Sjögren's Disease (SjD) presents a challenge as a chronic, progressive, and systemic autoimmune disorder affecting approximately 0.05% of the population. SjD primarily targets exocrine glands, leading to severe dryness of the mouth and eyes, but can impact multiple organs and increase lymphoma risk. Despite advances in B cell-targeted therapies, no cure exists, and treatment remains symptom-focused, highlighting gaps in our understanding of SjD inflammatory environment. One roadblock to study the heterogeneity of the landscape in affected salivary glands in humans is the limited amount of tissue that is typically available from biopsies of minor salivary glands (MSGs). Spatial transcriptomics has emerged as a cutting-edge approach to address this challenge by preserving tissue architecture and enabling precise mapping of immune and structural cell interactions. This chapter explores how the Visium HD protocol from 10X Genomics can be leveraged to dissect the cellular and molecular landscape of formalin-fixed paraffin-embedded human MSGs, uncovering insights into disease mechanisms and therapeutic opportunities.

Sjögren’s disease (SjD) is a chronic autoimmune condition marked by lymphocytic infiltration of exocrine glands and production of autoantibodies such as anti-SSA/Ro, anti-SSB/La, and rheumatoid factor. B lymphocytes play a central role in disease pathogenesis, driving autoantibody production and glandular damage, and contributing to lymphomagenesis. Despite promising therapies, no effective treatment is currently available, partly due to the biological and clinical heterogeneity of the disease. While interferon (IFN) signatures and B cell–related markers are used for patient stratification, their integration remains unexplored. This study analyzed B cell transcriptional and metabolic profiles using bulk transcriptomic, clinical, and flow cytometry data from the PRECISESADS consortium, alongside public single-cell RNA-sequencing datasets. A B cell–specific IFN-α signature was established to stratify patients into IFN-positive and IFN-negative groups. Both showed reduced oxidative phosphorylation (OXPHOS) and translation in B cell subsets, suggesting a shared pre-metabolic state. IFN-positive patients, however, displayed additional features, including enhanced glycolysis, amino acid and lipid metabolism, autophagy, and NF-κB signaling. They also showed an expansion of IFN-activated naïve (Naive IFN), Transitional, and double-negative (DN) B cells, particularly DN2 and DN2_CXCR3 subsets, which have been linked in the literature to autoreactivity and lymphoma development. The IFN signature in naïve B cells and DN2 correlated with elevated anti-SSA/Ro and anti-SSB/La titers, while only naïve B cells showed an association with increased histological focus scores. These findings support the relevance of a B cell–specific IFN signature in stratifying SjD patients and suggest new metabolic and transcriptional targets for disease monitoring and therapeutic development. ![Figure][1] ### Competing Interest Statement The authors have declared no competing interest. The research leading to these results has received support from the Innovative Medicines Initiative Joint Undertaking under Grant Agreement Number 115565, resources of which are composed of financial contributions from the European Union’s Seventh Framework Program (FP7/2007–2013) and EFPIA companies in-kind contributions. We would like to acknowledge 3TR and the PRECISESADS consortium, which guaranteed the availability of the data used in this article and the opportunity to analyse them. CI was funded by the Université de Brest and the Région Bretagne., 115565 [1]: pending:yes

Interpreting biological system changes requires interpreting vast amounts of multi-omics data. While user-friendly tools exist for single-omics analysis, integrating multiple omics still requires bioinformatics expertise, limiting accessibility for the broader scientific community. BiomiX tackles the bottleneck in high-throughput omics data analysis, enabling efficient and integrated analysis of multiomics data obtained from two cohorts. BiomiX incorporates diverse omics data, using DESeq2/Limma packages for transcriptomics, and quantifying metabolomics peak differences, evaluated via the Wilcoxon test with the False Discovery Rate correction. The metabolomics annotation for Liquid Chromatography-Mass Spectrometry untargeted metabolomics is additionally supported using the mass-to-charge ratio in the CEU Mass Mediator database and fragmentation spectra in the TidyMass package. Methylomics analysis is performed using the ChAMP R package. Finally, Multi-Omics Factor Analysis (MOFA) integration identifies shared sources of variation across omics data. BiomiX also generates statistics, report figures and integrates EnrichR and GSEA for biological process exploration and subgroup analysis based on user-defined gene panels enhancing condition subtyping. BiomiX fine-tunes MOFA models, to optimize factors number selection, distinguishing between cohorts and providing tools to interpret discriminative MOFA factors. The interpretation relies on innovative bibliography research on Pubmed, which provides the articles most related to the discriminant factor contributors. Furthermore, discriminant MOFA factors are correlated with clinical data, and the top contributing pathways are explored, all with the aim of guiding the user in factor interpretation. The analysis of single-omics and multi-omics integration in a standalone tool, along with MOFA implementation and its interpretability via literature, represents significant progress in the multi-omics field in line with the “Findable, Accessible, Interoperable, and Reusable” data principles. BiomiX offers a wide range of parameters and interactive data visualization, allowing for personalized analysis tailored to user needs. This R-based, user-friendly tool is compatible with multiple operating systems and aims to make multi-omics analysis accessible to non-experts in bioinformatics.

Objective: Sjogren's disease (SjD) is a systemic autoimmune disorder characterized by lymphocytic infiltration of exocrine glands, resulting in xerostomia, keratoconjunctivitis sicca, fatigue, arthralgia, and systemic organ involvement. This study aimed to characterize the metabolic and immune dysregulation of SjD using a multi-omics approach, focusing on the metabolic environment and B-cell transcriptomic responses. Methods: Transcriptomic, methylomic, and metabolomic datasets from whole blood, plasma, and urine of 293 SjD patients and 508 controls were analyzed from the PRECISESADS study. B-cell transcriptomes were included to link systemic metabolic alterations to cell-intrinsic immune programs. Multi-omics factor analysis (MOFA) was used to integrate data and identify discriminant molecular drivers. Results: Multi-omics integration revealed metabolic rewiring involving the urea cycle, glutamine/arginine metabolism, and NAD⁺ depletion linked to interferon signaling. Among the strongest contributors, plasma lysophosphatidic acids (LPA) emerged as key discriminants associated with interferon-driven activation. B-cell transcriptomes showed upregulation of LPA-related genes (CERS6, INPP1, TRIP6), and its receptor LPAR6. Importantly, in this study LPAR6 protein expression was confirmed in B cells for the first time. Secondary findings included alterations in sphingosine-1-phosphate (S1P) metabolism, suggesting a broader lysophospholipid signaling axis. Conclusions: This study identifies the LPA-LPAR6 signaling axis as a potential metabolic driver of B-cell activation and interferon-associated inflammation in SjD, highlighting a previously unrecognized immunometabolic pathway. These findings highlight LPA-LPAR6 as a candidate target for therapeutic modulation in SjD, while also implicating S1P signaling as a complementary regulatory mechanism. ### Competing Interest Statement The authors have declared no competing interest. Innovative Medicines Initiative Joint Undertaking, 115565

OBJECTIVE:To assess the prevalence of giant cell arteritis (GCA) or cancer detected via 18F-fluorodeoxyglucose positron emission tomography-computed tomography (PET-CT) in patients with polymyalgia rheumatica (PMR) without clinical signs suggestive of GCA. METHOD:This monocentric retrospective cohort study analyzed PET-CT scans performed for clinical suspicion of PMR since 2018. Patients meeting the 2012 ACR/EULAR criteria for PMR without clinical indications of GCA or cancer were included. Observations were divided into two groups: new-onset PMR and treatment failure. A nuclear medicine physician evaluated qualitative liver uptake scores at articular/periarticular sites and large vessels typically involved in PMR. Increased 18F-FDG uptake suggesting cancer was also assessed. RESULTS:Of 1223 PET-CT scans screened, 94 met the inclusion criteria: 38 for new-onset PMR and 56 for treatment failure. Subclinical GCA was identified in 10 (10.6%) patients, with a prevalence of 5.3% in the new-onset group and 14.2% in the treatment failure group. Aortic uptake was present in 90% of subclinical GCA cases. Sites with increased 18F-FDG uptake included hips (90%), shoulders, lumbar interspinous bursa (80%), and ischial tuberosity (60%). PET-CT identified eight (8.5%) cancer cases, equally distributed between the two groups. CONCLUSION:PET-CT detects subclinical GCA in approximately 10% of PMR patients without suggestive symptoms, three times more frequently in the treatment failure group. Aortitis is present in 90% of subclinical GCA cases. Cancer prevalence is 4.7%, with a heterogeneous spectrum and unclear association with PMR symptoms.

Background Autoreactive proteinase 3 (PR3+) B cells have recently been phenotypically and functionally characterized, and the presence of defective central antigen-independent and peripheral antigen-dependent checkpoints in patients with ANCA-associated vasculitis (AAV) has been shown. This work aimed to investigate the central tolerance-checkpoint controlling immature PR3+ B cells in the bone marrow (BM), before their migration into the periphery as transitional B cells. Objectives We investigated the presence and the specific phenotypic features of PR3+ B cells in BM mononuclear cells (BMMC) of non-vasculitis controls (No-AAV), comparing them to paired peripheral blood mononuclear cells (PBMC) of No-AAV and PBMC of PR3-AAV patients, and the central tolerance-checkpoint for PR3+ B cells. Methods We used a customized flow-cytometry assay, using PR3 as ligand to target autoreactive PR3+ B cells (PR3+B cells). Adult PR3-ANCA positive AAV (PR3-AAV) patients with a clinical diagnosis of granulomatosis with polyangiitis (GPA) or microscopic polyangiitis (MPA) were selected among consecutive subjects with AAV seen in our Rheumatology Unit. Subjects without vasculitis (No AAV) were selected among consecutive subjects undergoing bone marrow aspirate to exclude hematologic conditions of myeloid origin and eventually resulted healthy or in long-term complete remission during follow-up of myeloid neoplasms. PMBC from AAV patients and paired samples of BMMC and PBMC from No AAV were collected and analyzed. Results The proportion of PR3+ B cells within BMMC (median [IQR25-75%]; 1.98%[1.77-2.75]) was higher than within PBMC of No-AAV (0.9%[0.63-1.44], p<0.01 by paired comparison) and similar to their proportion within PBMC of PR3-AAV patients (1.82%[1.66-3.21]; p>0.05). When focusing on immature/transitional CD24++CD38++B cells only in No-AAV, we observed distinct phenotypes within BMMC versus PBMC (i.e. higher proportion of CD27-CD10+ and lower expression of CD21, IgD, IgM within BMMC versus PBMC), representing two separate developmental steps of B cell maturation. Within CD24++CD38++ B cells, BMMC contained the greatest proportion of PR3+ B cells as compared to PBMC (3.35%[1.99-4.92] versus 1.23%[0.62-1.55], p<0.01). We observed a significant decline of the PR3+ fraction from T1-like/immature subset (IgD-IgM+; 2.80%[1.23-4.02]) to T2-like/early transitional subset (IgD+IgM+; 1.76%[0.96-2.68], p<0.01) in BMMC, while no significant reduction was observed between the latter subset and the transitional compartment of PBMC (1.26%[0.62-1.56], p>0.05). Conclusion To prevent PR3-related autoimmunity, autoreactive PR3+ B cells pass a stringent selection in the BM, and their removal by central tolerance-checkpoint activity occurs mainly between T1-like/immature to T2-like/early transitional B cells of BMMC. References [1]Cornec D. Identification and phenotyping of circulating autoreactive proteinase 3-specific B cells in patients with PR3-ANCA associated vasculitis and healthy controls. J of Autoimmunity, 2017. [2]Berti A. Circulating autoreactive proteinase 3+ B cells and tolerance checkpoints in ANCA-associated vasculitis. JCI Insight, 2021. Acknowledgements: NIL. Disclosure of Interests Alvise Berti Speakers bureau: GSK, Michele Tomasi: None declared, Isabella Pesce: None declared, Enrico Lista: None declared, Anna Guella: None declared, Giuseppe Paolazzi: None declared, Roberto Bortolotti: None declared, Guido Grandi: None declared, Sophie Hillion: None declared, Ulrich Specks: None declared, Divi Cornec: None declared.

Evaluate the benefit of 2-deoxy-2-[18F]-fluoro-D-glucose ([18F] FDG) positron emission tomography/computed tomography (PET/CT) for the therapeutic assessment of Abatacept (ABA) as first-line therapy in early-onset polymyalgia rheumatica (PMR) patients. This was an ancillary study of ALORS trial (Abatacept in earLy Onset polymyalgia Rheumatica Study) assessing the ability of ABA versus placebo to achieve low disease activity (C-Reactive Protein PMR activity score (CRP PMR-AS) ≤ to 10) without glucocorticoid (GC) at week 12 in patients with early-onset PMR. The patients underwent [18F] FDG PET/CT at baseline and after 12 weeks of treatment. Responses to treatments were evaluated according to CRP PMR-AS, Erythrocyte Sedimentation Rate (ESR) PMR-AS, Clin PMR-AS, and CRP-Imputed (Imput-CRP) PMR-AS. Quantitative score by maximal standardized uptake value (SUVmax) and combined qualitative scores according to liver uptake (Leuven, Leuven/Groningen, and Besançon Scores) were used for assessment of [18F] FDG uptake in regions of interest (ROI) usually affected in PMR. Student’s t-test was applied to evaluate the clinical, biological, and [18F] FDG uptake variation difference in ABA and placebo groups between W0 and W12. Subgroup analysis by GC rescue was performed. At W12, there was no significant difference according to SUVmax between the ABA and the placebo groups in all ROI. Subgroup analysis according to GC administration demonstrated a significant (p 0.047) decrease in SUVmax within the left sternoclavicular joint ROI in the ABA group (− 0.8) compared to the placebo group (+ 0.6) without GC rescue. Other results did not reveal any significant difference between the ABA and placebo groups. According to combined qualitative scores, there was no significant difference between ABA and placebo groups for the direct comparison analysis and subgroup analysis according to GC rescue. [18F] FDG PET/CT uptake did not decrease significantly after ABA compared to placebo in anatomical areas usually affected in PMR patients. These results are correlated with the clinical-biological therapeutic assessment. The study was approved by the appropriate ethics committee (CPP Sud-Est II Ref CPP: 2018–33), and all patients gave their written informed consent before study enrollment. The protocol was registered on Clinicaltrials.gov (NCT03632187).

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BACKGROUND:Pemetrexed and gemcitabine are both antimetabolites drugs approved in advanced non-small cell lung cancer (NSCLC). Their toxicity profile is well known. However, rare vascular side effects can occur such as vascular acrosyndromes and especially digital ischemia. The cause of this disfiguring and painful event is still controversial. Amputation is frequently required and has been described as a predictor of poor survival outcomes.CASE PRESENTATION:This report presents two cases of vascular acrosyndrome in NSCLC patients during treatment with antimetabolites (pemetrexed and gemcitabine). Patients presented severe digital ischemia having required prostacyclin analog and chemotherapy discontinuation. In one case, symptoms improved while in the other case symptoms persisted. Both patients experienced prolonged tumor response. These findings suggest a multifactorial mechanism behind digital necrosis including an autoimmune process, which could lead to prolonged tumor control as described with immune checkpoint inhibitors.CONCLUSION:Severe vascular acrosyndrome such as digital ischemia can occur in lung cancer patients treated with antimetabolites. Awareness needs to be raised when using these drugs in patients with predisposing factors. Whether occurrence of chemotherapy-induced immune vascular side effects might explain prolonged tumor response deserves further investigations.

We read with interest the study by Posada et al. (1) evaluating the therapeutic effects of RSLV-132 (a new RNase compound) in patients with primary Sjogren syndrome (pSS). At first glance, one may be surprised by the enthusiastic conclusion of authors in spite of a biological effect opposite to what was expected, namely an increased expression of interferon (IFN)-stimulated genes (ISG), while pSS is considered as an acquired interferonopathy. However, therapeutic trials are also a unique opportunity to learn about the pathogenesis of such complex condition. Indeed, this study showed a significant improvement of fatigue (measured by various validated scales) in treated patients, surprisingly correlated with the increased expression of ISG.

Introduction: Because of their efficacy against numerous cancers, immune-checkpoint inhibitors (ICIs), anti-cytotoxic T-lymphocyte antigen-4, and anti-programmed cell death monoclonal antibodies are being used ever more often in oncology. However, some patients were excluded from clinical trials because of their comorbidities despite their potentially higher cancer frequencies, as is the case for immunocompromised patients.Areas covered: We analyzed reported preclinical and clinical information and evaluated the risk/benefit ratio for four immunocompromised populations: people living with human immunodeficiency virus (PLHs), solid-organ transplant recipients, recipients of hematopoietic stem-cell allografts, and patients with autoimmune diseases.Expert commentary: Information available in the literature is fragmentary and scarce, making it difficult to evaluate the risk/benefit ratio. It can, nonetheless, be noted that ICI use in PLHs seems possible. For solid-organ transplant recipients, the risk for the graft seems elevated. For the other two populations, it is difficult to conclude at this time.

Metalloproteinases (MMPs) contribute to tissue remodeling and acute inflammation not only by degrading extracellular matrix proteins but also by controlling the influx of chemokines through the regulation and shedding of syndecans. B-lymphocytes, in addition to their well-known function as antibody producing cells, participate in the innate immune response by secreting inflammatory cytokines and chemokines. However, there is little information about the role of B-lymphocytes in the regulation of MMPs; consequently, herein we investigated whether activated human circulating B-lymphocytes contributed to the secretion of MMPs. We demonstrate that B-lymphocytes activated by un-methylated CpG motifs, found in bacterial DNA, and β-glucans, found in the cell wall of fungi, both induced MMP-7. Interestingly, while CpG-stimulated cells activated the mTOR pathway via TLR9 receptor to induced MMP-7, β-glucan-stimulated cells were mTOR-independent and used Dectin-1 receptor. B-lymphocytes did not seem to have a major role in the secretion of tissue inhibitors of metalloproteinases (TIMPs). However, secreted MMP-7 participated in the shedding of Syndecan-4 from the surface of B-lymphocytes. In conclusion, circulating human B-lymphocytes contribute to the regulation of the innate immune system by participating in the secretion of MMP-7 which in turn is important for the shedding of Syndecan-4 in response to infectious stimuli.

To estimate the incidence and mortality rates, and their evolution over time, of physician-diagnosed primary Sjögren syndrome (pSS) in residents of Olmsted County, Minnesota.Medical records of patients with a diagnosis or suspicion of SS in Olmsted County from January 1, 2006, through December 31, 2015, were reviewed to identify incident cases of pSS (defined by physician diagnosis). These cases were combined with those from a 1976 through 2005 incident cohort (n=111) from the same population. Incidence rates were age and sex adjusted to the 2010 US white population. Survival rates were compared with the expected rates in the population of Minnesota.With 61 incident cases of pSS diagnosed in Olmsted County from 2006 through 2015, the total cohort included 172 patients with incident pSS from 1976 through 2015. Of the 172 patients, 151 (88%) were women and 161 (94%) were white, with a mean ± SD age at diagnosis of 58.3±16.7 years. The average age- and sex-adjusted annual incidence for 2006 through 2015 was 5.9 per 100,000 population (95% CI, 4.4-7.4 per 100,000 population), and the overall incidence for the entire period was 5.8 per 100,000 (95% CI, 4.9-6.6 per 100,000). The incidence increased with calendar time over the 40-year period (P=.005). There was no difference in mortality in the pSS cohort compared with expected (standardized mortality ratio, 1.15; 95% CI, 0.86-1.50).The average annual incidence of pSS in this population-based cohort was 5.8 per 100,000, with a progressive increase over the 40 years of the study. Overall survival of patients with pSS was not different from that of the general population.

We read with a great interest the recent contribution by Mossel et al ,1 which brings new important data on the diagnostic value of major salivary ultrasonography (SGUS) for primary Sjogren’s syndrome (pSS), as reported in previous studies2–10 and stresses the need to evaluate the inclusion of SGUS into future classification criteria for the disease. In their cohort of 103 consecutive patients clinically suspected for pSS, Mossel et al reported a good agreement between SGUS and labial and parotid salivary gland biopsy. Agreement was reported to be marginally higher for parotid gland biopsy (absolute agreement of 83%) than for minor labial salivary gland biopsy (absolute agreement 79%). However, the …

Background The goal of the IMI PRECISESADS project is to reclassify individuals affected by systemic autoimmune diseases (SADs) into clusters of molecular, instead of clinical entities. In this project, we investigated 40 individuals with primary Sjögren’s syndrome (pSS), as diagnosed by the revised American-European classification criteria and 53 healthy controls (HCs) to determine whether a fine flow cytometry analysis of T and B cell distribution in whole blood could cluster individuals according to disease activity. Methods Two flow cytometry panels were designed. The first panel was dedicated to T cells and combined CD57, CD45RA, CD62L, CD27, CD38, CD3, CD4, CD8 mAbs. The second panel was dedicated to B cells and combined IgD, TACI, CD5, CD24, CD27, CD38, CD19, CD11b mAbs. Disease activity was determined according to ESSDAI. A Luminex-based system was used to analyse up to 88 cytokines, chemokines and soluble factors in serum of pSS patients and HCs. Results A combined analysis of T and B cell distribution highlighted two groups of individuals with a strong cluster of pSS patients in one of them. Indeed, the first cluster (group A) gathers together 24 pSS patients and 5 HCs while group B consists in 16 pSS patients and 49 HCs. Group A was characterised by the association of an increase of activated naïve B cells (IgD+ CD24+ CD38+ CD27-), a decrease of memory B cells (IgD- CD38- CD27+), an increase of central memory CD4+ T cells (CD45RA- CD62L+ CD27+), an increase of effector memory CD8+ T cells (CD45RA- CD62L- CD27-) and a decrease of central memory CD8+ T cells (CD45RA- CD62L+ CD27+) compared to group B and 91% of HCs. Interestingly, group A patients have a higher disease activity score when compared to group B (Mean ESSDAI score 5.0±1.9 versus 3.2±0.9 respectively). Finally, serum pro-inflammatory molecules were markedly up-regulated in group A patients compared to HCs and group B patients. Conclusions A fine flow cytometry analysis of T and B cell subsets characterises pSS patients with a higher disease activity score and a pro-inflammatory signature. Similar approaches are ongoing in the context of the PRECISESADS project for other SADs. This work has received support from the EU/EFPIA Innovative Medicines Initiative Joint Undertaking PRECISESADS grant n° 115565.www.precisesads.eu