Short CommunicationProteomic profiling of antigens in circulating immune complexes associated with each of seven autoimmune diseases
Introduction
For a long time, immune complexes (ICs) assembly were thought to represent a common pathogenic pathway for several diseases (infections, vasculitis, and connective tissue autoimmune disorders). Actually, concentrations of circulating ICs (CICs) in sera from patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), or systemic scleroderma (SSc) were significantly higher than those in sera from healthy controls [1], [2]. Many researchers have investigated the mechanisms by which ICs could underlie pathogenicity [3]. An autoimmune response is directed against several autoantigens [4], [5]; therefore, comprehensive profiling of the autoantigens that actually assemble into ICs may provide insight into the pathophysiology of specific autoimmune diseases, and such profiling could form the basis for novel diagnosis and treatment strategies for these diseases. However, such comprehensive profiling studies for CICs are limited because tools for screening of ICs are lacking.
We developed a proteomic strategy, designated immune complexome analysis, in which ICs are separated from whole serum and then subjected to direct tryptic digestion and nano-liquid chromatography–tandem mass spectrometry (nano-LC–MS/MS) to comprehensively identify and profile constituent antigens in CICs [6]. We used this method to identify CIC-associated antigens in sera from patients with RA, and found that thrombospondin-1 is a constituent of CICs and is more highly specific and sensitive for established and early RA than other conventional diagnostic markers such as rheumatoid factor or anti-citrulline-containing protein/peptide antibody [6], [7].
In this report, we used immune complexome analysis of serum to study seven major autoimmune diseases—anti-neutrophil cytoplasmic antibody-associated vasculitis (AAV), Takayasu's arteritis (TA), mixed connective tissue disease (MCTD), dermatomyositis (DM), Sjögren's syndrome (SS), SSc, and SLE—to comprehensively identify antigens incorporated into CICs and find disease-specific antigens.
Section snippets
Materials and methods
Serum samples were collected from 66 patients; each patient had AAV (n = 7; 35–86 years; 3 females; 4 patients have microscopic polyantitis and 3 patients have granulomatosis with polyantitis), TA (n = 7; 28–63 years; 7 females), MCTD (n = 9; 43–77 years; 9 females), DM (n = 8; 30–69 years; 6 females), SS (n = 14; 35–78 years; 14 females), SSc (n = 7; 55–78 years; 6 females), or SLE (n = 14; 16–67 years; 13 females) as diagnosed based on classification criteria from the Ministry of Health, Labour and Welfare, Japan
Results and discussion
Here, we present the first comprehensive identification of the constituent antigens assembled into CICs from patients with AAV, TA, MCTD, DM, SS, SSc, or SLE (Table S1). We identified 341 and 264 human antigens via immune complexome analysis with Protein G or Protein A beads, respectively; 468 distinct antigens were identified in all, and each of these was found in independent samples from two or more patients (Table S2). Notably, only 29% (137) of these 468 distinct antigens were recovered
Conflict of interest
Research funding: K. Ohyama, Eisai Co., Ltd.
Acknowledgment
This work was supported by a grant-in-aid for Scientific Research (C) and Challenging Exploratory Research from the Ministry of Education, Culture, Sports, Science and Technology of Japan, a grant-in-aid for Scientific Research from Nagasaki University, the Joint Research Promotion Project of Nagasaki University Graduate School of Biomedical Sciences in 2013, the Research Foundation for Pharmaceutical Sciences and the Kato Memorial Trust For Nambyo Research.
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