Regulation of fibroblast growth factor 2 and fibroblast growth factor receptors by transforming growth factor β in human osteoblastic MG-63 cells

Abstract

Fibroblast growth factor 2 (FGF-2) and its receptors (FGFRs) are important regulators of bone cell function. Although FGF-2 is a major modulator of bone cell function, its expression and regulation in human osteoblasts have not been investigated. We examined FGF-2 messenger RNA (mRNA) expression and regulation in the human osteosarcoma MG-63 cells. Northern analysis revealed that MG-63 cells expressed FGF-2 mRNA transcripts of 7, 4, 2.2, and 1.3 kilobases (kb). In the absence of serum, treatment with transforming growth factor beta (TGF-β; 0.1-10 ng/ml) increased all FGF-2 mRNA transcripts. Maximal increase was seen with 1 ng/ml of TGF-β. TGF-β increased FGF-2 mRNA expression within 2 h and this was sustained for 24 h. Phorbal myristate acetate (PMA; 1 μM) also increased FGF-2 mRNA at 6 h. Time course studies showed that TGF-β did not significantly alter FGFR1 or FGFR2 mRNA expression in MG-63 cells. Western blotting with anti-human FGF-2 revealed that MG-63 cells synthesize three isoforms of FGF-2 protein of ∼18, 22/23, and 24 kDa, which were increased after either 6 h or 24 h of treatment with TGF-β. Increased FGF-2 mRNA and protein expression in response to TGF-β was markedly reduced by the protein kinase A (PKA) inhibitor H-89. Immunogold labeling of MG-63 cells treated with TGF-β showed increased labeling for FGF-2 and FGFR2 in the nuclei. In contrast, TGF-β treatment significantly decreased FGFR1 labeling in the nuclei. These data show that TGF-β regulates FGF-2 gene expression in human osteosarcoma cells. Furthermore, TGF-β modulates the cellular localization of FGF-2 and its receptors.