A mutation in a CD44 variant of inflammatory cells enhances the mitogenic interaction of FGF with its receptor
J. Clin. Invest. Shlomo Nedvetzki, et al. 111:1211 doi:10.1172/JCI17100 [Go to this article.]

Figure 4
Enhanced binding of FGFR-1 to cell surface CD44vRA. (a) Binding of FGFR-1 to Namalwa transfectants. The ability of the indicated cells to bind soluble FGFR-1 or FGF-2 was analyzed as described in Methods. (b) Kinetics of FGFR-1 binding to FGF-2–associated Namalwa transfectants. Namalwa-CD44v3-v10 and Namalwa-CD44vRA were incubated in the presence of the indicated concentrations of FGF-2 with soluble FGFR-1 conjugated to AP. Binding of the soluble receptor to Namalwa transfectants was assessed as indicated in a and demonstrates 50% effective binding at 100 pM of FGF-2. *P < 0.05. (c) FGF-2 bound to Namalwa-CD44vRA induces enhanced proliferation of BaF-32 cells expressing FGFR-1. The indicated fixed Namalwa transfectants were incubated in the presence of FGF-2 with BaF-32 cells. The ability of the bound FGF-2 to induce proliferation in BaF-32 cells was analyzed by a colorimetric assay at OD 490. Positive control: BaF-32 cells incubated with FGF-2 and heparin. Negative controls: BaF-32 cells incubated with FGF-2 alone or with heparin alone. Inset: A similar experiment, except that the proliferation of the positive-control BaF-32 cells is identical to the proliferation of BaF-32 cells incubated with Namalwa-CD44vRA cells. The numbers beneath the bars correspond to the numbered treatments shown under the bar of the main figure (Figure 4c). (d) Analysis of BaF-32 cell proliferation induced by fixed Namalwa transfectants or RA synovial fluid cells. Inhibition of BaF-32 cell proliferation by anti-CD44v3 mAb. The intensity of BaF-32 cell proliferation, as indicated by a colorimetric assay, following incubation with: bar 1, Namalwa-Neo cells; bar 2, Namalwa-CD44s cells; bar 3, Namalwa-CD44v3-v10 cells; bar 4, Namalwa-CD44v3-v10 cells plus anti-CD44v3 mAb; bar 5, Namalwa-CD44v3-v10 cells plus isotype-matched control mAb: bar 6, Namalwa-CD44vRA cells; bar 7, Namalwa-CD44vRA cells plus anti-CD44v3 mAb; bar 8, Namalwa-CD44vRA cells plus isotype-matched control mAb; bar 9, FGF-2 plus heparin (positive control); bar 10, FGF-2 alone (negative control); bar 11, heparin alone (negative control); bar 12, medium alone (negative control); bars 13–15, proliferation of BaF-32 cells following incubation with fixed RA synovial fluid cells derived from three different patients (RA16, RA17, RA18). Note that during the proliferation assay, Namalwa transfectants were loaded with FGF-2, whereas synovial fluid cells were not, because they contain endogenous FGF-2 (see inset, Figure 5b). Statistical analysis of the principal groups is shown.