Immunoglobulin G4-related sialadenitis (IgG4-RS) is an immune-mediated fibro-inflammatory disease and the pathogenesis is still not fully understood. The aim of this study was to explore the role and mechanism of interleukin-13 (IL-13) in the cellular senescence during the progress of IgG4-RS. We found that the expression of IL-13 and IL-13 receptor α1 (IL-13Rα1) as well as the number of senescent cells were significantly higher in the submandibular glands (SMGs) of IgG4-RS patients. IL-13 directly induced senescence as shown by the elevated activity of senescence-associated β-galactosidase (SA-β-gal), the decreased cell proliferation, and the upregulation of senescence markers (p53 and p16) and senescence-associated secretory phenotype (SASP) factors (IL-1β and IL-6) in SMG-C6 cells. Mechanistically, IL-13 increased the level of phosphorylated signal transducer and activator of transcription 6 (p-STAT6) and mitochondrial-reactive oxygen species (mtROS), while decreased the mitochondrial membrane potential, ATP level, and the expression and activity of superoxide dismutase 2 (SOD2). Notably, the IL-13-induced cellular senescence and mitochondrial dysfunction could be inhibited by pretreatment with either STAT6 inhibitor AS1517499 or mitochondria-targeted ROS scavenger MitoTEMPO. Moreover, IL-13 increased the interaction between p-STAT6 and cAMP-response element binding protein (CREB)-binding protein (CBP) and decreased the transcriptional activity of CREB on SOD2. Taken together, our findings revealed a critical role of IL-13 in the induction of salivary gland epithelial cell senescence through the elevated mitochondrial oxidative stress in a STAT6–CREB–SOD2-dependent pathway in IgG4-RS.

Objective IgG4‐related sialadenitis (IgG4‐RS) is a chronic fibroinflammatory disease characterized by glandular fibrosis and hyposalivation. This study was undertaken to explore the role of cellular senescence in the pathogenesis of IgG4‐RS–related fibrosis. Methods The expression of senescence markers and proinflammatory cytokines in the submandibular glands (SMGs) of IgG4‐RS patients (n = 18) and controls (n = 14) was determined by proteomics, real‐time polymerase chain reaction, Western blotting, and immunohistochemistry. After interleukin‐4 (IL‐4) treatment, high‐throughput RNA sequencing was performed to identify the differentially expressed genes in SMG‐C6 cells. A glandular fibrosis model was established by the intraglandular injection of IL‐4 into mouse SMGs (n = 8 per group). Results Salivary acinar and ductal epithelial cells underwent senescence in IgG4‐RS patients, as indicated by the elevated activity of senescence‐associated β‐galactosidase, lipofuscin accumulation, enhanced expression of senescence markers (p53 and p16INK4A), and up‐regulation of senescence‐associated secretory phenotype factors. Moreover, there was a significant increase in IL‐4 levels in SMGs from IgG4‐RS patients (P < 0.01), which positively correlated with p16INK4A expression and the fibrosis score. Incubation with IL‐4 exacerbated salivary epithelial cell senescence by increasing the expression of p16INK4A through the reactive oxygen species (ROS)/p38 MAPK pathway. Supernatant collected from IL‐4–induced senescent SMG‐C6 cells enhanced fibroblast activation and matrix protein production (P < 0.05). Furthermore, injecting mice with IL‐4 promoted fibrosis and senescence phenotypes in SMGs in vivo. Conclusion The cellular senescence induced by IL‐4 through the ROS/p38 MAPK‐p16INK4A pathway promotes fibrogenesis in IgG4‐RS. Our data suggest that cellular senescence could serve as a novel therapeutic target for treating IgG4‐RS.

Tight junctions (TJs) are cell–cell interactions that localize at the most apical portion of epithelial/endothelial cells. One of the predominant functions of TJs is to regulate material transport through paracellular pathway, which serves as a selective barrier. In recent years, the expression and function of TJs in salivary glands has attracted great interest. The characteristics of multiple salivary gland TJ proteins have been identified. During salivation, the activation of muscarinic acetylcholine receptor and transient receptor potential vanilloid subtype 1, as well as other stimuli, promote the opening of acinar TJs by inducing internalization of TJs, thereby contributing to increased paracellular permeability. Besides, endothelial TJs are also redistributed with leakage of blood vessels in cholinergic‐stimulated submandibular glands. Furthermore, under pathological conditions, such as Sjögren's syndrome, diabetes mellitus, immunoglobulin G4‐related sialadenitis, and autotransplantation, the integrity and barrier function of TJ complex are impaired and may contribute to hyposalivation. Moreover, in submandibular glands of Sjögren's syndrome mouse model and patients, the endothelial barrier is disrupted and involved in hyposecretion and lymphocytic infiltration. These findings enrich our understanding of the secretory mechanisms that link the importance of epithelial and endothelial TJ functions to salivation under both physiological and pathophysiological conditions.