Deciphering the role of cDC2s in Sjögren's syndrome: transcriptomic profile links altered antigen processes with IFN signature and autoimmunity.

OBJECTIVE: Type 2 conventional dendritic cells (cDC2s) are key orchestrators of inflammatory responses, linking innate and adaptative immunity. Here we explored the regulation of immunological pathways in cDC2s from patients with primary Sjögren's syndrome (pSS). METHODS: RNA sequencing of circulating cDC2s from patients with pSS, patients with non-Sjögren's sicca and healthy controls (HCs) was exploited to establish transcriptional signatures. Phenotypical and functional validation was performed in independent cohorts. RESULTS: Transcriptome of cDC2s from patients with pSS revealed alterations in type I interferon (IFN), toll-like receptor (TLR), antigen processing and presentation pathways. Phenotypical validation showed increased CX3CR1 expression and decreased integrin beta-2 and plexin-B2 on pSS cDC2s. Functional validation confirmed impaired capacity of pSS cDC2s to degrade antigens and increased antigen uptake, including self-antigens derived from salivary gland epithelial cells. These changes in antigen uptake and degradation were linked to anti-SSA/Ro (SSA) autoantibodies and the presence of type I IFNs. In line with this, in vitro IFN-α priming enhanced the uptake of antigens by HC cDC2s, reflecting the pSS cDC2 profile. Finally, pSS cDC2s compared with HC cDC2s increased the proliferation and the expression of CXCR3 and CXCR5 on proliferating CD4+ T cells. CONCLUSIONS: pSS cDC2s are transcriptionally altered, and the aberrant antigen uptake and processing, including (auto-)antigens, together with increased proliferation of tissue-homing CD4+ T cells, suggest altered antigen presentation by pSS cDC2s. These functional alterations were strongly linked to anti-SSA positivity and the presence of type I IFNs. Thus, we demonstrate novel molecular and functional pieces of evidence for the role of cDC2s in orchestrating immune response in pSS, which may yield novel avenues for treatment.

Cytometry by time of flight identifies distinct signatures in patients with systemic sclerosis, systemic lupus erythematosus and Sjögrens syndrome.

Systemic sclerosis (SSc), systemic lupus erythematosus (SLE) and primary Sjögrens syndrome (pSS) are clinically distinct systemic autoimmune diseases (SADs) that share molecular pathways. We quantified the frequency of circulating immune-cells in 169 patients with these SADs and 44 healty controls (HC) using mass-cytometry and assessed the diagnostic value of these results. Alterations in the frequency of immune-cell subsets were present in all SADs compared to HC. Most alterations, including a decrease of CD56hi NK-cells in SSc and IgM+ Bcells in pSS, were disease specific; only a reduced frequency of plasmacytoid dendritic cells was common between all SADs Strikingly, hierarchical clustering of SSc patients identified 4 clusters associated with different clinical phenotypes, and 9 of the 12 cell subset-alterations in SSc were also present during the preclinical-phase of the disease. Additionally, we found a strong association between the use of prednisone and alterations in B-cell subsets. Although differences in immune-cell frequencies between these SADs are apparent, the discriminative value thereof is too low for diagnostic purposes. Within each disease, mass cytometry analyses revealed distinct patterns between endophenotypes. Given the lack of tools enabling early diagnosis of SSc, our results justify further research into the value of cellular phenotyping as a diagnostic aid.

Epigenetically quantified immune cells in salivary glands of Sjögren's syndrome patients: a novel tool that detects robust correlations of T follicular helper cells with immunopathology.

OBJECTIVE: To investigate whether epigenetic cell counting represents a novel method to quantify immune cells in salivary glands of patients with different forms of Sjögren's and sicca syndrome and to capture immunopathology and potentially aid in diagnosis. METHODS: DNA from frozen salivary gland tissue sections of sicca patients was used for bisulphite conversion of demethylated DNA cytosine residues, followed by cell-specific quantitative PCR to calculate cell percentages in relation to total tissue cell numbers as quantified by housekeeping gene demethylation. The percentages of epigenetically quantified cells were correlated to RNA expression of matched salivary gland tissue and histological and clinical parameters. RESULTS: The percentages of epigenetically quantified CD3, CD4, CD8, T follicular helper (Tfh) cells, FoxP3+ regulatory T cells and B cells were significantly increased in the salivary glands of patients with SS. Unsupervised clustering using these percentages identified patient subsets with an increased lymphocytic focus score and local B cell hyperactivity and classifies patients different from conventional classification criteria. In particular, Tfh cells were shown to strongly correlate with the expression of CXCL13, lymphocytic focus scores, local B cell hyperactivity and anti-SSA positivity. CONCLUSION: Epigenetic cell counting is a promising novel tool to objectively and easily quantify immune cells in the labial salivary gland of sicca patients, with a relatively small amount of tissue needed. In view of the potential of this technique to include a huge number of (cell-specific) biomarkers, this opens up new standardized ways of salivary gland analysis with high relevance for patient classification, understanding of immunopathology and monitoring of drug responses in clinical trials.

Dysregulated miRNome of plasmacytoid dendritic cells from patients with Sjögren's syndrome is associated with processes at the centre of their function.

OBJECTIVE: A considerable body of evidence supports a role for type-I IFN in the pathogenesis of primary SS (pSS). As plasmacytoid dendritic cells (pDCs) are a major source of type-I IFN, we investigated their molecular regulation by measuring expression of a large set of miRNAs. METHODS: pDCs were isolated from peripheral blood of pSS patients (n = 30) and healthy controls (n = 16) divided into two independent cohorts (discovery and replication). Screening of 758 miRNAs was assessed by an OpenArray quantitative PCR-based technique; replication of a set of identified miRNAs was performed by custom array. Functional annotation of miRNA targets was performed using pathway enrichment. Novel targets of miR-29a and miR-29c were identified using a proteomic approach (stable isotope labelling with amino acids in cell culture). RESULTS: In the discovery cohort, 20 miRNAs were differentially expressed in pSS pDCs compared with healthy control pDCs. Of these, differential expression of 10 miRNAs was confirmed in the replication cohort. The dysregulated miRNAs were involved in phosphoinositide 3-kinase-Ak strain transforming and mammalian target of rapamycin signalling, as well as regulation of cell death. In addition, a set of novel protein targets of miR-29a and miR-29c were identified, including five targets that were regulated by both miRs. CONCLUSION: The dysregulated miRNome in pDCs of patients with pSS is associated with aberrant regulation of processes at the centre of pDC function, including type-I IFN production and cell death. As miR-29a and miR-29c are pro-apoptotic factors and several of the novel targets identified here are regulators of apoptosis, their downregulation in patients with pSS is associated with enhanced pDC survival.

Serum microRNA screening and functional studies reveal miR-483-5p as a potential driver of fibrosis in systemic sclerosis.

OBJECTIVE: MicroRNAs (miRNAs) are regulatory molecules, which have been addressed as potential biomarkers and therapeutic targets in rheumatic diseases. Here, we investigated the miRNA signature in the serum of systemic sclerosis (SSc) patients and we further assessed their expression in early stages of the disease. METHODS: The levels of 758 miRNAs were evaluated in the serum of 26 SSc patients as compared to 9 healthy controls by using an Openarray platform. Three miRNAs were examined in an additional cohort of 107 SSc patients and 24 healthy donors by single qPCR. MiR-483-5p expression was further analysed in the serum of patients with localized scleroderma (LoS) (n = 22), systemic lupus erythematosus (SLE) (n = 33) and primary Sjögren's syndrome (pSS) (n = 23). The function of miR-483-5p was examined by transfecting miR-483-5p into primary human dermal fibroblasts and pulmonary endothelial cells. RESULTS: 30 miRNAs were significantly increased in patients with SSc. Of these, miR-483-5p showed reproducibly higher levels in an independent SSc cohort and was also elevated in patients with preclinical-SSc symptoms (early SSc). Notably, miR-483-5p was not differentially expressed in patients with SLE or pSS, whereas it was up-regulated in LoS, indicating that this miRNA could be involved in the development of skin fibrosis. Consistently, miR-483-5p overexpression in fibroblasts and endothelial cells modulated the expression of fibrosis-related genes. CONCLUSIONS: Our findings showed that miR-483-5p is up-regulated in the serum of SSc patients, from the early stages of the disease onwards, and indicated its potential function as a fine regulator of fibrosis in SSc.

Decreased expression of thymic stromal lymphopoietin in salivary glands of patients with primary Sjögren's syndrome is associated with increased disease activity.

OBJECTIVES: Thymic Stromal Lymphopoietin (TSLP) is a potent immunomodulatory cytokine involved in Th2- and Th17-mediated immune responses in different autoimmune diseases. TSLP expression in relation to disease activity was studied in salivary glands of primary Sjögren's syndrome (pSS) patients as compared to non-SS sicca (nSS) controls. METHODS: Tissue sections of minor salivary glands from pSS and nSS patients were stained with monoclonal antibodies against human TSLP, CD3, CD19 and cytokeratin high molecular weight (CK HMW) or stained for Alcian blue to detect mucus production. The number of TSLP-expressing cells was quantified and expression was correlated to local and systemic disease parameters. RESULTS: The number of TSLP-expressing cells was significantly lower in pSS patients than in nSS controls and correlated with a range of disease markers. In pSS patients, TSLP was expressed outside of lymphocytic infiltrates at sections that also encompassed high numbers of intact acinar cells. This difference was independent of tissue destruction. CONCLUSIONS: Reduced TSLP expression in pSS patients is associated with increased local and systemic inflammatory markers. Loss of TSLP expression may contribute to Th1/Th17-associated immunopathology in pSS, in line with previous studies demonstrating that TSLP promotes a protective Th2 milieu at mucosal sites.

Thymic stromal lymphopoietin as a novel mediator amplifying immunopathology in rheumatic disease.

Thymic stromal lymphopoietin (TSLP) is an IL-7-related cytokine that has been studied extensively in atopic diseases and more recently in various rheumatic disorders. It is involved in T cell development in the thymus and promotes homeostatic T cell expansion by classical dendritic cells. However, deregulated TSLP expression in various rheumatic diseases has implicated this cytokine as a strong mediator in immunopathology. Overexpressed TSLP induces strong T cell activation and production of pro-inflammatory cytokines in human cells and animal models for RA, SSc and LN, underscoring the therapeutic potential of targeting the TSLP-TSLP receptor axis.

Leflunomide/hydroxychloroquine combination therapy targets type I IFN-associated proteins in patients with Sjögren's syndrome that show potential to predict and monitor clinical response.

OBJECTIVES: To assess to what extent leflunomide (LEF) and hydroxychloroquine (HCQ) therapy in patients with primary Sjögren's syndrome (RepurpSS-I) targets type I IFN-associated responses and to study the potential of several interferon associated RNA-based and protein-based biomarkers to predict and monitor treatment. METHODS: In 21 patients treated with LEF/HCQ and 8 patients treated with placebo, blood was drawn at baseline, 8, 16 and 24 weeks. IFN-signatures based on RNA expression of five IFN-associated genes were quantified in circulating mononuclear cells and in whole blood. MxA protein levels were measured in whole blood, and protein levels of CXCL10 and Galectin-9 were quantified in serum. Differences between responders and non-responders were assessed and receiver operating characteristic analysis was used to determine the capacity of baseline expression and early changes (after 8 weeks of treatment) in biomarkers to predict treatment response at the clinical endpoint. RESULTS: IFN-signatures in peripheral blood mononuclear cell and whole blood decreased after 24 weeks of LEF/HCQ treatment, however, changes in IFN signatures only poorly correlated with changes in disease activity. In contrast to baseline IFN signatures, baseline protein concentrations of galectin-9 and decreases in circulating MxA and Galectin-9 were robustly associated with clinical response. Early changes in serum Galectin-9 best predicted clinical response at 24 weeks (area under the curve 0.90). CONCLUSIONS: LEF/HCQ combination therapy targets type-I IFN-associated proteins that are associated with strongly decreased B cell hyperactivity and disease activity. IFN-associated Galectin-9 is a promising biomarker for treatment prediction and monitoring in pSS patients treated with LEF/HCQ.

The Transcriptomic Profile of Monocytes from Patients With Sjögren's Syndrome Is Associated With Inflammatory Parameters and Is Mimicked by Circulating Mediators.

Primary Sjögren's syndrome (pSS) is a systemic autoimmune disease characterized by infiltration of the exocrine glands and prominent B cell hyperactivity. Considering the key role of monocytes in promoting B cell hyperactivity, we performed RNA-sequencing analysis of CD14+ monocytes from patients with pSS, non-Sjögren's sicca (nSS), and healthy controls (HC). We demonstrated that the transcriptomic profile of pSS patients is enriched in intermediate and non-classical monocyte profiles, and confirmed the increased frequency of non-classical monocytes in pSS patients by flow-cytometry analysis. Weighted gene co-expression network analysis identified four molecular signatures in monocytes from pSS patients, functionally annotated for processes related with translation, IFN-signaling, and toll-like receptor signaling. Systemic and local inflammatory features significantly correlated with the expression of these signatures. Furthermore, genes highly associated with clinical features in pSS were identified as hub-genes for each signature. Unsupervised hierarchical cluster analysis of the hub-genes identified four clusters of nSS and pSS patients, each with distinct inflammatory and transcriptomic profiles. One cluster showed a significantly higher percentage of pSS patients with higher prevalence of anti-SSA autoantibodies, interferon-score, and erythrocyte sedimentation rate compared to the other clusters. Finally, we showed that the identified transcriptomic differences in pSS monocytes were induced in monocytes of healthy controls by exposure to serum of pSS patients. Representative hub-genes of all four signatures were partially inhibited by interferon-α/ß receptor blockade, indicating that the circulating inflammatory mediators, including type I interferons have a significant contribution to the altered transcriptional profile of pSS-monocytes. Our study suggests that targeting key circulating inflammatory mediators, such as type I interferons, could offer new insights into the important pathways and mechanisms driving pSS, and holds promise for halting immunopathology in Sjögren's Syndrome.

Implication of miR-126 and miR-139-5p in Plasmacytoid Dendritic Cell Dysregulation in Systemic Sclerosis.

Compelling evidence shows the involvement of plasmacytoid dendritic cells (pDCs) in systemic sclerosis (SSc) pathogenesis. This study investigated whether microRNAs (miRNAs) are involved in the dysregulation of pDCs in SSc patients already at early stages. RNA from circulating pDCs was isolated from two independent cohorts of SSc patients with different disease phenotypes, and individuals with Raynaud's phenomenon, for microRNA profiling and RNA-sequencing analysis. Proteomic analysis was exploited to identify novel direct miRNA targets at the protein level. Twelve and fifteen miRNAs were differentially expressed in at least one group of patients compared to healthy controls in discovery cohort I and II, respectively. Of note, miR-126 and miR-139-5p were upregulated in both preclinical and definite SSc patients and correlated with the expression of type I interferon (IFN)-responsive genes. Toll-like receptor 9 (TLR9) stimulation of healthy pDCs upregulated the expression of both miRNAs, similarly to what was observed in patients. The proteomic analysis identified USP24 as a novel target of miR-139-5p. The expression level of USP24 was inversely correlated with miR-139-5p expression in SSc patients and induced by TLR9 stimulation in healthy pDCs. These findings demonstrated that the miRNA profile is altered in pDCs of SSc patients already at early stages of the disease and indicate their potential contribution to pDC activation observed in patients.

Plasmacytoid DCs From Patients With Sjögren's Syndrome Are Transcriptionally Primed for Enhanced Pro-inflammatory Cytokine Production.

Primary Sjögren's syndrome (pSS) is a systemic auto-immune disease typified by dryness of the mouth and eyes. A majority of patients with pSS have a type-I interferon (IFN)-signature, which is defined as the increased expression of IFN-induced genes in circulating immune cells and is associated with increased disease activity. As plasmacytoid dendritic cells (pDC) are the premier type-I IFN-producing cells and are present at the site of inflammation, they are thought to play a significant role in pSS pathogenesis. Considering the lack of data on pDC regulation and function in pSS patients, we here provided the first in-depth molecular characterization of pSS pDCs. In addition, a group of patients with non-Sjögren's sicca (nSS) was included; these poorly studied patients suffer from complaints similar to pSS patients, but are not diagnosed with Sjögren's syndrome. We isolated circulating pDCs from two independent cohorts of patients and controls (each n = 31) and performed RNA-sequencing, after which data-driven networks and modular analysis were used to identify robustly reproducible transcriptional "signatures" of differential and co-expressed genes. Four signatures were identified, including an IFN-induced gene signature and a ribosomal protein gene-signature, that indicated pDC activation. Comparison with a dataset of in vitro activated pDCs showed that pSS pDCs have higher expression of many genes also upregulated upon pDC activation. Corroborating this transcriptional profile, pSS pDCs produced higher levels of pro-inflammatory cytokines, including type-I IFN, upon in vitro stimulation with endosomal Toll-like receptor ligands. In this setting, cytokine production was associated with expression of hub-genes from the IFN-induced and ribosomal protein gene-signatures, indicating that the transcriptional profile of pSS pDCs underlies their enhanced cytokine production. In all transcriptional analyses, nSS patients formed an intermediate group in which some patients were molecularly similar to pSS patients. Furthermore, we used the identified transcriptional signatures to develop a discriminative classifier for molecular stratification of patients with sicca. Altogether, our data provide in-depth characterization of the aberrant regulation of pDCs from patients with nSS and pSS and substantiate their perceived role in the immunopathology of pSS, supporting studies that target pDCs, type-I IFNs, or IFN-signaling in pSS.

Salivary gland secretome: a novel tool towards molecular stratification of patients with primary Sjögren's syndrome and non-autoimmune sicca.

Objective: To explore the potential of salivary gland biopsy supernatants (the secretome) as a novel tool to aid in stratification of patients with sicca syndrome and to study local immunopathology in Sjögren's syndrome. Methods: Labial salivary gland biopsies were incubated in saline for 1 hour. In these tissue supernatants from a discovery cohort (n=16) of patients with primary Sjögren's syndrome (pSS) and non-Sjögren's sicca (nSS), 101 inflammatory mediators were measured by Luminex. Results were validated in a replication cohort (n=57) encompassing patients with pSS, incomplete SS and nSS. Results: The levels of 23 cytokines were significantly increased in patients with pSS versus nSS in the discovery cohort. These 23 and 3 additional cytokines were measured in a second cohort. Elevated concentrations of 11 cytokines were validated and the majority correlated with clinical parameters. Classification tree analysis indicated that the concentrations of CXCL13, IL-21, sIL-2R and sIL-7Rα could be used to classify 95.8% of patients with pSS correctly. Conclusion: Labial salivary gland secretomes can be used to reliably assess mediators involved in immunopathology of patients with pSS, potentially contributing to patient classification. As such, this method represents a novel tool to identify therapeutic targets and markers for diagnosis, prognosis and treatment response.

MicroRNA-130a Contributes to Type-2 Classical DC-activation in Sjögren's Syndrome by Targeting Mitogen- and Stress-Activated Protein Kinase-1.

Objectives: Considering the critical role of microRNAs (miRNAs) in regulation of cell activation, we investigated their role in circulating type-2 conventional dendritic cells (cDC2s) of patients with primary Sjögren's syndrome (pSS) compared to healthy controls (HC). Methods: CD1c-expressing cDC2s were isolated from peripheral blood. A discovery cohort (15 pSS, 6 HC) was used to screen the expression of 758 miRNAs and a replication cohort (15 pSS, 11 HC) was used to confirm differential expression of 18 identified targets. Novel targets for two replicated miRNAs were identified by SILAC in HEK-293T cells and validated in primary cDC2s. Differences in cytokine production between pSS and HC cDC2s were evaluated by intracellular flow-cytometry. cDC2s were cultured in the presence of MSK1-inhibitors to investigate their effect on cytokine production. Results: Expression of miR-130a and miR-708 was significantly decreased in cDC2s from pSS patients compared to HC in both cohorts, and both miRNAs were downregulated upon stimulation via endosomal TLRs. Upstream mediator of cytokine production MSK1 was identified as a novel target of miR-130a and overexpression of miR-130a reduced MSK1 expression in cDC2s. pSS cDC2s showed higher MSK1 expression and an increased fraction of IL-12 and TNF-α-producing cells. MSK1-inhibition reduced cDC2 activation and production of IL-12, TNF-α, and IL-6. Conclusions: The decreased expression of miR-130a and miR-708 in pSS cDC2s seems to reflect cell activation. miR-130a targets MSK1, which regulates pro-inflammatory cytokine production, and we provide proof-of-concept for MSK1-inhibition as a therapeutic avenue to impede cDC2 activity in pSS.

Circulating small non-coding RNAs reflect IFN status and B cell hyperactivity in patients with primary Sjögren's syndrome.

BACKGROUND: Considering the important role of miRNAs in the regulation of post-transcriptional expression of target genes, we investigated circulating small non-coding RNAs (snc)RNA levels in patients with primary Sjögren's syndrome (pSS). In addition we assessed if serum sncRNA levels can be used to differentiate patients with specific disease features. METHODS: Serum RNA was isolated from 37 pSS patients as well as 21 patients with incomplete Sjögren's Syndrome (iSS) and 17 healthy controls (HC) allocated to two independent cohorts: discovery and validation. OpenArray profiling of 758 sncRNAs was performed in the discovery cohort. Selected sncRNAs were measured in the validation cohort using single-assay RT-qPCR. In addition, unsupervised hierarchical clustering was performed within the pSS group. RESULTS: Ten sncRNAs were differentially expressed between the groups in the array. In the validation cohort, we confirmed the increased expression of U6-snRNA and miR-661 in the iSS group as compared to HC. We were unable to validate differential expression of any miRNAs in the pSS group. However, within this group several miRNAs correlated with laboratory parameters. Unsupervised clustering distinguished three clusters of pSS patients. Patients in one cluster showed significantly higher serum IgG, prevalence of anti-SSB autoantibodies, IFN-score, and decreased leukocyte counts compared to the two other clusters. CONCLUSION: We were unable to identify any serum sncRNAs with differential expression in pSS patients. However, we show that circulating miRNA levels are associated with disease parameters in pSS patients and can be used to distinguish pSS patients with more severe B cell hyperactivity. As several of these miRNAs are implicated in the regulation of B cells, they may play a role in the perpetuation of the disease.

Increased CCL25 and T Helper Cells Expressing CCR9 in the Salivary Glands of Patients With Primary Sjögren's Syndrome: Potential New Axis in Lymphoid Neogenesis.

OBJECTIVE: Follicular helper T (Tfh) cells play a critical role in germinal center formation and B cell activation, both of which are hallmarks of primary Sjögren's syndrome (SS). CCR9-expressing T helper cells have "Tfh-like" characteristics and their numbers are increased at mucosa-associated sites in several inflammatory conditions. Because the characteristics of these cells are unique and evaluation has been limited, this study was undertaken to investigate the local and systemic CCL25/CCR9 axis in patients with primary SS. METHODS: Levels of CCL25 protein and messenger RNA (mRNA) and CCR9+ T helper cells were evaluated in the labial salivary glands (LSGs) of patients with primary SS and patients with sicca syndrome without a diagnosis of primary SS (non-SS sicca controls). CCL25 levels were assessed for correlation with parameters of inflammation and clinical features. Circulating CCR9+ and CXCR5+ T helper cells were compared on the basis of phenotypic and functional properties. RESULTS: CCL25 protein and mRNA levels were elevated in the LSGs of patients with primary SS as compared to non-SS sicca controls. Increased levels of CCL25 were associated with B cell hyperactivity, autoimmunity, and levels of interleukin-21 (IL-21) and soluble IL-7 receptor α-chain (IL-7Rα). Furthermore, the frequency of CCR9-expressing cells in the LSGs was increased and levels of circulating CCR9+ T helper cells expressing programmed death 1 and inducible T cell costimulator were elevated in patients with primary SS. CCR9+ T helper cells displayed higher expression of IL-7Rα and secreted higher levels of interferon-γ, IL-17, IL-4, and IL-21 as compared to CXCR5+ T helper cells, ex vivo and upon triggering with antigen or IL-7. Both CCR9+ and CXCR5+ T helper cells induced IgG production by B cells more potently than that induced in the cultures with CCR9-CXCR5- T helper cells. CONCLUSION: Enhanced expression of CCL25 in LSGs of patients with primary SS can facilitate attraction of CCR9+ T helper cells, and these cells secrete high levels of proinflammatory cytokines when triggered with antigen or IL-7. The observed associations with B cell hyperactivity, autoimmunity, and markers of lymphoid neogenesis indicate that the CCL25/CCR9 axis plays a significant role in the immunopathology of primary SS, suggesting that this axis could represent a novel therapeutic target for the disease.

Salivary Gland Pathology in Sjögren's Syndrome.

Primary Sjögren syndrome (pSS) can be considered a systemic autoimmune disease with a strong organ bias. The involvement of the exocrine glands is prevalent and drives the pathognomonic manifestations of dryness that define the sicca syndrome. The salivary glands also represent the hub of pSS pathology. Elements belonging to both innate and acquired immune responses have been described at this site that contribute to disease establishment and progression. The interaction between those elements and their relative contributions to the clinical manifestations and lymphoma progression largely remain to be addressed.