title: Synergistic effect of fibroblast growth factor-4 in ectopic bone formation induced by bone morphogenetic protein-2.
abstract: Bone morphogenetic protein family members (BMPs) are essential signaling molecules during limb development and, in this process, fibroblast growth factor family members (FGFs) cooperate with BMPs. FGFs also exert anabolic effects in bone when systemically or locally applied. Thus, it is likely that the cooperation with FGFs also occurs in BMP-induced ectopic bone formation and that the exogenous FGF application would promote this bone formation. In the present study, after subcutaneously implanting recombinant human BMP-2 (rhBMP-2) in rats, we examined the expression of FGF-4 and FGF receptors (FGFRs) mRNAs and the effect of exogenous recombinant human FGF-4 (rhFGF-4) on bone formation. Three days after implantation, the pellets containing rhBMP-2 were surrounded by fibroblastic mesenchymal cells; on day 7, cartilage tissue appeared; on day 10, hypertrophic chondrocytes and a small amount of mineralized tissue were observed; and, on day 14, the amount of mineralized tissue increased. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that FGF-4 expression appeared at early stages (days 3 and 7) and its expression decreased at later stages (days 10, 14, and 21), whereas FGFRs were expressed continuously. In situ hybridization revealed that, on days 3 and 7, FGF-4, and FGFR subtypes 1 and 2 (FGFR-1 and FGFR-2) were expressed in mesenchymal cells and chondrocytes, and in the area of alkaline phosphatase (ALP) expression. On day 10, FGF-4 was not detected, whereas the expression of FGFR-1 and FGFR-2 was detectable in the area of alkaline phosphatase (ALP) expression. Injection of rhFGF-4 on days 2, 3, and 4 enhanced the mineralized tissue formation induced by rhBMP-2; however, neither rhFGF-4 treatment on days 6, 7, and 8 nor rhFGF-4 treatment on days 9, 10, and 11 influenced the amount of rhBMP-2-induced mineralization. Our results indicate that FGF-4 and FGFR signals play important roles during rhBMP-2-induced bone formation. We further suggest that the combination of rhBMP-2 and rhFGF-4 would be useful for bone augmentation.
authors.affiliation: Department of Periodontology, Hard Tissue Engineering, Tokyo Dental and Medical University, Japan. kaorikubo513@aol.com
authors.forename: K
authors.initials: K
authors.lastname: Kubota
authors.forename: S
authors.initials: S
authors.lastname: Iseki
authors.forename: S
authors.initials: S
authors.lastname: Kuroda
authors.forename: S
authors.initials: S
authors.lastname: Oida
authors.forename: T
authors.initials: T
authors.lastname: Iimura
authors.forename: W R
authors.initials: WR
authors.lastname: Duarte
authors.forename: K
authors.initials: K
authors.lastname: Ohya
authors.forename: I
authors.initials: I
authors.lastname: Ishikawa
authors.forename: S
authors.initials: S
authors.lastname: Kasugai
mesh_terms: D000818:Animals; D001483:Base Sequence; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins / Q000502:physiology*; D017931:DNA Primers; D051519:Fibroblast Growth Factor 4; D005346:Fibroblast Growth Factors / Q000502:physiology*; D010012:Osteogenesis / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000502:physiology*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016212:Transforming Growth Factor beta
chemical_list: C525718:BMP2 protein, human; C525720:Bmp2 protein, rat; D055396:Bone Morphogenetic Protein 2; D019485:Bone Morphogenetic Proteins; D017931:DNA Primers; C496447:Fgf4 protein, rat; D051519:Fibroblast Growth Factor 4; D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D005346:Fibroblast Growth Factors