Peroxiredoxin 2 is a novel autoantigen for anti-endothelial cell antibodies in systemic vasculitis

Abstract

SummaryAnti-endothelial cell antibodies (AECA) have been frequently detected in systemic vasculitis, which affects blood vessels of various sizes. To understand the pathogenic roles of AECA in systemic vasculitis, we attempted to identify target antigens for AECA comprehensively by a proteomic approach. Proteins extracted from human umbilical vein endothelial cells (HUVEC) were separated by two-dimensional electrophoresis, and Western blotting was subsequently conducted using sera from patients with systemic vasculitis. As a result, 53 autoantigenic protein spots for AECA were detected, nine of which were identified by mass spectrometry. One of the identified proteins was peroxiredoxin 2 (Prx2), an anti-oxidant enzyme. Frequency of anti-Prx2 autoantibodies, measured by enzyme-linked immunosorbent assay (ELISA), was significantly higher in systemic vasculitis (60%) compared to those in collagen diseases without clinical vasculitis (7%, P < 0·01) and healthy individuals (0%, P < 0·01). Further, the titres changed in parallel with the disease activity during time-courses. The presence of anti-Prx2 autoantibodies correlated significantly with elevation of serum d-dimers and thrombinantithrombin complex (P < 0·05). Immunocytochemical analysis revealed that live endothelial cells expressed Prx2 on their surface. Interestingly, stimulation of HUVEC with rabbit anti-Prx2 antibodies increased secretion of interleukin (IL)-6, IL-1b, IL-1ra, growth regulated oncogene (GRO)-a, granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colonystimulating factor (GM-CSF), IL-8 and monocyte chemoattractant protein (MCP)-1 more than twofold compared to that of with rabbit immunoglobulin (Ig)G. Taken together, our data suggest that anti-Prx2 autoantibodies would be a useful marker for systemic vasculitis and would be involved in the inflammatory processes of systemic vasculitis.

Thioredoxin and Thioredoxin Target Proteins: From Molecular Mechanisms to Functional Significance

“…In addition to its role in diseases associated with increased ROS, autoantibodies against Prxs have been associated with autoimmune diseases such as Kawasaki disease, systemic sclerosis, and systemic vasculitis (84,125,139). In these studies, sera from patients with and without autoimmune diseases were screened for autoantibodies against different isoforms of Prx.…”

Persistent Endothelial Damage after Intravenous Immunoglobulin Therapy in Kawasaki Disease

“…Recognition of autoantigens by antiendothelial cell antibodies has been reported [8,9]. We previously demonstrated a marked elevation of a B-cell-activating factor belonging to the TNF family (BAFF) [10] before intravenous immunoglobulin therapy (IVIG), and its steep drop after the therapy [11], which suggest that B-cell activation in the acute phase might be involved in the pathogenesis of KD.…”

Validation of a New Biomarker in Patients with Kawasaki Disease Identified by Proteomics

“…Currently, we have detected more than 150 candidate antigenic spots, and identified more than 50 protein spots. In our earlier study, we identified Peroxiredoxin-2 (Prx2), a member of the Peroxiredoxin (Prx) family of peroxidases, as a target antigen for AECA [9]. In mammalian cells, the Prx family has at least six members (Table 1), and Prx2 is one of the most rapid and potent responders to oxidative stress.…”

“…To evaluate the roles of AECA in detail, it is essential that their target antigens are identified and that the role of individual target antigens is assessed. Various approaches, such as expression libraries and proteomic, have been developed to identify biomarkers [6][7][8][9][10][11]. In KD, two target antigens for the AECA, tropomyosin and T-plastin, were identified using serological analysis of a recombinant cDNA expression library [11].…”

“…These kinds of analyses may lead to clinical applications. To identify the target proteins for AECA in patients with vasculitis, we used a proteomics approach that included 2-dimensional electrophoresis and Western Blotting, followed by mass spectrometry [9]. Briefly, we extracted the proteins from HUVEC and HeLa cells (control), and separated them by 2-dimensional electrophoresis to detect AECA antigens specific for ECs.…”