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. 2018 Feb 12;20(1):30.
doi: 10.1186/s13075-017-1508-5.

Identification of a novel autoantibody against self-vimentin specific in secondary Sjögren's syndrome

Yu-Hui Li  1 , Ya-Ping Gao  2 , Jie Dong  2 , Lian-Jie Shi  1 , Xiao-Lin Sun  1 , Ru Li  1 , Xue-Wu Zhang  1 , Yu Liu  1 , Li Long  1 , Jing He  1 , Qun-Jie Zhong  3 , Eric Morand  4 , Guang Yang  5 , Zhan-Guo Li  6
Affiliations

Identification of a novel autoantibody against self-vimentin specific in secondary Sjögren's syndrome

Yu-Hui Li et al. Arthritis Res Ther. .

Abstract

Background: Sjögren's syndrome (SS) is a primary autoimmune disease (pSS) or secondarily associated with other autoimmune diseases (sSS). The mechanisms underlying immune dysregulation in this syndrome remain unknown, and clinically it is difficult to diagnose owing to a lack of specific biomarkers.

Methods: We extracted immunoglobulins (Igs) from the sera of patients with sSS associated with rheumatoid arthritis (RA) and used them to screen a phage display library of peptides with random sequences.

Results: Our results show that an sSS-specific peptide, designated 3S-P, was recognized by sera of 68.2% (60 of 88) patients with sSS, 66.2% of patients with RA-sSS, and 76.5% of patients with systemic lupus erythematosus (SLE)-sSS. The anti-3S-P antibody was scarcely found in patients with pSS (1.8%), RA (1.3%), SLE (4.2%), ankylosing spondylitis (0%), and gout (3.3%), as well as in healthy donors (2%). The 3S-P-binding Igs (antibodies) were used to identify antigens from salivary glands and synovial tissues from patients with sSS. A putative target autoantigen expressed in the synovium and salivary gland recognized by anti-3S-P antibody was identified as self-vimentin.

Conclusions: This novel autoantibody is highly specific in the diagnosis of sSS, and the underlying molecular mechanism of the disease might be epitope spreading involved with vimentin.

Keywords: Autoantigen; Biomarker; Sjögren’s syndrome; Vimentin.

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Conflict of interest statement

Ethics approval and consent to participate

All the procedures involving specimens obtained from human subjects were performed under protocols approved by the Human Research Protective Committee of People’s Hospital Peking University, and all the methods were carried out in accordance with the guidelines and regulations of the Ethics Committee of People’s Hospital Peking University. Informed consent was also provided by all subjects before the study protocol was initiated.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

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Figures

Fig. 1
Fig. 1
Antibodies to 3S-P in serum samples from patients and healthy control subjects. a ELISA results of detection of anti-3S-P IgG in the subjects’ sera. Each circle represents one patient. b Sensitivity and specificity of the assay of IgG antibodies against 3S-P for differentiating between patients with sSS and control subjects. The ROC curve indicates the level of antibodies against 3S-P in patients with sSS compared with that in patients with RA, pSS, or SLE. sSS Secondary Sjögren’s syndrome associated with rheumatoid arthritis, RA Rheumatoid arthritis, pSS Primary Sjögren’s syndrome, SLE Systemic lupus erythematosus, AS Ankylosing spondylitis, HC Healthy control subjects, 3S-P Secondary Sjögren’s syndrome-associated peptide, IgG Immunoglobulin G
Fig. 2
Fig. 2
Immunohistochemical analysis of tissues and cells using anti-3S-P antibodies. a Expression of anti-3S-P target antigens in RA-sSS synovial tissue and salivary tissue. RA-sSS synovium was stained using antibodies against KLH and R-anti-3S-P. RA-sSS salivary gland tissue was stained using R-anti-3S-P and anti-KLH as control (original magnification × 400). b Confocal immunofluorescence microscopic images of FLSs reacted with R-anti-3S-P or anti-KLH antibody. Antibody staining (FITC, green), and nuclear staining (DAPI, blue) are shown. Scale bar represents 20 μm. 3S-P Secondary Sjögren’s syndrome-associated peptide, DAPI 4′6-Diamidino-2-phenylindole, FITC Fluorescein isothiocyanate, FLS Fibroblast-like synoviocyte, KLH Keyhole limpet hemocyanin, RA Rheumatoid arthritis, R-anti-3S-P Rabbit-anti-secondary Sjögren’s syndrome-associated peptide, sSS Secondary Sjögren’s syndrome
Fig. 3
Fig. 3
Two-dimensional PAGE of total proteins extracted from FLSs of patients with RA-sSS and Western blot analysis. a Western blot showing immunostaining of FLS proteins with R-anti-3S-P and anti-KLH, respectively. b Colloidal Coomassie Blue staining of two-dimensional gel of FLS proteins. Protein spots indicated by circles were further identified using matrix-assisted laser desorption/ionization time-of-flight MS. Mascot search results showed protein scores of 580, 1113, and 343 for human caldesmon isoform 2 (Cal), vimentin (Vim), and 60 kDa heat shock protein (HSP-60), respectively. FLS Fibroblast-like synoviocyte, KLH Keyhole limpet hemocyanin, RA Rheumatoid arthritis, R-anti-3S-P Rabbit-anti-secondary Sjögren’s syndrome-associated peptide, sSS Secondary Sjögren’s syndrome
Fig. 4
Fig. 4
Western blot analysis of proteins extracted from FLSs using antibodies against caldesmon (lane 1), HSP60 (lane 2), and antivimentin (lane 3) as probes. β-Actin was used as an internal control. FLS Fibroblast-like synoviocyte, HSP60 Heat shock protein 60
Fig. 5
Fig. 5
Identification of vimentin as the target autoantigen of anti-3S-P autoantibodies. a Autoantigens of anti-3S-P and vimentin are present in immunoprecipitates (IP) obtained by using antivimentin antibody from lysates of FLSs, whereas no band was seen in negative controls. b ELISA data for the detection of recombinant human vimentin using the R-anti-3S-P antibody diluted 1:50 to 1:800. The anti-KLH antibody was used as an isotype control and was diluted 1:50 to 1:800. c Western blot (WB) of recombinant human vimentin probed with antivimentin, R-anti-3S-P, and anti-KLH antibodies. ELISA Enzyme-linked immunosorbent assay, FLS Fibroblast-like synoviocyte, KLH Keyhole limpet hemocyanin, R-anti-3S-P Rabbit-anti-secondary Sjögren’s syndrome-associated peptide, SDS Sodium dodecyl sulfate
Fig. 6
Fig. 6
Peptide sequence homologies. Identity is indicated by solid lines, and a conservative substitution is indicated by the dotted line
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