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Presenting Author(s) Phyllis LuValle
Abstract Title Exogenous Fibroblast Growth Factor and Diminished Raf Kinase Signalling Result in Impaired Endochondral Bone Formation in Embryonic Limb Rudiments.
Full author List Todd Leask, Christal Stieb, and Phyllis LuValle
Text of abstract The intracellular signalling involved in the initiation and coordination of longitudinal bone growth is not well understood. Raf-1 kinase is expressed only by post-mitotic growth plate chondrocytes and is therefore an excellent candidate for coordinating intracellular signalling events. We and others have previously demonstrated in vitro that Raf-1 and the MAP kinase pathway are involved in transcriptional activation of hypertrophic chondrocyte-specific genes. Thyroid hormone-stimulated Fibroblast growth factor (FGF) signalling is also involved in bone formation. Ligands for FGFR3 found in cartilage include FGF9 and FGF18. We have previously shown in vitro that FGF9 and FGF18 are expressed by chondrocytes undergoing maturation and can themselves promote chondrocyte maturation and activation of Raf-1 and the MAP kinase pathway. Using an ex vivo system of normal embryonic mouse bone rudiment cultures, we have investigated the roles that Raf-1 and FGFR3 signalling play in the maturation of chondrocytes in the growth plate. Treatment with a Raf-1 Kinase Inhibitor (Calbiochem) resulted in a marked dose-dependent decrease in both the length of the rudiment and chondrocyte hypertrophy. Mineralization was inhibited as evidenced by both alkaline phosphatase activity and alizarin red stain. These results indicate that Raf-1 is required for accurate maturation and mineralization of growth plate cartilage. Surprisingly, addition of FGF9 or FGF18 resulted in dose-dependent reductions in rudiment length, diminished size of the hypertrophic zone, and inhibition of mineralization. These results suggest that exogenous FGFR3 signalling acts at different thresholds on the processes of maturation.. An activating mutation in FGF receptor 3 (FGFR3) results in Achondroplasia, a common form of dwarfism that exhibits a decrease in growth plate chondrocyte proliferation in mice. We are currently repeating our experiments in mice carrying this mutation. Taken together, these results imply that both Raf-1 and FGFR3 signalling are necessary for proper bone formation.
Which session is your work most relevant to: Tissue differentiation