Targeted Synovial Fluid Proteomics for Biomarker Discovery in Rheumatoid Arthritis |
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Authors: | Leticia Cano Daniel G Arkfeld |
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Institution: | 1. Division of Immunology, Beckman Research Institute of the City of Hope National Medical Center, Duarte, CA, 91010, USA 3. Laboratory of Applied Mass Spectrometry, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, MD, USA 2. Division of Rheumatology, Department of Medicine, University of Southern California Keck School of Medicine and the Los Angeles County and University of Southern California Medical Center, Los Angeles, CA, 90033, USA
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Abstract: | Objective Rheumatoid arthritis (RA) is an autoimmune disease that targets the synovium. The autoantigens involved in the autoantibody
responses in RA are unknown. A targeted proteomics approach was used to identify proteins in RA synovial fluid (SF) that are
recognized by autoantibodies in RA sera.
Methods RA SF, depleted of abundant proteins, was fractionated by two-dimensional liquid chromatography (chromatofocusing followed
by reverse phase HPLC). Protein arrays constructed from these fractions were probed with RA and normal control sera, and proteins
within reactive fractions were identified by mass spectrometry. The reactivity of RA sera to an identified peptide was confirmed
by ELISA.
Results RA sera specifically reacted to a SF fraction containing fibrin. Mass spectrometry analyses established the presence of a
citrullinated arginine at position 271 of the fibrin fragment present in RA SF. A synthetic peptide corresponding to fibrin
residues 259–287, containing the citrulline substitution at Arg 271, was recognized by 10 of 12 RA sera, but by two of 18
normal control sera and one of 10 systemic lupus erythematosus sera.
Conclusion Proteomics methodology can be used to directly characterize post-translational modifications in candidate autoantigens isolated
from sites of disease activity. The finding that RA sera contain antibodies to the citrullinated fibrin 259–287 peptide may
ultimately lead to improved diagnostic tests for RA and/or biomarkers for disease activity.
Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users. |
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Keywords: | Rheumatoid arthritis Synovial fluid Clinical proteomics Biomarker discovery Citrullination Fibrinogen ELISA protein macroarrays Autoimmune disease Autoantibodies Autoantigens Mass spectrometry Post-translational modification |
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