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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Namalwa-CD44v3-v10 and Namalwa-CD44vRA bind FGF-2 to a similar extent. (...
Namalwa-CD44v3-v10 and Namalwa-CD44vRA bind FGF-2 to a similar extent. (a) Flow cytometry. The indicated Namalwa transfectants were incubated with biotinylated FGF-2 in the absence (inset) or presence of 0.2 M NaCl and then analyzed by flow cytometry for their ability to bind this growth factor, detected by staining with streptavidin-PE. Control: Namalwa cells transfected with empty vector (Namalwa-neo) and incubated with biotinylated FGF-2. (b) Western blot analysis. Western blots of cell extracts from Namalwa transfectants with anti-FGF Ab confirmed the flow-cytometry analysis. The Namalwa transfectants were preincubated with FGF-2 before being subjected to cell extraction and gel electrophoresis. The anti-FGF Ab showed that FGF-2 was bound to a similar extent to CD44v3-v10 and CD44vRA, whereas CD44s did not bind FGF-2. Actin, a housekeeping gene product, is equally expressed in all transfectant extracts. (c) Excess soluble heparin blocks the binding of FGF-2 to Namalwa-CD44vRA. Namalwa-CD44vRA cells were coincubated with biotinylated FGF-2 and an excess of soluble heparin or soluble chondroitin sulfate A plus C, then analyzed by flow cytometry for their ability to bind the growth factor. The binding of biotinylated FGF-2 was detected with streptavidin-PE. The extreme left-hand histogram depicts Namalwa-CD44vRA cells incubated with streptavidin-PE only. Similar results were obtained using three Namalwa-CD44vRA clones (not shown). (d) Heparinase treatment reduces FGF-2 binding to Namalwa-CD44vRA. Namalwa-CD44vRA cells were treated with heparinase or chondroitinase ABC and then analyzed by flow cytometry for their ability to bind biotinylated FGF-2. Detection and control as in c. Similar results were observed in three Namalwa-CD44vRA clones (not shown).