Physiopathologie de la polyarthrite rhumatoïde : effet inflammatoire des auto-anticorps anti-protéines citrullinées et du facteur rhumatoïde

La polyarthrite rhumatoïde (PR) est une maladie auto-immune qui touche 0,5 à 1% de la population adulte mondiale. Bien que des facteurs génétiques et environnementaux de susceptibilité soient identifiés, son étiologie reste inconnue. Sur le plan clinique, elle se caractérise par une inflammation chronique des articulations synoviales menant à une destruction ostéo-cartilagineuse. Sur le plan biologique, elle est marquée par la présence d'auto-anticorps chez la majorité des patients. Parmi ceux-ci, les auto-anticorps anti-protéines citrullinées (ACPA) et le facteur rhumatoïde (FR) sont détectés chez environ 80% des patients et associés aux formes les plus actives et les plus sévères de la maladie. Outre leur intérêt diagnostique et, à un moindre degré, pronostique, les ACPA et le FR sont synthétisés dans le tissu synovial où leurs cibles respectives, la fibrine citrullinée principalement et les IgG (fragments Fc), sont également abondantes. Au sein des articulations, ces auto-anticorps sont donc susceptibles de former des complexes immuns avec leurs antigènes-cibles articulaires, d'activer des voies effectrices mettant en jeu des récepteurs aux fragments Fc des immunoglobulines (FcR) et/ou l'activation du complément, et de participer ainsi à la promotion de l'inflammation articulaire. Notre laboratoire s'est précédemment intéressé au rôle des ACPA dans l'inflammation du tissu synovial rhumatoïde. Un modèle cellulaire entièrement humain a été développé dans le but de mimer l'interaction entre macrophages et complexes immuns issus de la fixation d'ACPA à la fibrine citrullinée, présente sous forme de dépôts dans le tissu synovial rhumatoïde. Ce modèle utilise des macrophages différenciés à partir de monocytes d'individus sains stimulés par des complexes immuns reconstitués par immunocapture sélective d'IgG ACPA de patients atteints de PR par du fibrinogène citrulliné immobilisé (CI-ACPA). Il a permis d'établir la capacité de ces complexes immuns à induire une sécrétion macrophagique de TNF-α via la fixation à des récepteurs aux fragments Fc des IgG (FcγR) activateurs, parmi lesquels le récepteur FcγRIIa joue un rôle prépondérant. Le TNF-α étant une cytokine-clé de la synovite rhumatoïde, ces résultats ont renforcé l'hypothèse selon laquelle les ACPA seraient un inducteur majeur de sa sécrétion. Nous avons poursuivi l'étude de l'implication des auto-anticorps dans la physiopathologie de la PR, en étudiant l'influence du FR sur l'effet pro inflammatoire des ACPA dans le même modèle in vitro. Nous avons tout d'abord évalué l'influence de deux FR de classe IgM (FR IgM) monoclonaux purifiés à partir de sérums de patients atteints de cryoglobulinémie de type II. Nous avons observé que l'incorporation de chacun de ces FR aux CI-ACPA amplifiait fortement la sécrétion macrophagique de TNF-α. Plus globalement, ceci s'accompagnait d'un déséquilibre de la réponse cytokinique des macrophages en faveur de l'inflammation : la sécrétion d'autres cytokines pro-inflammatoires telles que l'IL-1β, l'IL-6 ou l'IL-8 et le rapport de sécrétion TNF α:IL-10 étaient augmentés, alors que le rapport IL1-Ra:IL-1β était diminué. En outre, le mélange de cytokines sécrétées par les macrophages a induit une sécrétion accrue d'IL-6 par des fibroblastes synoviaux provenant de patients atteints de PR. De plus, nous avons confirmé que les CI-ACPA n'induisaient pas la sécrétion de TNF α par les monocytes circulants dont étaient dérivés les macrophages et nous avons montré que l'incorporation de FR IgM dans les complexes immuns ne permettait pas non plus de l'induire. Par ailleurs, nous avons établi qu'à lui seul, le FR IgM ne provoquait pas de sécrétion de TNF α par les macrophages. L'amplification de la réponse cytokinique aux CI-ACPA qu'induit le FR IgM provient très probablement d'une augmentation de la signalisation activatrice en aval de FcγR, puisqu'elle est dépendante du recrutement d'un nombre accru d'IgG dans les complexes immuns, captées par l'IgM multivalente. Enfin, nous avons observé l'activation de la cascade du complément par les CI-ACPA, et la majoration de ce phénomène en présence de FR IgM. Dans un deuxième temps, nous avons étudié l'influence de « l'authentique » FR IgM, c'est-à-dire celui présent chez les patients atteints de PR, et posé la question de l'influence du FR de classe IgA (FR IgA) de la même provenance. Nous avons purifié ces deux classes de FR par différentes étapes de chromatographie d'affinité, à partir de plusieurs mélanges de sérums de patients atteints de PR. Avec chacune des deux classes de FR, nous avons montré que leur incorporation aux CI-ACPA induisait systématiquement l'amplification de la réponse TNF-α macrophagique et que la réponse cytokinique était globalement déséquilibrée en faveur de l'inflammation. Poursuivant l'étude des mécanismes de l'amplification de la réponse cytokinique par le FR IgM, nous avons définitivement écarté l'hypothèse d'une signalisation en aval d’un récepteur aux IgM. En effet, aucun des récepteurs aux IgM leucocytaires déjà décrits, c'est à dire ni le récepteur FcµR, ni le récepteur Fcα/µR, n'était exprimé à la surface des macrophages de notre modèle. En outre, la stimulation des macrophages par des IgM ou leur portion Fc ni n'a induit la sécrétion de TNF-α, ni n'a modulé celle induite par des complexes immuns contenant des IgG. Par contre, nous avons confirmé que le récepteur FcαRI aux IgA était exprimé à la surface des macrophages et, par blocage sélectif de la fixation des FR IgA à ce récepteur, nous avons montré sa participation dans l'activation des macrophages par les CI-ACPA ayant incorporé le FR IgA. Enfin, nous avons observé que l'amplification de la réponse TNF-α par les FR IgM et IgA augmentait drastiquement en présence de LPS. Ceci montre une possible coopération des voies des FcR et du TLR4 dans la synovite, en cas de présence simultanée de complexes immuns et de ligands de TLR4. Une telle coopération est probable puisque divers ligands endogènes potentiels de TLR4 ont été décrits au sein du tissu synovial rhumatoïde. Au total, FR IgM et FR IgA apparaissent comme des amplificateurs majeurs de la réaction inflammatoire induite par les ACPA au sein du tissu synovial rhumatoïde. Nous avons précisé leur mode d'action qui correspond à l'interaction des CI-ACPA avec les macrophages via leurs FcR mais aussi, très probablement, au moins pour le FR IgM, via les récepteurs du complément. Ces résultats apportent un éclairage inédit sur le rôle, suspecté mais jamais démontré du FR IgM mais aussi du FR IgA, dans la physiopathogénie de la PR. La pertinence de stratégies thérapeutiques qui viseraient à contrecarrer l'activation des mécanismes effecteurs pro-inflammatoires induits par les CI contenant ACPA et FR, ou à inhiber la production de ces auto-anticorps, est ainsi fortement renforcée.Rheumatoid Arthritis (RA) is the most frequent human auto-immune disease affecting 0.5 to 1% of the population worldwide. The exact cause of RA is unknown, but joint inflammation is thought to be the result of an uncontrolled autoimmune response arising from a combination of environmental and genetic factors. Among autoantibodies found in RA patients, autoantibodies to citrullinated proteins (ACPA) and rheumatoid factors (RF) are present in the serum of about 80% of patients and are frequently associated with active and severe forms of RA. In addition to their diagnostic and prognostic interest, ACPA and RF are synthesized by plasma cells of the rheumatoid synovial membrane at the very place where their respective target, citrullinated fibrin and Fc fragment of IgG are also present and abundant. In the joints, these autoantibodies are likely to form immune complexes with their articular antigenic targets able to trigger inflammatory effector responses depending on Fc receptors to immunoglobulins and/or complement activation, thus to promote joint inflammation. During the last few years, our laboratory dedicated itself to the understanding of the role of ACPA in RA synovial tissue inflammation. A totally human in vitro model was developed in order to mimic the interactions between synovial macrophages and immune complexes formed by ACPA and citrullinated fibrin, their major target in the synovial tissue. In this model, ACPA immune complexes (ACPA-IC) are generated by selective immunocapture of IgG ACPA from the serum of RA patients by in vitro citrullinated human fibrinogen immobilized onto culture wells into which macrophages, differentiated from CD14-positive monocytes of healthy human donors, are then seeded. This model allowed demonstrating the ability of ACPA-IC to induce TNF-α secretion by macrophages through signalling downstream of activating Fc receptors to IgG (FcγR), predominantly the FcγRIIa receptor. Considering that TNF-α is a key pro-inflammatory cytokine in the disease, these results strengthened the hypothesis that ACPA have an inflammatory potential and a major role in RA. To further study the involvement of autoantibodies in the pathophysiology of RA we undertook to assess the influence of RF on the pro-inflammatory role of ACPA in the in vitro model. We first evaluated the influence of two monoclonal RF IgM paraproteins from patients with type II cryoglobulinemia. Each RF IgM strongly amplified the macrophage TNF-α secretion induced by ACPA-IC. Overall, incorporation into ACPA-IC of RF IgM shifted the macrophage secretion of cytokines toward an even more pronounced proinflammatory profile as it increased the TNF-α:IL 10 ratio and the IL 6 and IL-8 secretions and decreased the IL 1Ra:IL 1β ratio. Moreover, the secreted cytokine cocktail exhibited an enhanced capacity to prompt IL-6 secretion by RA synovial fibroblasts. Then, we confirmed that ACPA-IC did not induce TNF-α secretion by monocytes and showed that they remained unresponsive even after incorporation of RF IgM. Besides, we showed that RF IgM alone did not induce TNF-α macrophage secretion. The amplification of the macrophage cytokine responses probably arises from an increase in the activating signals downstream of FcγRs due to an RF IgM-mediated recruitment of more IgG into ACPA-IC. Finally, in vitro complement activation by immobilized ACPA-IC markedly increased when their formation occurred in the presence of RF IgM. Secondly, we evaluated the influence of the RF IgM and RF IgA found in RA patients. These were purified from RA serum pools by serial affinity chromatographies. Incorporation of each class of RF systematically amplified the macrophage cytokine secretion and skewed it in favour of proinflammatory cytokines. Further exploration of the mechanism of the RF IgM-mediated amplification allowed ruling out the involvement of any signalling downstream of an IgM receptor. Indeed, neither FcµR nor Fcα/µR, the only recognized leucocyte IgM receptors, was expressed by the macrophages of our model. Moreover, macrophages did not secrete any TNF α in response to IgM or IgM Fc portions and their TNF α response to IgG-containing immune complexes was not modulated by the simultaneous presence of IgM or Fc5µ fragments. However, we confirmed that the receptor FcαRI for IgA Fc was expressed and that it participates in the activating signalling cascade induced by ACPA-IC formed in the presence of RF IgA IC since selective blockade of the binding of RF IgA to this receptor inhibited the TNF-α response to these immune complexes. Finally, LPS substantially enhanced induction of macrophage TNF-α by ACPA-IC containing RF IgM or RF IgA. Considering that numerous endogenous TLR4 ligands have been described in the RA synovial tissue, this indicates that simultaneous FcR engagement by RA-associated IC and ligation of TLR4 by such endogenous ligands likely cooperate in promoting synovial tissue inflammation. Therefore, RF IgM and RF IgA appear as major enhancers of the inflammatory reaction induced by ACPA in the RA synovial tissue. We show that this is most probably mediated by FcR triggering but also, concerning RF IgM, by activation of the complement cascade. Our studies shed new light on the largely accepted but poorly mechanistically documented role of RF IgM and RF IgA in RA pathophysiology. They also emphasize the relevance of therapeutic strategies aiming at downregulating the proinflammatory effectors mechanisms triggered by immunes complexes containing ACPA and RF or seeking inhibition of the production of theses autoantibodies

CD23, molécule de l immunité innée ?

ANGERS-BU Médecine-Pharmacie (490072105) / SudocSudocFranceF

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IgM and IgA Rheumatoid Factors Purified from Rheumatoid Arthritis Sera Boost the Fc Receptor– and Complement-Dependent Effector Functions of the Disease-Specific Anti–Citrullinated Protein Autoantibodies

International audienceRheumatoid factors (RF) and the disease-specific anti-citrullinated protein autoantibodies (ACPA) coexist in the joints of rheumatoid arthritis (RA) patients where they probably contribute to synovitis. We investigated the influence of IgM and IgA RF on the FcR- and complement-dependent effects of ACPA immune complexes (ACPA-IC). When stimulated by ACPA-IC formed in the presence of IgM RF or IgA RF fractions purified from RA serum pools, M-CSF-generated macrophages skewed their cytokine response toward inflammation, with increases in the TNF-α/IL-10 ratio and in IL-6 and IL-8 secretion, and decreases in the IL-1Ra/IL-1β ratio. In the IgM RF-mediated amplification of the inflammatory response of macrophages, the participation of an IgM receptor was excluded, notably by showing that they did not express any established receptor for IgM. Rather, this amplification depended on the IgM RF-mediated recruitment of more IgG into the ACPA-IC. However, the macrophages expressed FcαRI and blocking its interaction with IgA inhibited the IgA RF-mediated amplification of TNF-α secretion induced by ACPA-IC, showing its major implication in the effects of RF of the IgA class. LPS further amplified the TNF-α response of macrophages to RF-containing ACPA-IC. Lastly, the presence of IgM or IgA RF increased the capacity of ACPA-IC to activate the complement cascade. Therefore, specifically using autoantibodies from RA patients, the strong FcR-mediated or complement-dependent pathogenic potential of IC including both ACPA and IgM or IgA RF was established. Simultaneous FcR triggering by these RF-containing ACPA-IC and TLR4 ligation possibly makes a major contribution to RA synovitis

IgM rheumatoid factor amplifies the inflammatory response of macrophages induced by the rheumatoid arthritis-specific immune complexes containing anticitrullinated protein antibodies

International audienceObjectives: Anticitrullinated protein antibodies (ACPA) are specifically associated with rheumatoid arthritis (RA) and produced in inflamed synovial membranes where citrullinated fibrin, their antigenic target, is abundant. We showed that immune complexes containing IgG ACPA (ACPA-IC) induce FcγR-mediated tumour necrosis factor (TNF)-α secretion in macrophages. Since IgM rheumatoid factor (RF), an autoantibody directed to the Fc fragment of IgG, is also produced and concentrated in the rheumatoid synovial tissue, we evaluated its influence on macrophage stimulation by ACPA-IC.Methods: With monocyte-derived macrophages from more than 40 healthy individuals and different human IgM cryoglobulins with RF activity, using a previously developed human in vitro model, we evaluated the effect of the incorporation of IgM RF into ACPA-IC. Results: IgM RF induced an important amplification of the TNF-α secretion. This effect was not observed in monocytes and depended on an increase in the number of IgG-engaged FcγR. It extended to the secretion of interleukin (IL)-1β and IL-6, was paralleled by IL-8 secretion and was not associated with overwhelming secretion of IL-10 or IL-1Ra. Moreover, the RF-induced increased proinflammatory bioactivity of the cytokine response to ACPA-IC was confirmed by an enhanced, not entirely TNF-dependent, capacity of the secreted cytokine cocktail to prompt IL-6 secretion by RA synoviocytes. Conclusions: By showing that it can greatly enhance the proinflammatory cytokine response induced in macrophages by the RA-specific ACPA-IC, these results highlight a previously undescribed, FcγR-dependent strong proinflammatory potential of IgM RF. They clarify the pathophysiological link between the presence of ACPA and IgM RF, and RA severity

Heterogeneity and Lobularity of Pancreatic Pathology in Type 1 Diabetes during the Prediabetic Phase

Type 1 diabetes (T1D) is an autoimmune disease in which insulin-producing beta cells are destroyed in the islets of Langerhans. One of its main pathological manifestations is the hyper-expression of Major Histocompatibility Complex I (MHC-I) by beta cells, which was first described over 3 decades ago yet its cause remains unknown. It might not only be a sign of beta cell dysfunction but could also render the cells susceptible to autoimmune destruction; for example, by islet-infiltrating CD8 T cells. In this report, we studied pancreas tissue from a 22-year-old non-diabetic male cadaveric organ donor who had been at high risk of developing T1D, in which autoantibodies against GAD and IA-2 were detected. Pancreas sections were analyzed for signs of inflammation. Multiple insulin-containing islets were identified, which hyper-expressed MHC-I. However, islet density and MHC-I expression exhibited a highly lobular and heterogeneous pattern even within the same section. In addition, many islets with high expression of MHC-I presented higher levels of CD8 T cell infiltration than normal islets. These results demonstrate the heterogeneity of human pathology that occurs early during the pre-diabetic, autoantibody positive phase, and should contribute to the understanding of human T1D

Loss of IDO1 Expression From Human Pancreatic β-Cells Precedes Their Destruction During the Development of Type 1 Diabetes

Indoleamine 2,3 dioxygenase-1 (IDO1) is a powerful immunoregulatory enzyme that is deficient in patients with type 1 diabetes (T1D). In this study, we present the first systematic evaluation of IDO1 expression and localization in human pancreatic tissue. Although IDO1 was constitutively expressed in β-cells from donors without diabetes, less IDO1 was expressed in insulin-containing islets from double autoantibody-positive donors and patients with recent-onset T1D, although it was virtually absent in insulin-deficient islets from donors with T1D. Scatter plot analysis suggested that IDO1 decay occurred in individuals with multiple autoantibodies, prior to β-cell demise. IDO1 impairment might therefore contribute to β-cell demise and could potentially emerge as a promising therapeutic target

Plasma microRNA signature in presymptomatic and symptomatic subjects with C9orf72-associated frontotemporal dementia and amyotrophic lateral sclerosis

International audienceObjective: To identify potential biomarkers of preclinical and clinical progression in chromosome 9 open reading frame 72 gene (C9orf72)-associated disease by assessing the expression levels of plasma microRNAs (miRNAs) in C9orf72 patients and presymptomatic carriers. Methods: The PREV-DEMALS study is a prospective study including 22 C9orf72 patients, 45 presymptomatic C9orf72 mutation carriers and 43 controls. We assessed the expression levels of 2576 miRNAs, among which 589 were above noise level, in plasma samples of all participants using RNA sequencing. The expression levels of the differentially expressed miRNAs between patients, presymptomatic carriers and controls were further used to build logistic regression classifiers. Results: Four miRNAs were differentially expressed between patients and controls: miR-34a-5p and miR-345-5p were overexpressed, while miR-200c-3p and miR-10a-3p were underexpressed in patients. MiR-34a-5p was also overexpressed in presymptomatic carriers compared with healthy controls, suggesting that miR-34a-5p expression is deregulated in cases with C9orf72 mutation. Moreover, miR-345-5p was also overexpressed in patients compared with presymptomatic carriers, which supports the correlation of miR-345-5p expression with the progression of C9orf72-associated disease. Together, miR-200c-3p and miR-10a-3p underexpression might be associated with full-blown disease. Four presymptomatic subjects in transitional/prodromal stage, close to the disease conversion, exhibited a stronger similarity with the expression levels of patients. Conclusions: We identified a signature of four miRNAs differentially expressed in plasma between clinical conditions that have potential to represent progression biomarkers for C9orf72-associated frontotemporal dementia and amyotrophic lateral sclerosis. This study suggests that dysregulation of miRNAs is dynamically altered throughout neurodegenerative diseases progression, and can be detectable even long before clinical onset. Trial registration number NCT02590276. copyright

MP_Database.v1.1 : première base de données permettant l’intercomparaison des contaminations en microplastiques dans les dépôts sédimentaires à différentes échelles spatiales et temporelles

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