title: Anti-citrullinated fibronectin antibodies in rheumatoid arthritis are associated with human leukocyte antigen-DRB1 shared epitope alleles.
abstract: BACKGROUND
  Fibronectin is one of the most abundant proteins present in the inflamed joint. Here, we characterized the citrullination of fibronectin in the joints of rheumatoid arthritis (RA) patients and studied the prevalence, epitope specificity and human leukocyte antigen (HLA) association of autoantibodies against citrullinated fibronectin in RA.
  
  
  METHODS
  Citrullinated residues in fibronectin isolated from RA patient synovial fluid were identified by mass spectrometry. The corresponding citrullinated and non-citrullinated peptides were synthesized and used to analyze the presence of autoantibodies to these peptides in RA sera and sera from other diseases and healthy controls by ELISA. The data were compared with risk factors like shared epitope HLA alleles and smoking, and with clinical features.
  
  
  RESULTS
  Five citrullinated residues were identified in fibronectin from RA synovial fluid. RA sera reacted in a citrulline-dependent manner with two out of four citrullinated fibronectin peptides, one of which contains two adjacent citrulline residues, in contrast to non-RA sera, which were not reactive. The most frequently recognized peptide (FN-Cit1035,1036, LTVGLTXXGQPRQY, in which × represents citrulline) was primarily targeted by anti-CCP (cyclic citrullinated peptide) 2-positive RA patients. Anti-FN-Cit1035,1036 autoantibodies were detected in 50% of established anti-CCP2-positive RA patients and in 45% of such patients from a early arthritis clinic. These antibodies appeared to be predominantly of the immunoglobulin G (IgG) isotype and to be associated with HLA shared epitope alleles (odds ratio = 2.11).
  
  
  CONCLUSIONS
  Fibronectin in the inflamed synovia of RA patients can be citrullinated at least at five positions. Together with the flanking amino acids, three of these citrullinated residues comprise two epitopes recognized by RA autoantibodies. Anti-citrullinated fibronectin peptide antibodies are associated with HLA shared epitope alleles.
authors.affiliation: Department of Biomolecular Chemistry, Institute for Molecules and Materials, Nijmegen Center for Molecular Life Sciences, Radboud University Nijmegen, P,O, Box 9101, 6500 HB Nijmegen, The Netherlands. J.vanbeers@ncmls.ru.nl
authors.forename: Joyce J B C
authors.initials: JJ
authors.lastname: van Beers
authors.forename: Annemiek
authors.initials: A
authors.lastname: Willemze
authors.forename: Judith
authors.initials: J
authors.lastname: Stammen-Vogelzangs
authors.forename: Jan W
authors.initials: JW
authors.lastname: Drijfhout
authors.forename: René E M
authors.initials: RE
authors.lastname: Toes
authors.forename: Ger J M
authors.initials: GJ
authors.lastname: Pruijn
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D000704:Analysis of Variance; D001172:Arthritis, Rheumatoid / Q000097:blood / Q000235:genetics / Q000276:immunology*; D001323:Autoantibodies / Q000097:blood / Q000276:immunology*; D001665:Binding Sites / Q000235:genetics; D002956:Citrulline / Q000276:immunology* / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000235:genetics / Q000276:immunology*; D005353:Fibronectins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D059811:HLA-DRB1 Chains / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood / Q000276:immunology; D016015:Logistic Models; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012907:Smoking / Q000276:immunology; D013582:Synovial Fluid / Q000276:immunology / Q000378:metabolism; D053719:Tandem Mass Spectrometry
chemical_list: D001323:Autoantibodies; D000939:Epitopes; D005353:Fibronectins; D059811:HLA-DRB1 Chains; D007074:Immunoglobulin G; D010455:Peptides; D002956:Citrulline