Background: Sjögren’s disease (SjD) is an autoimmune disorder characterized by impaired exocrine gland function, primarily due to lymphocyte infiltration and subsequent damage to the salivary glands. Macrophage migration inhibitory factor (MIF), as a pleiotropic pro-inflammatory cytokine, plays a critical role in autoimmune diseases such as rheumatoid arthritis and systemic lupus erythematosus through its interaction with its receptor, CD74. However, the specific mechanisms of the MIF-CD74 signaling axis in the pathogenesis of SjD remain unclear.

Objectives: This study aims to investigate the role of the MIF-CD74 axis in the pathogenesis of SjD and to elucidate its underlying molecular mechanisms in driving inflammatory responses in salivary gland epithelial cells and promoting B-cell infiltration.

Methods: Labial gland samples from patients with SjD and non-SjD controls were analyzed using single-cell RNA sequencing (scRNA-seq). This approach revealed distinct gene expression profiles, transcription factor activities, and cell-cell communication networks, highlighting pathogenic pathways potentially involved in SjD. Subsequently, ELISA and immunofluorescence confocal microscopy were employed to detect the expression and localization of MIF and CD74 in the serum and labial glands of these cohorts, followed by correlation analysis with clinical parameters. To further investigate these findings in vivo , an experimental SjD (ESjD) mouse model was established. Serum MIF levels were compared between model and control mice, and the expression and localization of MIF and CD74 in the submandibular glands were examined. Flow cytometry was then used to analyze the expression of MIF and CD74 across different immune cell subsets within the submandibular glands of ESjD mice versus controls. Finally, to assess the therapeutic potential of targeting this axis, NOD mice were intraperitoneally administered the MIF inhibitor ISO-1, and lymphocyte infiltration in the submandibular glands was evaluated using flow cytometry and immunofluorescence.

Results: Single-cell RNA sequencing analysis of labial glands from 11 patients with SjD and 5 non-SS controls revealed significant cellular heterogeneity. Specifically, MIF expression was markedly elevated in ductal epithelial cells of SjD patients compared to controls (P < 0.0001), while its receptor CD74 was highly expressed on infiltrating B cells (P < 0.0001). Immunofluorescence and flow cytometry further confirmed that MIF was primarily secreted by ductal epithelial cells and was significantly upregulated in pSS compared to patients with sicca symptoms alone. Correspondingly, CD74 expression on B cells was also significantly higher in the SjD group. Consistently, serum MIF levels were significantly elevated in SjD patients compared to healthy controls (P < 0.01) and showed positive correlations with serum immunoglobulin levels and the ESSDAI score. In an experimental SjD (ESjD) mouse model, the expression levels of MIF in both serum and submandibular gland epithelial cell culture supernatants were significantly higher than those in naive mice. Mif mRNA expression was significantly upregulated in submandibular glands compared to controls (P < 0.0001). Immunofluorescence staining localized MIF predominantly to ductal epithelial cells and CD74 to B cells. Flow cytometry confirmed increased MIF expression in ductal epithelial cells (P < 0.05) and elevated CD74 on B cells (P < 0.01) in ESjD mice versus controls. Therapeutic targeting of the MIF-CD74 axis was evaluated in NOD mice administered the MIF inhibitor ISO-1 (20 mg/kg/day, i.p.) for 8 weeks. This treatment significantly reduced lymphocyte infiltration in submandibular glands, increased salivary flow rate, and effectively ameliorated disease pathology, thereby restoring salivary function.

Conclusions: This study demonstrates that the MIF-CD74 signaling axis is aberrantly activated in SjD. Epithelial-derived MIF contributes to and exacerbates local salivary gland inflammation and B cell infiltration by engaging the CD74 receptor on B cells. Furthermore, administration of an MIF inhibitor was shown to alleviate B cell infiltration in the submandibular glands of NOD mice and delay disease progression. These results collectively suggest that targeted inhibition of the MIF-CD74 axis may represent a promising novel therapeutic strategy for SjD.

REFERENCES: NIL.

Acknowledgments: NIL.

Disclosure of Interests: None declared.