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La signalisation calcique est un mécanisme physiologique essentiel pour l’homéostasie cellulaire, et en particulier pour les cellules immunitaires telles que les lymphocytes B (LB). Chez l’homme, la caractérisation des molécules impliquées dans la signalisation calcique des LB est encore restreinte. Nous avons donc choisi d’étudier les molécules STIM1, STIM2, Orai1 et TRPC1 afin de rendre compte des modifications de la signalisation calcique observées dans deux modèles physiopathologiques : le Lupus Erythémateux Systémique (LES) et la Leucémie Lymphoïde Chronique (LLC). Pour les LB de LES, par rapport à des LB de témoins, ce travail a permis de montrer que la surexpression de la molécule STIM1 dans les LB de LES, surtout au stade des LB transitionnels, était associée avec un influx constitutif et extracellulaire de calcium et que cet influx était responsable d’une phosphorylation modérée de la MAPK Erk1/2 dans les cellules au repos. Cependant, cet effet positif est contrebalancé par un défaut de l’influx SOCE (strore operated calcium entry) et d’un défaut d’activation de la phosphorylation de la MAPK Erk après stimulation du récepteur à l’antigène des LB (BCR). Les effets positifs et négatifs observés sur la phosphorylation d.Erk1/2 sont directement attribuables au niveau d’expression de STIM1 comme nous l’avons démontré en sur-exprimant STIM1 dans des LB de témoins (vecteur pSTIM1-STIM1) ou en ayant recours à un siRNA spécifique de STIM1 dans les LB de LES. Le défaut d’activité régulatrice des LB de LES sur la prolifération des LT dans un modèle de co-culture autologue LT/LB en présence de CpG (activation du TLR9 des LB), et d’anticorps anti-CD3/CD28 (activation des LT) est levé en inhibant l’expression de STIM1 dans ces cellules ce qui permet de restaurer la production d.IL-10 par les LB de LES et la génération de LT régulateurs. Le défaut d’activation de la voie Erk/DNMT1/méthylation de l’ADN dans les LB de LES semble également contrôlée par le niveau d’expression de la molécule STIM1.Pour les LB CD5+ de LLC, l’analyse des partenaires calciques nous a permis de distinguer deux groupes de patients : un premier groupe qui exprime le complexe membranaire STIM1/TRPC1/Orai1 et un second groupe qui n’exprime qu’Orai1. La présence du complexe membranaire STIM1/TRPC1/Orai1 est associée avec un influx constitutif de calcium, la phosphorylation d’Erk1/2, et un défaut d’activation du signal SOCE après stimulation du BCR. Le rôle de la molécule CD5 sur l’influx constitutif des LLC du groupe I a été démontré en utilisant des siRNA spécifiques pourCD5 et en analysant la lignée B humaine Jok-1 transfectée avec CD5. Enfin, l’utilisation d’un anticorps anti-STIM1 est capable d’induire l’apoptose in vitro des LB de LLC du groupe I, alors que l’anticorps anti-CD20, rituximab, n’est efficace que sur les LLC du groupe II.En conclusion, cette étude ouvre de nouvelles perspectives diagnostiques et thérapeutiques en auto-immunité dans le LES et en cancérologie dans la LLC.Calcium signaling is a central mechanism for cellular homeostasis, including for immune cells such as B lymphocytes (B cells). In human, the characterization of the molecules involved in B cell calcium signalisation is still limited and we have focused our study on STIM1, STIM2, Orai1 and TRPC1. In order to better understanding the role of these molecules, two pathologies, with known calcium dysregulations, were chosen : Systemic Lupus Erythematosus (SLE) and Chronic Lymphocytic Leukemia (CLL).For SLE B cells, compared to healthy control (HC) B cells, this work has established that overexpression of the STIM1 molecule characterizes SLE B cells, especially at the transitional B cell stage. STIM1 overexpression in SLE B cells was associated with a constitutive extracellular calcium influx that in turn contributes to the moderate phosphorylation of the MAPK Erk1/2. However, this positive effect is counterbalanced by a defective strore operated calcium entry (SOCE), and a defective Erk phosphorylation after B cell receptor (BCR) activation. The positive and negative effects on the MAPK Erk1/2 in SLE B cells are directly linked to level of STIM1 as demonstrated in HC B cells transfected with a STIM1 vector (pSTIM1-STIM1), or by using a specific siRNA to STIM1 (siSTIM1) in SLE B cells.The incapacity of SLE B cells to regulate T cell proliferation in an autologous co-culture T/B cell model is reversed by using siSTIM1 in SLE B cells. In this model, using siSTIM1 in SLE B cells also restores the production of IL-10 by B cells, and the generation of regulatory T cells. The defective Erk/DNMT1/DNA methylation pathway in SLE B cells is suspected to be under the control of STIM1 overexpression.For CD5+ CLL B cells, analysis of calcium partners has enabled us to distinguish two groups of patients: one group that expresses at the plasma membrane the complex STIM1/TRPC1/Orai1 and a second group that expresses onlyOrai1. The presence of the membrane complex STIM1/TRPC1/Orai1 in CLL group I is associated with a constitutive extracellular calcium influx, the basal phosphorylation of Erk1/2, and a defaut to mobilize SOCE upon BCR engagement. In the CLL group I patients, a role of CD5 on the constitutive extracellular calcium influx was shown using siRNA specific to CD5 and using the transfected CD5 expressing B cell line Jok-1. Finally, the in vitro use of an anti-STIM1 antibody is able to induce apoptosis in CLL B cells from group I patients, while the anti-CD20 antibody (rituxan) was only effective in the CLL B cells from group II patients.In conclusion, this study opens new diagnostic and therapeutic perspectives in autoimmunity and in cancer

Impact of stress on aged immune system compartments: Overview from fundamental to clinical data

International audienceLife expectancy is continuously increasing due to major progress in preventing, delaying or curing various pathologies normally encountered in old age. However, both scientific and medical advances are still required to understand underlying cause of the disparate comorbidities occurrence with aging. In one hand, aging profoundly impairs the immune system; it is characterized by many changes in haematopoiesis, adaptive and innate systems, associated with pro-inflammatory environment. In another hand, stressful events (acute or chronic) can also impact the immune system through the secretion of hormones, which are also altered with aging. The field of psychoneuroimmunology is now providing evidences that in acute medical conditions, elderly people are not equal in their responses to stressors depending on many extrinsic and intrinsic factors. These parameters could interfere with elderly's ability to mount an effective immune response. The objective of this review is to provide an overview of the literature (from fundamental to clinical observations) to draw a parallel between immune dysregulation caused by stress or by aging. Understanding this entanglement could enable us to target fundamental age-related pathways and thus open new avenues in improving both lifespan and health span

A case for the graft-versus-host disease as a model for B cell-mediated autoimmunity.

International audienceFollowing allogenic hematopoietic stem cell transplantation (HSCT), patients with autoimmune disease or hematopoietic malignancy may develop acute or chronic graft-versus-host (GvH) disease. B lymphocytes, from the recipient as well as from the donor, have recently been implicated in the pathogenesis of such disturbances. Their deleterious effects are accounted for by other tasks B lymphocytes accomplish than the antibody production. We highlight herein some recent observations in the context of B cells in the GvH disease

Metabolic regulation by PPARγ is required for IL-33-mediated activation of ILC2s in lung and adipose tissue

International audienceType 2 innate lymphoid cells (ILC2s) play a critical role early in the response to infection by helminths and in the development of allergic reactions. ILC2s are also involved in the physiologic regulation of adipose tissue and its metabolic response to cold shock. We find that the metabolic sensor peroxisome proliferator-activated receptor gamma (PPARγ) is highly expressed in ILC2s of the lung and adipose tissue and increases responsiveness to IL-33. In turn, activation of ILC2 by IL-33 leads to increased expression of PPARγ, a prerequisite for proliferation and expression of the effector cytokines IL-5 and IL-13. In contrast, pharmacological inhibition of PPARγ leads to decreased expression of CD36 and fatty acid uptake, a necessary source of energy for ILC2s and of potential ligands for PPARγ. As a consequence, treatment of mice with a PPARγ antagonist reduces the severity of an ILC2-dependent acute airway inflammation. Together, our results demonstrate the critical role of the metabolic sensor PPARγ for the functions of ILC2s

New Insights into Lymphocyte Differentiation and Aging from Telomere Length and Telomerase Activity Measurements

International audienceαβ CD8+, γδ, and NK lymphocytes are fundamental effector cells against viruses and tumors. These cells can be divided into multiple subsets according to their phenotype. Based on progressive telomere attrition from naive to late effector memory cells, human CD8+ T cell subsets have been positioned along a pathway of differentiation, which is also considered as a process of lymphocyte aging or senescence. A similar categorization has not been clearly established for γδ and NK cell populations. Moreover, the distinction between the aging of these populations due to cellular differentiation or due to the chronological age of the donor has not been formally considered. In this study, we performed systematic measurements of telomere length and telomerase activity in human αβ CD8+, γδ, and NK lymphocytes based on subset division and across age to address these points and better understand the dichotomy between differentiation and temporal aging. This approach enables us to position phenotypically distinct γδ or NK subsets along a putative pathway of differentiation, such as for CD8+ T cells. Moreover, our data show that both cellular differentiation and donor aging have profound but independent effects on telomere length and telomerase activity of lymphocyte subpopulations, implying distinct mechanisms and consequences on the immune system

Rationale for treating primary Sjögren's syndrome patients with an anti-CD6 monoclonal antibody (Itolizumab).

International audienceCD6 is a cell surface receptor expressed on the majority of T cells and a subset of B cells. When expressed, CD6 contributes to lymphocyte activation through its extracellular domain 1, while adhesion and cellular migration are related to the extracellular scavenger receptor cysteine-rich domain (SRCR-D)-3 of CD6. Itolizumab, clone T1h, is a newly developed humanized IgG1 monoclonal antibody that targets CD6 SRCR-D1 and blocks immune activation. Itolizumab has been proposed to be effective in autoimmune diseases such as rheumatoid arthritis, Sjögren's syndrome and multiple sclerosis. In Sjögren's syndrome, the utilization of itolizumab as therapeutic option is reinforced by our recent observation that ALCAM, the CD6 ligand, is overexpressed and that CD6-positive T and B cells are detected within salivary glands from Sjögren's syndrome patients. In this study, itolizumab-positive target cells were characterized within both peripheral blood and salivary glands in order to provide rational for anti-CD6 treatment in Sjögren's syndrome