A mutation in a CD44 variant of inflammatory cells enhances the mitogenic interaction of FGF with its receptor
Shlomo Nedvetzki, … , Avner Yayon, David Naor
Shlomo Nedvetzki, … , Avner Yayon, David Naor
Published April 15, 2003
Citation Information: J Clin Invest. 2003;111(8):1211-1220. https://doi.org/10.1172/JCI17100.
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Article Autoimmunity

A mutation in a CD44 variant of inflammatory cells enhances the mitogenic interaction of FGF with its receptor

Abstract

Synovial fluid cells from joints of rheumatoid arthritis (RA) patients express a novel variant of CD44 (designated CD44vRA), encoding an extra trinucleotide (CAG) transcribed from intronic sequences flanking a variant exon. The CD44vRA mutant was detected in 23 out of 30 RA patients. CD44-negative Namalwa cells transfected with CD44vRA cDNA or with CD44v3-v10 (CD44vRA wild type) cDNA bound FGF-2 to an equal extent via their associated heparan sulfate chains. However, Namalwa cells, immobilizing FGF-2 via their cell surface CD44vRA, bound substantially more soluble FGF receptor-1 (FGFR-1) than did Namalwa cells immobilizing the same amount of FGF-2 via their cell surface CD44v3-v10. The former cells stimulated the proliferation of BaF-32 cells, bearing FGFR-1, more efficiently than did the latter cells. Finally, isolated primary synovial fluid cells from RA patients expressing CD44vRA bound more soluble FGFR-1 to their cell surface–associated FGF-2 than did corresponding synovial cells expressing CD44v3-v10 or synovial cells from osteoarthritis patients. The binding of soluble FGFR-1 to RA synovial cells could be specifically reduced by their preincubation with Ab’s against the v3 exon product of CD44. Hence, FGF-2 attached to the heparan sulfate moiety expressed by the novel CD44 variant of RA synovium cells exhibits an augmented ability to stimulate FGFR-1–mediated activities. A similar mechanism may foster the destructive inflammatory cascade not only in RA, but also in other autoimmune diseases.

Authors

Shlomo Nedvetzki, Itshak Golan, Nathalie Assayag, Erez Gonen, Dan Caspi, Micha Gladnikoff, Avner Yayon, David Naor

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Figure 4

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Enhanced binding of FGFR-1 to cell surface CD44vRA. (a) Binding of FGFR-...
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.