Antibodies specific for carbamylated proteins precede the onset of clinical symptoms in mice with collagen induced arthritis

PLoS One. 2014 Jul 15;9(7):e102163. doi: 10.1371/journal.pone.0102163. eCollection 2014.

Abstract

Objective: The immune response to post-translationally modified antigens is a key characteristic of rheumatoid arthritis. Carbamylation is such a posttranslational modification. Recently, we demonstrated that autoantibodies recognizing carbamylated proteins are present in sera of rheumatoid arthritis. The molecular mechanisms underlying the break of tolerance and hence the induction of anti-CarP antibody responses are unknown as well as their appearance in mouse models for systemic arthritis. Therefore we analyzed their appearance in the mouse collagen-induced arthritis model.

Methods: collagen induced arthritis was induced by immunization with type II collagen in complete Freund's adjuvant. Arthritis severity was monitored by clinical scoring and anti-CarP antibody levels were determined by ELISA.

Results: Anti-CarP antibodies were detectable in mice with collagen induced arthritis. We did not detect ACPA in mice with collagen induced arthritis. The specificity of the antibodies for carbamylated proteins was confirmed by inhibition assays and immunoblotting. Injection with complete Freund's adjuvant without type II collagen could also induce anti-CarP antibodies, however, in mice with arthritis, the anti-CarP antibody response was stronger and developed more rapidly. The onset of collagen induced arthritis was preceded by an increase of anti-CarP IgG2a levels in the serum.

Conclusion: In mice with collagen induced arthritis we did not observe an immune response against citrullinated antigens, but we did observe an immune response against carbamylated antigens. This anti-CarP response already appeared before disease onset, indicating that collagen induced arthritis can be used as an in vivo model to study anti-CarP antibodies. Our data also indicate that the tolerance to carbamylated proteins, in contrast to the response to citrullinated proteins, is easily broken and that arthritis boosts the immune response against these proteins. The anti-CarP response in mice with CIA can be used as a model for immune responses to post-translationally modified proteins.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibody Specificity / immunology*
  • Arthritis, Experimental / immunology*
  • Autoantibodies / blood
  • Autoantibodies / immunology*
  • Carbamates / metabolism*
  • Enzyme-Linked Immunosorbent Assay
  • Immunoglobulin Isotypes / blood
  • Immunoglobulin Isotypes / immunology
  • Mice
  • Protein Processing, Post-Translational
  • Proteins / immunology*
  • Proteins / metabolism*

Substances

  • Autoantibodies
  • Carbamates
  • Immunoglobulin Isotypes
  • Proteins

Grant support

This work was supported by the IMI JU funded project BeTheCure, contract no 115142-2, the Dutch Arthritis Foundation, The Netherlands Organization for Scientific Research and Pfizer. Leendert Trouw receives a fellowship from Janssen biologics BV. Martin Hegen (who is an employee of Pfizer) has been involved in the design of the study and in the preparation of the manuscript in a scientific collaboration. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.