Autoantibody recognition of collagen type II in arthritis

Autoantibodies against collagen type II (CII), a protein localized in the joint cartilage, play a major role in collagen-induced arthritis (CIA), one of the most commonly used animal models for rheumatoid arthritis (RA). The studies included in this thesis were undertaken to elucidate structural and functional requirements for B and T cells to recognize native CII structures during experimental arthritis as well as in human RA. To reveal in detail how CII-specific autoantibodies recognize CII, we determined the crystal structure of two pathogenic immune complexes occurring in arthritis. The two crystal structures reveal how autoantibodies target triple-helical CII and specifically which epitope sequences and CDR residues that are crucial for binding. Interestingly, although amino acid residues in the hypervariable regions of both autoantibodies were somatically mutated, the majority of the contacts with the epitope involve germlineencoded structures. A recombinant CII peptide library was also generated in order to reveal the CII epitope specificity of polyclonal autoantibodies in CIA. This library was used to confirm already known CII epitopes occurring in CIA but also to investigate new CII epitope regions that could play a role in arthritis. Several new regions targeted by autoantibodies were found and interestingly, antibodies to these regions were also identified in non-human primate species with arthritis. Subsequently, autoantibodies to the major CII epitopes C1, U1, and J1 were analyzed in serum and synovial fluid from RA patients. The autoantibody levels to all three CII epitopes were significantly higher in synovial fluid compared to serum, in particular autoantibodies to the U1 epitope. Finally, we studied the CII-specific antibody responses as well as CII-specific T cell responses during different timepoints of CIA in two mouse strains, B10.Q and a humanized mouse strain B10.DR4. Ncf1*/*. The obtained data showed that the antibody and T cell responses investigated stayed relatively constant during the disease course and were dominated by T cells specific for non-modified (K264) and hydroxylated (HOK264) CII259-273 peptides in the humanized mice and to the galactosylated (GalOK264) CII259-273 peptide in the B10.Q mice. C1 specific antibodies dominated in both strains while antibodies to the U1 epitope was characteristic for the humanized mouse. LIST OF PUBLICATIONS I. Analysis of autoantibodies to the joint-specific protein collagen II in experimental and rheumatoid arthritis Ingrid Lindh, Hüseyin Uysal, Omri Snir, Erik Lönnblom, Ida Andersson, Franziska Lange, Michel Vierboom, Kutty Selva Nandakumar, Vivianne Malmström, and Rikard Holmdahl Manuscript II. Crystal structure of an arthritogenic anticollagen immune complex Doreen Dobritzsch*, Ingrid Lindh*, Hüseyin Uysal, Kutty Selva Nandakumar, Harald Burkhardt, Gunter Schneider, and Rikard Holmdahl Arthritis Rheum. 2011 Dec;63(12):3740-8. III. Comparative analysis of CII-specific immune responses during development of collagen induced arthritis in two B10 strains Tsvetelina Batsalova*, Ingrid Lindh*, Johan Bäcklund, Balik Dzhambazov, and Rikard Holmdahl Arthritis Res Ther. 2012 Nov 1;14(6):R237. IV. Immunoglobulin heavy chain V gene polymorphism controls epitopespecific antibody response Bruno Raposo, Doreen Dobritzsch, Ingrid Lindh, Diana Ekman, Changrong Ge, Michael Förster, Hüseyin Uysal, Gunter Schneider, and Rikard Holmdahl Manuscript *These authors contributed equally to the work Additional publication not included in the thesis: Tissue transglutaminase enhances collagen type II-induced arthritis and modifies the immunodominant T-cell epitope CII260-270 Balik Dzhambazov, Ingrid Lindh, Åke Engström, and Rikard Holmdahl Eur J Immunol. 2009 Sep;39(9):2412-23.