Elsevier

Clinical Biochemistry

Volume 48, Issue 3, February 2015, Pages 181-185
Clinical Biochemistry

Short Communication
Proteomic profiling of antigens in circulating immune complexes associated with each of seven autoimmune diseases

https://doi.org/10.1016/j.clinbiochem.2014.11.008Get rights and content

Highlights

  • We applied immune complexome analysis to the study of seven autoimmune diseases.

  • We identified 468 distinct immune complexes-associated antigens using this method.

  • Importantly, 62 of those antigens were disease-specific antigens.

Abstract

Objective

Immune complexes (ICs) trigger humoral immune responses. Therefore, the identification of constituent antigens within ICs would have very different clinical significance than identification of free antigens.

Design and methods

Here, we applied immune complexome analysis of serum to the study of seven major autoimmune diseases—anti-neutrophil cytoplasmic antibody-associated vasculitis, Takayasu's arteritis, mixed connective tissue disease, dermatomyositis, Sjögren's syndrome, systemic scleroderma, and systemic lupus erythematosus—and healthy donors to comprehensively identify antigens incorporated into circulating ICs and to find disease-specific antigens.

Results

We identified 468 distinct IC-associated antigens using this method. Importantly, 62 of those antigens were disease-specific antigens, and there were at least three disease-specific antigens for each of the seven autoimmune diseases. Of the disease-specific antigens identified, coiled-coil domain-containing protein 158 and spectrin were identified as potential autoantigens important to SSc and SS pathogenesis, respectively; notable titin and spectrin autoantibodies are reportedly found in SSc and SS patients, respectively.

Conclusion

Immune complexome analysis may be generally applicable to the study of the relationship between ICs and autoimmune diseases in animals and humans.

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.

References (15)

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