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. 2020 Dec;9(6):2054-2063.
doi: 10.21037/gs-20-814.

Proteomics-based analysis indicating α-enolase as a potential biomarker in primary Sjögren's syndrome

Pan Wei  1 , Yixiao Xing  1 , Boya Li  1 , Feng Chen  2 , Hong Hua  1
Affiliations

Proteomics-based analysis indicating α-enolase as a potential biomarker in primary Sjögren's syndrome

Pan Wei et al. Gland Surg. 2020 Dec.

Abstract

Background: Primary Sjögren's syndrome (pSS) is a chronic autoimmune disease. Its etiology is not well understood. Salivary glands are the main target organ in pSS, investigating the changes of salivary protein in pSS patients may not only be a valuable way of identifying new biomarkers/antigens for pSS, but also of revealing the pathogenesis underlying this autoimmune disease. In the present study, we aimed to investigate new biomarkers and explore their potential role in pSS.

Methods: In this study, α-enolase (ENO1) was found to be overexpressed in pSS by 1D gel electrophoresis/mass spectrometry. The finding was verified by Western blots, immunohistochemistry (IHC), and polymerase chain reaction (PCR) results in both saliva and labial salivary glands. The expression level of immunoglobulin G (IgG) antibody to ENO1 was then tested by enzyme-linked immunosorbent assay (ELISA).

Results: ENO1 autoantibody was found to be overexpressed in pSS compared with healthy controls. The effects of ENO1 overexpression on rat submandibular gland cell line SMG-C6 was investigated in vitro. The expressions of proteins related to saliva secretion and immunomodulatory were upregulated in ENO1 overexpressed SMG-C6 cells.

Conclusions: Both ENO1 and anti-ENO1 autoantibody are overexpressed in pSS patients. Nevertheless, their potential role in the pathogenesis of pSS warrants further study.

Keywords: Primary Sjögren’s syndrome (pSS); mass spectrometry; proteomics-based analysis; α-enolase (ENO1).

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Conflict of interest statement

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at http://dx.doi.org/10.21037/gs-20-814). The authors have no conflicts of interest to declare.

Figures

Figure 1
Figure 1
Western blot analysis of salivary ENO1 in saliva from pSS patients (n=8) and healthy controls (n=8). Results indicate that the salivary ENO1 level was much higher in pSS patients compared with healthy controls. ENO1, α-enolase-1; pSS, primary Sjögren’s syndrome; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.
Figure 2
Figure 2
Reverse transcription-polymerase chain reaction analysis of ENO1 mRNA in the labial glands of pSS patients (n=5) and healthy controls (HC; n=4). Results showed that ENO1 mRNA was significantly overexpressed in the labial glands of pSS patients compared with healthy controls. *P<0.05. ENO1, α-enolase-1; pSS, primary Sjögren’s syndrome; HC, healthy controls.
Figure 3
Figure 3
Immunohistochemical staining of labial glands. (A) ENO1 proteins were highly expressed in the acinar and duct epithelial cells in pSS patients. (B) ENO1 expression was lower in healthy controls. (C) ENO1 was not expressed in infiltrated lymphocytes. ENO1, α-enolase-1; pSS, primary Sjögren’s syndrome. The scale is 1:100 µm.
Figure 4
Figure 4
Analyses of anti-ENO1 autoantibody in serum samples from pSS patients and healthy controls. (A) Levels of serum anti-ENO1 autoantibody were significantly higher in pSS patients compared with healthy controls (P<0.01). (B) ROC curve analysis was performed to evaluate the performance of anti-ENO1 in distinguishing between pSS and healthy controls. Area under the curve value of anti-ENO1 was 0.811. (C) Linear regression analysis showed that anti-ENO1 was moderately associated with salivary flow rate by linear regression (n=26, R2=0.5056). **, P<0.01. ENO1, α-enolase; pSS, primary Sjögren’s syndrome; ROC, receiver-operating characteristic.
Figure 5
Figure 5
Biologic process of differentially regulated proteins with PANTHER (http://www.pantherdb.org).
See this image and copyright information in PMC

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