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This article is part of the supplement: 25th European Workshop for Rheumatology Research

Poster presentation

Amelioration of joint inflammation by a PAR-2-specific monoclonal antibody

V King1, EB Kelso1, JC Lockhart2, L Dunning2, WR Ferrell1, JA Gracie1 and IB McInnes1

Author Affiliations

1 Centre for Rheumatic Diseases, University of Glasgow, UK

2 Biological Sciences, University of Paisley, UK

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Arthritis Research & Therapy 2005, 7(Suppl 1):P160  doi:10.1186/ar1681


The electronic version of this article is the complete one and can be found online at:


Received:11 January 2005
Published:17 February 2005

© 2005 BioMed Central Ltd

Background

Protease-activated receptor-2 (PAR-2) is a G-protein-coupled receptor recognized to mediate inflammatory responses. Using a PAR-2 'knockout' mouse, we previously demonstrated this receptor to play a crucial role in chronic joint inflammation [1]. Inhibition of PAR-2 activation therefore potentially represents a novel therapeutic target in treatment of arthritis, but selective antagonists are not yet available.

Objective

To test the hypothesis that acute joint inflammation could be inhibited by targeting the 'tethered ligand' sequence of PAR-2 using a specific monoclonal antibody (SAM-11).

Methods

The presence of synovial PAR-2 was confirmed in wild-type (PAR-2+/+) C57BL/6J mice by western blotting using SAM-11 (Santa Cruz, USA). The anti-inflammatory potential of this antibody, which inhibits PAR-2 activation by preventing release of its activating ligand, was investigated by intra-articular administration of SAM-11 to mice prior to induction of acute joint inflammation (under halothane/O2/N2O anaesthesia) by injection of 20 μl 2% carrageenan and 4% kaolin (C/K) into the knee joint. Joint swelling was assessed by comparing caliper-measured knee joint diameter pre and 24 hours post-injection. The swelling response was compared in untreated, and in two parallel groups of inflamed mice that had received intra-articular injection of SAM-11 at 5 ng or 10 ng, 5 min prior to C/K administration. Selectivity of SAM-11 for PAR-2 was confirmed by immunohistochemical analysis (in combination with the Animal Research Kit Dako, USA) of the wild-type mouse brain, which expresses PAR-1, PAR-2, PAR-3 and PAR-4 compared with that of PAR-2 gene-disrupted (PAR-2-/-) mice.

Results

Western blotting demonstrated the presence of PAR-2 in normal murine synovium and substantial upregulation in acutely inflamed joints. Immunohistochemical analysis of the murine brain with SAM-11 revealed staining in PAR-2+/+ mice but not in PAR-2-/- mice, confirming PAR-2 specificity of this monoclonal antibody. C/K resulted in substantial knee joint swelling, which was significantly and dose-dependently (P < 0.00001; one-way analysis of variance; n = 4 per group) inhibited by SAM-11 pretreatment (Fig. 1). These findings are consistent with a proinflammatory role for PAR-2 in arthritis, and demonstrate that antibody inhibition of PAR-2 activation ameliorates acute joint inflammation.

thumbnailFigure 1. Twenty-four hours after intra-articular injection of kaolin, knee joint swelling is dose-dependently reduced by SAM-11 pre-treatment. ** P < 0.00001, * P < 0.01 (n = 4 per group). NS, no significantly.

References

  1. Ferrell WR, Lockhart JC, Kelso EB, Dunning L, Plevin R, et al.: Essential role for proteinase-activated receptor-2 in arthritis.

    J Clin Invest 2003, 111:35-49. PubMed Abstract | Publisher Full Text | PubMed Central Full Text OpenURL