Rheumatoid Arthritis: Charlionet R

Goëb V, Thomas-L'Otellier M, Daveau R, Charlionet R, Fardellone P, Le Loët X, Tron F, Gilbert D, Vittecoq O. · Department of Rheumatology and Inserm Unit 905, IFRMP 23, Institute for Biomedical Research, University of Rouen, Rouen University Hospital, Rouen 76031 cedex, France. · Arthritis Res Ther. · Pubmed #19284558 links to  free full text

Abstract: INTRODUCTION: The aim of our study was to identify new early rheumatoid arthritis (RA) autoantibodies. METHODS: Sera obtained from 110 early untreated RA patients (<6 months) were analyzed by western blot using HL-60 cell extract, separated on one-dimensional and two-dimensional gel electrophoresis (1-DE, 2-DE). Sera from 50 healthy blood donors and 20 patients with non-RA rheumatisms were used as controls for 1-DE and 2-DE, respectively. The immunoreactive proteins were identified by MALDI-TOF mass spectrometric analysis and the presence of potential sites of citrullination in each of these proteins was evaluated. FT-ICR mass spectrometry was used to verify experimentally the effect of citrullination upon the mass profile observed by MALDI-TOF analysis. RESULTS: The 110 1-DE patterns allowed detection of 10 recurrent immunoreactive bands of 33, 39, 43, 46, 51, 54, 58, 62, 67 and 70 kDa, which were further characterized by 2-DE and proteomic analysis. Six proteins were already described RA antigens: heterogeneous nuclear ribonucleoprotein A2/B1, aldolase, alpha-enolase, calreticulin, 60 kDa heat shock protein (HSP60) and BiP. Phosphoglycerate kinase 1 (PGK1), stress-induced phosphoprotein 1 and the far upstream element-binding proteins (FUSE-BP) 1 and 2 were identified as new antigens. Post-translational protein modifications were analyzed and potentially deiminated peptides were found on aldolase, alpha-enolase, PGK1, calreticulin, HSP60 and the FUSE-BPs. We compared the reactivity of RA sera with citrullinated and noncitrullinated alpha-enolase and FUSE-BP linear peptides, and showed that antigenicity of the FUSE-BP peptide was highly dependent on citrullination. Interestingly, the anti-cyclic citrullinated peptide antibody (anti-CCP2) status in RA serum at inclusion was not correlated to the reactivity directed against FUSE-BP citrullinated peptide. CONCLUSIONS: Two categories of antigens, enzymes of the glycolytic family and molecular chaperones are also targeted by the early untreated RA autoantibody response. For some of them, and notably the FUSE-BPs, citrullination is involved in the immunological tolerance breakdown observed earlier in RA patients. Autoantibodies recognizing a citrullinated peptide from FUSE-BP may enhance the sensibility for RA of the currently available anti-CCP2 test.

Saulot V, Vittecoq O, Charlionet R, Fardellone P, Lange C, Marvin L, Machour N, Le Loët X, Gilbert D, Tron F. · Institut National de la Santé et de la Recherche Médicale, Rouen, France. · Arthritis Rheum. · Pubmed #12115223 links to  free full text

Abstract: OBJECTIVE: To identify a new autoantigen/autoantibody population in rheumatoid arthritis (RA) sera. METHODS: Following a population-based recruitment effort, 255 patients with very early arthritis (median disease duration 4 months) were studied using different clinical, biologic, and radiologic assessments. After a followup period of 1 year, patients were classified as having RA (n = 145), non-RA rheumatic diseases (n = 70), and undifferentiated arthritis (n = 40). Patients' sera were analyzed by one-dimensional (1D) and 2D Western blotting. The recognized 50-kd protein was analyzed by matrix-assisted laser desorption ionization-time-of-flight (MALDI-TOF) mass spectrometry (MS). RA serum reactivities were evaluated against the recombinant protein synthesized by an in vitro coupled transcription-translation system. RESULTS: On 1D Western blots, 36 of the 145 RA sera bound to a 50-kd polypeptide. On 2D Western blots, anti-50-kd+ RA sera recognized a triplet of isoelectric point 6.5-7.0 and a molecular mass of 50 kd. The 3 spots of the triplet were analyzed by MALDI-TOF MS and were shown to correspond to human alpha-enolase. A goat anti-enolase antiserum, which recognized a band comigrating with the 50-kd antigen on 1D Western blots, gave a labeling pattern on 2D Western blots similar to that observed with anti-50-kd+ RA sera. Among the 36 RA sera that identified alpha-enolase in protein maps, only 8 recognized the recombinant (unmodified) alpha-enolase. The specificity of anti-alpha -enolase antibodies for RA was 97.1%. Half of the anti-alpha -enolase-positive RA patients were negative for both rheumatoid factor and antifilaggrin antibodies. The presence of anti-alpha-enolase antibodies was the greatest predictive factor of radiologic progression in the first 66 RA patients included. CONCLUSION: Autoantibodies to alpha-enolase, an enzyme of the glycolytic pathway, are present in the sera of patients with very early RA and have potential diagnostic and prognostic value for RA.