Investigation of a Set of Dysregulated Tear Proteins of Sjögren’s Syndrome Patients Using In Vitro Models Uncovers New Insights and Potential Candidate Biomarkers

Abstract

Purpose : The validated biomarkers in dry eye disease and Sjögren’s syndrome (SS) are still an unmet need. Therefore, we investigated dysregulations in the tear proteome of primary SS (pSS) patients to identify disease-associated potential candidate biomarkers.

Methods : Schirmer strips were collected from pSS patients (n=12) and healthy controls (n=6). Proteome from each strip was analyzed using timsTOF Pro mass spectrometry. Proteins were identified using MaxQuant® and their functional analysis was performed using Cytoscap®e and Reactome®. A panel of targets of interest (TOI) was selected based on a set of criteria. Immortalized human corneal epithelial cells (HCE) were exposed to hyperosmolarity (HO, 500 mOsm/l) or proinflammatory cytokines (10 ng/ml IL-1β and IFN-γ) for 4h and 24h to analyze gene expressions of TOI using RT-qPCR. Statistical analysis was performed using Limma statistical test (proteomics data) and Student's t-test.

Results : Over a hundred proteins were significantly modulated in pSS patients. Tear glycoproteins (LACRT, CLU, PRR3 and PRP27) were highly downregulated, suggesting a disruption of cell-protecting mechanisms. Functional analysis of all dysregulated proteins highlighted an alteration in cytoskeleton organization and cell metabolism. Apoptosis-initiator protein CASP3 was overexpressed in pSS patients. Of the selected 25 TOI, PLA2G2A, CXCL17, PRDX6, TMSB4X and MYH9 were modulated in the in vitro models. HO significantly increased the gene expressions of PLA2G2A, PRDX6, CXCL17 and TMSB4X in HCE after 24h. Likewise, IFN-γ and IL-1β significantly increased the gene expression of CXCL17 (4.5-fold) and MYH9 (1.5-fold), respectively.

Conclusions : PLA2G2A, PRDX6, CXCL17, TMSB4X and MYH9 were modulated both in pSS patients and in the in vitro models, suggesting that they might contribute to the disease pathophysiology and could possess characteristics of potential tears biomarkers.

This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.