact that rhBMP2 can both increase ALPase activity and promote in vitro mineralization in PDLF cells and HBDC raises the possibility that rhBMP2 may be useful in directing the regeneration of periodontal mineralized tissue during wound healing. Further study using signal such as rhBMP2 in tissue scaffold to modulate PDLF cells differentiation may be needed in tissue engineering of periodontal tissue.