Importance Of Toll-Like Receptors For B Lymphocyte Survival In Primary Sjögren’s Syndrome

The Sjögren's syndrome is a systemic autoimmune disease characterized by lymphocytic infiltration of the glands responsible for mouth and eyes dryness. A minority of infiltrating B cells is organized as germinal centers while the majority is aggregated into clusters of transitional and marginal zone B cells. The Toll-like receptor 9 (TLR9) recognizes microbial DNA but also, sometimes, the self DNA. It appears to be a key determinant of the survival and differentiation of B lymphocytes. After laser micro-dissection of B cells from salivary glands, analyses by quantitative RT-PCR showed that transitional B cells express high level of TLR9 mRNA unlike B cells from germinal centers. B lymphocytes from healthy donors were sorted by flow cytometry and stimulated in vitro with their TLR9. It induces survival, activation and proliferation associated with phenotypic changes. Transitional B cells exhibited characteristics of the marginal zone, whereas mature B cells expressed follicular germinal center specificities. Finally, IgM and IgG were secreted by both population, but with elevated production of autoantibodies by the transitional B cells. Increased expression of TLR9 by transitional B cells suggests that they may be highly sensitive to differentiate into autoantibody secreting cells through maturation into the marginal zone into the salivary glands. TLR9 might be a target for forthcoming biotherapies. Le syndrome de Gougerot-Sjögren est une maladie autoimmune systémique caractérisée par une infiltration lymphocytaire des glandes responsable d'une sécheresse buccale et oculaire. Une minorité des lymphocytes B infiltrants est organisée en centres germinatifs tandis que la majorité est regroupée en agrégats de lymphocytes B transitionnels et de la zone marginale. Le Toll-like receptor 9 (TLR9) reconnaît l'ADN microbien mais aussi, parfois, l'ADN du soi. Il apparaît donc comme un élément déterminant de la survie et la différenciation des lymphocytes B. Après micro-dissection laser des lymphocytes B des glandes salivaires, une analyse par RT-PCR quantitative a montré que les lymphocytes B transitionnels expriment fortement l'ARNm de TLR9 contrairement à ceux des centres germinatifs. Des lymphocytes B de donneurs sains ont été triés par cytométrie en flux puis stimulés in vitro par leur TLR9. Il s’ensuit une survie, une activation et une prolifération associées à des modifications phénotypiques. Les lymphocytes B transitionnels présentent des caractéristiques de la zone marginale, tandis que les lymphocytes B matures expriment des spécificités folliculaires des centres germinatifs. Enfin, des IgM et des IgG sont sécrétées par les deux types de population, mais avec une production d'auto-anticorps plus élevée issue de la différenciation des lymphocytes B transitionnels. L’expression accrue de TLR9 par les lymphocytes B transitionnels suggère qu'ils pourraient être particulièrement sensibles à une différenciation en cellules sécrétrices d'auto-anticorps par une maturation vers la zone marginale au sein des glandes salivaires. Le TLR9 pourrait bien devenir la cible des futures biothérapies.The Sjögren's syndrome is a systemic autoimmune disease characterized by lymphocytic infiltration of the glands responsible for mouth and eyes dryness. A minority of infiltrating B cells is organized as germinal centers while the majority is aggregated into clusters of transitional and marginal zone B cells. The Toll-like receptor 9 (TLR9) recognizes microbial DNA but also, sometimes, the self DNA. It appears to be a key determinant of the survival and differentiation of B lymphocytes. After laser micro-dissection of B cells from salivary glands, analyses by quantitative RT-PCR showed that transitional B cells express high level of TLR9 mRNA unlike B cells from germinal centers. B lymphocytes from healthy donors were sorted by flow cytometry and stimulated in vitro with their TLR9. It induces survival, activation and proliferation associated with phenotypic changes. Transitional B cells exhibited characteristics of the marginal zone, whereas mature B cells expressed follicular germinal center specificities. Finally, IgM and IgG were secreted by both population, but with elevated production of autoantibodies by the transitional B cells. Increased expression of TLR9 by transitional B cells suggests that they may be highly sensitive to differentiate into autoantibody secreting cells through maturation into the marginal zone into the salivary glands. TLR9 might be a target for forthcoming biotherapies. Le syndrome de Gougerot-Sjögren est une maladie autoimmune systémique caractérisée par une infiltration lymphocytaire des glandes responsable d'une sécheresse buccale et oculaire. Une minorité des lymphocytes B infiltrants est organisée en centres germinatifs tandis que la majorité est regroupée en agrégats de lymphocytes B transitionnels et de la zone marginale. Le Toll-like receptor 9 (TLR9) reconnaît l'ADN microbien mais aussi, parfois, l'ADN du soi. Il apparaît donc comme un élément déterminant de la survie et la différenciation des lymphocytes B. Après micro-dissection laser des lymphocytes B des glandes salivaires, une analyse par RT-PCR quantitative a montré que les lymphocytes B transitionnels expriment fortement l'ARNm de TLR9 contrairement à ceux des centres germinatifs. Des lymphocytes B de donneurs sains ont été triés par cytométrie en flux puis stimulés in vitro par leur TLR9. Il s’ensuit une survie, une activation et une prolifération associées à des modifications phénotypiques. Les lymphocytes B transitionnels présentent des caractéristiques de la zone marginale, tandis que les lymphocytes B matures expriment des spécificités folliculaires des centres germinatifs. Enfin, des IgM et des IgG sont sécrétées par les deux types de population, mais avec une production d'auto-anticorps plus élevée issue de la différenciation des lymphocytes B transitionnels. L’expression accrue de TLR9 par les lymphocytes B transitionnels suggère qu'ils pourraient être particulièrement sensibles à une différenciation en cellules sécrétrices d'auto-anticorps par une maturation vers la zone marginale au sein des glandes salivaires. Le TLR9 pourrait bien devenir la cible des futures biothérapies

Potential infection of grazing cattle via contaminated water: a theoretical modelling approach

Wastewater discharge and agricultural activities may pose microbial risks to natural water sources. The impact of different sources can be assessed by water quality modelling. The aim of this study was to use hydrological and hydrodynamic models to illustrate the risk of exposing grazing animals to faecal pollutants in natural water sources, using three zoonotic faecal pathogens as model microbes and fictitious pastures in Sweden as examples. Microbial contamination by manure from fertilisation and grazing was modelled by use of a hydrological model (HYPE) and a hydrodynamic model (MIKE 3 FM), and microbial contamination from human wastewater was modelled by application of both models in a backwards process. The faecal pathogens Salmonella spp., verotoxin-producing Escherichia coli O157:H7 (VTEC) and Cryptosporidium parvum were chosen as model organisms. The pathogen loads on arable land and pastures were estimated based on pathogen concentration in cattle faeces, herd prevalence and within-herd prevalence. Contamination from human wastewater discharge was simulated by estimating the number of pathogens required from a fictitious wastewater discharge to reach a concentration high enough to cause infection in cattle using the points on the fictitious pastures as their primary source of drinking water. In the scenarios for pathogens from animal sources, none of the simulated concentrations of salmonella exceeded the concentrations needed to infect adult cattle. For VTEC, most of the simulated concentrations exceeded the concentration needed to infect calves. For C. parvum, all the simulated concentrations exceeded the concentration needed to infect calves. The pathogen loads needed at the release points for human wastewater to achieve infectious doses for cattle were mostly above the potential loads of salmonella and VTEC estimated to be present in a 24-h overflow from a medium-size Swedish wastewater treatment plant, while the required pathogen loads of C. parvum at the release points were below the potential loads of C. parvum in a 24-h wastewater overflow. Most estimates in this study assume a worst-case scenario. Controlling zoonotic infections at herd level prevents environmental contamination and subsequent human exposure. The potential for infection of grazing animals with faecal pathogens has implications for keeping animals on pastures with access to natural water sources. As the infectious dose for most pathogens is more easily reached for calves than for adult animals, and young calves are also the main shedders of C. parvum, keeping young calves on pastures adjacent to natural water sources is best avoided

TOLL-LIKE RECEPTOR 9 DRIVES THE MATURATION OF B LYMPHOCYTES IN THE SALIVARY GLANDS OF PATIENTS WITH SJÖGREN’S SYNDROME

Oral Communication presented at the ";Forum des Jeunes Chercheurs";, Brest (France) 2011

Can pain be self-managed?:Pain change in vulnerable participants to a health education programme

Chronic pain exerts a significant impact on the quality of life, giving rise to both physical and psycho-social vulnerabilities. It not only leads to direct costs associated with treatments, but also results in indirect costs due to the reduced productivity of affected individuals. Chronic conditions can be improved by reducing modifiable risk factors. Various educational programs, including the Chronic Disease Self-Management Programme (CDSMP), have demonstrated the advantages of enhancing patient empowerment and health literacy. Nevertheless, their efficacy in addressing pain symptoms has received limited attention, especially concerning vulnerable populations. This research aims to assess the effectiveness of the CDSMP in alleviating pain among socio-economically vulnerable participants with chronic conditions. By accounting for a wide range of variables, and using data from the EFFICHRONIC project (EU health programme), we investigated the changes in pain levels after the intervention, among 1070 participants from five European countries. Our analyses revealed a significant reduction in pain following the intervention. This finding supports the notion that training programs can effectively ameliorate pain and alleviate its impact on the quality of life, particularly in vulnerable populations. Younger participants, as well as those with higher education levels and individuals experiencing higher levels of pain at baseline, were more likely to experience a reduction in their pain levels. These findings underscore the importance of recognising the social determinants of health. The study was registered at ClinicalTrials.gov (ISRCTN70517103).</p

New ELISA for B Cell-Activating Factor

Item does not contain fulltextWorkshop Theory and Methodology - 26th ISAGA Conference, 18 juli 1995Valencia, Spain : [S.n.]24 p

Comparative Network Analysis of Preterm vs. Full-Term Infant-Mother Interactions

Several studies have reported that interactions of mothers with preterm infants show differential characteristics compared to that of mothers with full-term infants. Interaction of preterm dyads is often reported as less harmonious. However, observations and explanations concerning the underlying mechanisms are inconsistent. In this work 30 preterm and 42 full-term mother-infant dyads were observed at one year of age. Free play interactions were videotaped and coded using a micro-analytic coding system. The video records were coded at one second resolution and studied by a novel approach using network analysis tools. The advantage of our approach is that it reveals the patterns of behavioral transitions in the interactions. We found that the most frequent behavioral transitions are the same in the two groups. However, we have identified several high and lower frequency transitions which occur significantly more often in the preterm or full-term group. Our analysis also suggests that the variability of behavioral transitions is significantly higher in the preterm group. This higher variability is mostly resulted from the diversity of transitions involving non-harmonious behaviors. We have identified a maladaptive pattern in the maternal behavior in the preterm group, involving intrusiveness and disengagement. Application of the approach reported in this paper to longitudinal data could elucidate whether these maladaptive maternal behavioral changes place the infant at risk for later emotional, cognitive and behavioral disturbance

OP0137 GENOME-WIDE WHOLE-BLOOD TRANSCRIPTOME PROFILING IN A LARGE EUROPEAN COHORT OF SYSTEMIC SCLEROSIS PATIENTS

Background:The analysis of annotated transcripts from genome-wide expression studies data is of paramount importance to understand the molecular phenomena underlying the occurrence of complex diseases, such as systemic sclerosis (SSc).Objectives:To perform whole-blood transcriptome and pathway analysis on whole-blood (WB) RNA collected in two cohorts of European SSc patients. Via a discovery and validation strategy we aimed at characterizing the molecular pathways that differentiate SSc from controls and that are reproducible in geographically diverse populations.Methods:WB samples from 252 controls and 162 SSc patients were collected in RNA stabilizers. Patients were divided into a discovery (n=79; Southern Europe) and validation cohort (n=83; Central-Western Europe). RNA sequencing was performed by an Illumina assay. Functional annotations of Reactome pathways were performed with the FAIME algorithm. In parallel, a immunophenotyping analysis on 28 circulating cell populations was assessed. We then tested: the presence of differentially expressed genes or pathways and the correlation between absolute cell counts and RNA transcripts/FAIME scores in regression models. Results significant in both populations were considered as replicated.Results:A total of 15224 genes and 1277 related functional pathways were available for analysis. Among these, 99 genes and 225 pathways were significant in both sets. The heatmap in figure shows the relative expression of replicated pathways and the distribution of cases and controls (red and green bars). Among the significant pathways we found a deregulation in: type-I IFN, TLR-cascade and signalling, function of the tumor suppressor p53 protein, platelet degranulation and activation. Correlation analysis showed that the count of several cell subtypes is jointly associated with RNA transcripts or FAIME scores with strong differences in relation to the geographical origin of samples; neutrophils emerged as the major determinant of gene expression in SSc-whole-blood samples.Conclusion:We discovered a set of differentially expressed genes and pathways that could be validated in two independent sets of SSc patients highlighting a number of deregulated molecular processes that have relevance for the pathogenesis of autoimmunity and SSc.Acknowledgments:This work was supported by EU/EFPIA/Innovative Medicines Initiative Joint Undertaking PRECISESADS grant No. 115565.Disclosure of Interests:Lorenzo Beretta Grant/research support from: Pfizer, Guillermo Barturen: None declared, Barbara Vigone: None declared, Chiara Bellocchi: None declared, Nicolas Hunzelmann: None declared, Ellen Delanghe: None declared, László Kovács: None declared, Ricard Cervera: None declared, Maria Gerosa: None declared, Rafaela Ortega Castro: None declared, Isabel Almeida: None declared, Divi Cornec: None declared, Carlo Chizzolini Consultant of: Boehringer Ingelheim, Roche, Jacques-Olivier Pers: None declared, Zuzanna Makowska Employee of: Bayer AG, Anne buttgereit Employee of: Bayer AG, Ralf Lesche Employee of: Bayer, Martin Kerick: None declared, Marta Alarcon-Riquelme: None declared, Javier Martin Ibanez: None declare

The Future of Biologic Agents in the Treatment of Sjögren’s Syndrome

The gain in knowledge regarding the cellular mechanisms of T and B lymphocyte activity in the pathogenesis of Sjögren’s syndrome (SS) and the current availability of various biological agents (anti-TNF-α, IFN- α, anti-CD20, and anti-CD22) have resulted in new strategies for therapeutic intervention. In SS, various phase I and II studies have been performed to evaluate these new strategies. Currently, B cell-directed therapies seem to be more promising than T cell-related therapies. However, large, randomized, placebo-controlled clinical trials are needed to confirm the promising results of these early studies. When performing these trials, special attention has to be paid to prevent the occasional occurrence of the severe side effects