Part 2 of this thesis focuses on the role of posttranslational modifications in breaking tolerance to self-‐antigens in susceptible individuals. Oxidative modifications of known autoantigens were identified in lysates from hydrogen peroxide-‐induced necrotic cells. It is unclear if oxidative modifications are involved in the pathogenesis of autoimmune diseases. Several questions still need to be answered. Are the observed in vitro modifications also present in vivo and, more importantly, are proteins that are modified upon oxidative stress present in affected tissues of autoimmune patients? Do healthy individuals also have such proteins and do these differ between healthy and diseased individuals? Can these modified proteins induce autoimmunity in animal models? CVB-‐3 infected Min6 cell lysates contain a Mr 100,000 protein which was recognized more frequently by T1D sera than by control sera. This Mr 100,000 protein most likely is a cellular Min6 protein. The data described in this thesis may facilitate follow-‐up studies to obtain more insight in the role of posttranslational modifications in autoimmunity.
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Chapter 8
Summary
Samenvatting
List of publications
Curriculum vitae
Dankwoord
Summary
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Summary
Autoimmune diseases arise from an inappropriate immune response towards substances and tissues normally present in the body. One of the characteristic features of autoimmune diseases is the occurrence of high titers of autoantibodies in the sera of these patients. In sera of patients suffering from rheumatoid arthritis (RA) autoantibodies are found targeting citrullinated proteins, and these autoantibodies sometimes can be detected years before disease onset. These anti-‐citrullinated protein antibodies (ACPA) are exclusively found in RA patients and recently it was shown that the RA patients who don’t produce ACPA represent a specific subpopulation of patients with a different prognosis than ACPA-‐positive RA patients, making ACPA a very good diagnostic and prognostic tool.
Although the clinical utility of these autoantibodies is appreciated, little is known about the mechanisms related to their production and the regulation of the autoimmune response. One model hypothesizes that posttranslational modifications occurring during cell death can result in the generation of neo-‐ epitopes and the loss of immunological tolerance to self-‐antigens.
The increasing number of distinct autoantibodies and the diversity of autoantibodies with similar or related specificities motivate the need for assays in which multiple autoantibodies in the same sample can be detected simultaneously. The goal of the first studies described in this thesis was to develop a multiplex assay based on imaging surface plasmon resonance (iSPR), which can be used to subdivide RA patients based on their autoantibody profile (described in Part 1: Chapters 2 to 4). Another goal was to gain more insight in the development of autoimmune diseases and to investigate the role of cell death-‐ associated protein modifications in the initiation of the autoimmune response (described in Part 2: Chapters 5 and 6).
This thesis starts with an introductory chapter, Chapter 1, which gives an overview of our current knowledge on the generation of autoantibodies, with a special focus on the proposed role of cell death-‐associated protein modifications in breaking immunological tolerance. In addition, the iSPR technology for measuring autoantibody profiles in a multiplex format is introduced.
The presence of (auto)antibodies in biofluids can be measured by various experimental approaches, such as enzyme linked immunosorbent assay (ELISA), immunodiffusion, immunoprecipitation, immunoblotting, etc. Multiplex platforms have the advantage to test for the presence of multiple autoantibodies