Abstract
IntroductionThe aim of our study was to identify new early rheumatoid arthritis (RA) autoantibodies.MethodsSera 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.ResultsThe 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, α-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, α-enolase, PGK1, calreticulin, HSP60 and the FUSE-BPs. We compared the reactivity of RA sera with citrullinated and noncitrullinated α-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.ConclusionsTwo 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.
Highlights
The aim of our study was to identify new early rheumatoid arthritis (RA) autoantibodies
Post-translational protein modifications were analyzed and potentially deiminated peptides were found on aldolase, enolase, Phosphoglycerate kinase 1 (PGK1), calreticulin, HSP60 and the far upstream elementbinding proteins (FUSE-BP)
We compared the reactivity of RA sera with citrullinated and noncitrullinated -enolase and FUSE-BP linear peptides, and showed that antigenicity of the FUSE-BP peptide was highly dependent on citrullination
Summary
The aim of our study was to identify new early rheumatoid arthritis (RA) autoantibodies. The autoimmune response appears early, often prior to the apparition of clinical symptoms, and leads to the production of various autoantibodies (autoAb) detectable in serum. These autoAb help to understand pathological mechanisms and constitute biological markers of the disease [1]. Whether filaggrin is the true autoantigen of ACPA is unlikely since it is exclusively expressed in epithelial cells, and other citrullinated proteins – such as fibrinogen [8], vimentin [9], enolase [10], collagen type I [11], fibronectin [12], a translational initiation factor [13] and even a viral protein, EBNA-1 [14] – have been shown to be the target of the autoimmune response. The deimination of proteins is mediated by peptidylarginine deiminase (PADI) and occurs notably during cell death and oxidative stress [15,16], both events observed in RA synovium
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
