FIBROBLAST-GROWTH-FACTOR-2 AND FIBROBLAST-GROWTH-FACTOR-4 STIMULATE MIGRATION OF MOUSE EMBRYONIC LIMB MYOGENIC CELLS

Fibroblast growth factors (FGFs) are believed to be vital for limb out growth and patterning during embryonic development. Although the effec t of FGFs on the formation of the skeletal elements has been studied i n detail, their effect on the development of the limb musculature is s till uncertain. In this study, we used Blindwell chemotactic chambers to examine the effect of FGF-2 and FGF-4 on the motility of myogenic c ells obtained from the proximal region of day 11.5 mouse forelimbs, Th e limb myogenic cells were found to be chemotactically attracted to FG F-2 and FGF-4 at 1-50 ng/ml, Both FGFs increased myogenic cell migrati on in a dose-dependent manner with maximal responses attained at 10-50 ng/ml for FGF-2 and at 10 ng/ml. for FGF-4; however, FGF-2 was found to be a more potent chemoattractant than FGF-4, It was possible to inh ibit the myogenic cells' response to FGF-2 and FGF-4 by the addition o f the appropriate neutralizing antibody The effects of FGF-2 on cell m igration were further investigated by loading this cytokine into Affi- Gel blue beads and transplanting them into day 11.5 forelimb buds, The results showed that FGF-2 attracted DiI-labelled proximal cells to mi grate toward the implanted beads and that the migration was more exten sive than that observed in the absence of FGF-2, A checkerboard assay was performed in which various concentrations of FGF-2 and FGF-4 were introduced to both the upper and lower wells of the Blindwell chambers , The results indicated that both FGF isoforms can stimulate chemokine sis as well as chemotaxis in myogenic cells, In addition, the effect o f FGF-2 and FGF-4 on other aspects of skeletal muscle development was investigated, FGF-S at 0.1-10 ng/ml stimulated. a significant increase in the number of myocytes expressing sarcomeric myosin on examination after 48 hr in culture, but the effect of FGF-4 was negligible at all concentrations analyzed; however, both FGF-2 and FGF-4 inhibited myoc yte fusion compared with the spontaneous fusion observed ill control c ultures, Finally we used in situ hybridization and immunohistochemical techniques to determine the distribution of myogenic cells and FGF-2 protein in the day 11.5 mouse forelimbs. (C) 1997 Wiley-Liss, Inc.