Search

Background Salivary glands of primary Sjogren’s syndrome (pSS) are characterised by complex leukocyte infiltration organised into tertiary lymphoid structures (TLS). The mechanisms regulating leukocyte trafficking into inflamed salivary glands are poorly described, but dysregulated T-cell recruitment during inflammation is believed to contribute to disease onset and chronicity. We recently described a homeostatic pathway in which a B cell-derived peptide (PEPITEM), secreted in response to adiponectin, regulates T-cell trafficking during inflammation via sphingosine-1-phosphate activity on endothelial cells1. Disruption of this pathway by downregulation of adiponectin receptors on circulating B cells has been demonstrated in type-1 diabetes and rheumatoid arthritis, suggesting a potential role for PEPITEM in the pathogenesis of autoimmune diseases, and indicating a role for adiponectin receptors as biomarkers in these conditions1. Objectives We aimed to investigate the efficacy of PEPITEM as an inhibitor of T-cell trafficking in an inducible animal model of salivary gland inflammation that mimics the histological features of pSS, and to investigate the potential translatability of this pathway in patients with pSS. Methods Submandibular salivary glands of C57BL/6 mice were intra-ductally cannulated with luciferase-encoding replication-deficient adenovirus to induce TLS formation as previously described2. Mice were administered daily either PBS or PEPITEM by intraperitoneal injection from day 0, and their salivary glands dissected at day 5 post cannulation. T-cell infiltration into salivary glands was assessed using a combination of flow cytometry, immunofluorescence, and qRT-PCR for inflammatory chemokines. Results B cells in sera from cannulated animals express lower levels of both adiponectin receptors 1 and 2 in comparison with non-inflamed control mice. In cannulated animals treated with PEPITEM, histological analysis of salivary glands revealed fewer, as well as less aggregated, infiltrating T cells. Both CD4 +and CD8+numbers were significantly lower in the salivary glands of PEPITEM-treated animals. Furthermore, administration of PEPITEM also decreased mRNA transcripts for lymphotoxin beta, IL-7, lymphoid chemokines (CCL19 and CXCL13) and T-cell chemokine receptor CCR7, cytokines and chemokines known to regulate ectopic lymphoneogenesis in pSS. Samples from pSS patients are currently being assessed to validate the relevance of this pathway in pSS. Conclusions These results demonstrate that administration of exogenous PEPITEM can reduce T-cell influx into salivary glands. This may represent a rescue of the homeostatic regulation of leukocyte trafficking which is disrupted in inflammation. Our work suggests that PEPITEM should be considered to address therapeutic needs in chronic inflammatory conditions, and that the detection of decreased levels of adiponectin receptors could serve as a biomarker in pSS. References [1] Chimen, McGettrick, et al. Nat Med. 2015;21(5):467–475. [2] Bombardieri, Barone, et al. J. Immunol. 2012;189:3767–3776. Disclosure of Interest None declared

Introduction The RANKL (ligand)-RANK-OPG triad, members of the TNF(R) superfamily, is implicated in lymphoid organ development and bone homeostasis. It has recently been demonstrated that RANK-activated astrocytes release CCL20 and attract T cells to the central nervous system in a model of Multiple Sclerosis and that transgenic RANK expression in the skin promotes aberrant epithelial cell proliferation and is sufficient to induce ectopic formation of tertiary lymphoid structures (TLS). Ductal epithelial cells (SGEs) have been implicated in Sjogren’s Syndrome (SS) pathogenesis where they mediate immune recruitment by expression of pro-inflammatory chemokines and support the formation of pre-malignant myoepithelial lesions. Objectives To address the role of RANK-RANKL interaction in primary (p) SS. Methods A combination of human and mouse studies were used to explore the RANK-RANKL interaction in pSS. Salivary glands (SGs) and saliva samples from patients recruited in the OASIS cohort (University of Birmingham) were studied to evaluate this pathway in human disease. Consecutive stimulated saliva samples (n=69) were analysed using Proseek Multiplex INF96 × 96, covering 92 unique inflammation-related protein biomarkers. Taking advantage of a viral induced model of pSS we studied the effect of this pathway with a RANKL blocking antibody and by inducing gain of function with direct cannulation in the salivary glands of recombinant RANKL. Murine SGs were studied by immunofluorescence, flow cytometry and qPCR on total tissue and sorted cells. Results Fourteen proteins in saliva were significantly separated between pSS and sicca controls, and elevated levels of just two proteins, RANKL and TNFβ, could classify pSS or sicca with 75% accuracy. Levels of salivary RANKL and CCL20 were strongly correlated (r=0.6; p Conclusions In vivo RANK/RANKL interaction mediates recruitment of activated T cells that are skewed toward a Th2 phenotype. These, in turn, will favour the establishment of TLS in the SG. Those data were confirmed in human pSS, where expression of RANK is found in inflamed epithelium and RANKL detection in saliva is able to differentiate patients with pSS from sicca controls, thus candidates this pathway both for drug targeting and patient stratification. Disclosure of interest None declared