Rôle de la voie Wnt/β-catenine dans le mécanisme anti-inflammatoire employant une thérapie d’immunoglobulines normales

Les immunoglobulines polyclonales intraveineuses (IVIG) sont préparées à partir de plasmas provenant de plusieurs milliers de donneurs sains et utilisées comme traitement dans de nombreuses maladies inflammatoires et autoimmunes. Lors de ma thèse, j’ai investigué si cette thérapie pouvait interférer avec la détection sérique du virus Zika chez des patients atteints du syndrome de Guillain-Barré (GBS). J’ai démontré que la thérapie par IVIG n’interférait pas avec la détection sérique du virus dans le plasma des patients atteints de GBS suivant un traitement aux IVIG. Contrairement aux souris, les IVIG peuvent activer les basophiles humains par une voie différente que celle de l’IL-33. Les IVIG induisent la sécrétion d’Il-4, IL-6 et IL-8 par interaction directe avec les IgE à la surface des basophiles. Cette fonction est dépendante de la fraction F(ab’)2 et implique l’activation de Syk. Ces résultats montrent un nouveau mécanisme dans l’activation des basophiles humains par les IVIG. La dernière partie de ma thèse m’a permis d’étudier le rôle de la voie de signalisation β-caténine sur les effets anti-inflammatoires médiées pars les IVIG. La β-caténine, composante de la voie Wnt, joue un rôle important dans la tolérogénicité des cellules dendritiques (DC) et dans la protection contre l’encéphalomyélite auto-immune expérimentale (EAE). Les données générées montrent que les IVIG activent la voie β-caténine chez les DC humains en plus de la production de Wnt 5a nécessitant une IgG complète ainsi que les co-récepteurs LRP5/6. En dépit de l’induction de β-caténine par les IVIG, cette voie est dispensable pour ses actions anti-inflammatoires in vitro et in vivo dans le modèle EAE.Intravenous immunoglobulin (IVIG) is a therapeutic preparation of pooled normal IgG obtained from the several thousand healthy donors. It is established as first-line therapy for many autoimmune and inflammatory diseases. In the first part of my thesis, I have investigated if IVIG therapy interferes with the serological detection of Zika virus infection in Guillain–Barré syndrome (GBS) patients. By analyzing the plasma of GBS patients treated with IVIG for anti-Zika IgG, I have demonstrated that IVIG therapy in GBS patients does not interfere with the serological Zika detection. The second part addresses the immunoregulatory role of IVIG on human basophil function. Unlike in mice, IVIG does not require DC-SIGN-dependent IL-33 for the activation of human basophils. IVIG directly induces the activation of IL-3-primed human basophils and secretion of IL-4, IL-6, and IL-8 by directly interacting with the basophil surface-bound IgE. This function was F(ab’)2-dependent and involves Syk activation. These results demonstrate a novel mechanism of human basophil activation by IVIG. The last part unravels the signaling pathways associated with IVIG-mediated anti-inflammatory effects specifically the Wnt/β-catenin pathway, which imparts tolerogenic properties to dendritic cells (DCs) and protection against experimental autoimmune encephalomyelitis (EAE). My data shows that IVIG activates β-catenin in human DC along with upregulation of Wnt 5a. Activation of β-catenin requires intact IgG and LRP5/6 co-receptors. However, despite the activation of β-catenin by IVIG, this pathway is dispensable for its anti-inflammatory actions both in vitro and in vivo in the EAE model

Sonic hedgehog-responsive lipoxygenases and cyclooxygenase-2 modulate Dectin-1-induced inflammatory cytokines

Immune responses during fungal infections are predominately mediated by 5/15-lipoxygenases (LO)-or cyclooxygenase (COX)-2-catalysed bioactive eicosanoid metabolites like leukotrienes, lipoxins and prostaglandins. Although few host mediators of fungi-triggered eicosanoid production have been established, the molecular mechanism of expression and regulation of 5-LO, 15-LO and COX-2 are not well-defined. Here, we demonstrate that, macrophages infected with representative fungi Candida albicans, Aspergillus flavus or Aspergillus fumigatus or those treated with Curdlan, a selective agonist of pattern recognition receptor for fungi Dectin-1, displays increased expression of 5-LO, 15-LO and COX-2. Interestingly, Dectin-1-responsive Syk pathway activates mTOR-sonic hedgehog (SHH) signaling cascade to stimulate the expression of these lipid metabolizing enzymes. Loss-of-function analysis of the identified intermediaries indicates that while Syk-mTOR-SHH pathway-induced 5-LO and 15-LO suppressed the Dectin-l-responsive pro-inflammatory signature cytokines like TNE-alpha, IL-1 beta and IL-12, Syk-mTOR-SHH-induced COX-2 positively regulated these cytokines. Dectin-1-stimulated IL-6, however, is dependent on 5-LO, 15-LO and COX-2 activity. Together, the current study establishes Dectin-1-arbitrated host mediators that direct the differential regulation of immune responses during fungal infections and thus are potential candidates of therapeutic intervention. (C) 2015 Elsevier Ltd. All rights reserved

Vaccine-induced immune thrombotic thrombocytopenia: Consider IVIG batch in the treatment

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Wuchereria bancrofti filaria activates human dendritic cells and polarizes T helper 1 and regulatory T cells via toll-like receptor 4

International audienceInteraction between innate immune cells and parasite plays a key role in the immunopathogenesis of lymphatic filariasis. Despite being professional antigen presenting cells critical for the pathogen recognition, processing and presenting the antigens for mounting T cell responses, the dendritic cell response and its role in initiating CD4+ T cell response to filaria, in particular Wuchereria bancrofti, the most prevalent microfilaria is still not clear. Herein, we demonstrate that a 70 kDa phosphorylcholine-binding W. bancrofti sheath antigen induces human dendritic cell maturation and secretion of several pro-inflammatory cytokines. Further, microfilarial sheath antigen-stimulated dendritic cells drive predominantly Th1 and regulatory T cell responses while Th17 and Th2 responses are marginal. Mechanistically, sheath antigen-induced dendritic cell maturation, and Th1 and regulatory T cell responses are mediated via toll-like receptor 4 signaling. Our data suggest that W. bancrofti sheath antigen exploits dendritic cells to mediate distinct CD4+ T cell responses and immunopathogenesis of lymphatic filariasis

Acid Stripping of Surface IgE Antibodies Bound to FcεRI Is Unsuitable for the Functional Assays That Require Long-Term Culture of Basophils and Entire Removal of Surface IgE

International audienceBasophils are rare granulocytes and dysregulated functions of these cells are associated with several atopic and non-atopic allergic diseases of skin, respiratory system and gastrointestinal tract. Both cytokines and immunoglobulin E (IgE) are implicated in mediating the basophil activation and pathogenesis of these disorders. Several reports have shown that healthy individuals, and patients with allergic disorders display IgG autoantibodies to IgE and hence functional characterization of these anti-IgE IgG autoantibodies is critical. In general, anti-IgE IgG autoantibodies modulate basophil activation irrespective of allergen specificity by interacting with constant domains of IgE. Therefore, an ideal solution to prove the functions of such anti-IgE IgG autoantibodies would be to completely eliminate type I high affinity immunoglobulin E receptor (FcɛRI)-bound IgE from the surface of basophils and to demonstrate in an unequivocal manner the role of anti-IgE IgG autoantibodies. In line with previous reports, our data show that FcɛRI on peripheral blood basophils are almost saturated with IgE. Further, acetic acid buffer (pH 4) efficiently removes these FcɛRI-bound IgE. Although immediately following acetic acid-elution of IgE had no repercussion on the viability of basophils, following 24 hours culture with interleukin-3 (IL-3), the viability and yield of basophils were drastically reduced in acid-treated cells and had repercussion on the induction of activation markers. Lactic acid treatment on the other hand though had no adverse effects on the viability of basophils and IL-3-induced activation, it removed only a small fraction of the cell surface bound IgE. Thus, our results show that acid buffers could be used for the elution of FcɛRI-bound IgE on the basophil surface for the biochemical characterization of IgE antibodies or for the immediate use of basophils to determine their sensitivity to undergo degranulation by specific allergens. However, these methods are not utile for the functional assays of basophils that require longer duration of culture and entire removal of surface IgE to validate the role of anti-IgE IgG autoantibodies that interact with FcɛRI-bound IgE irrespective of allergen specificity

Potential of regulatory T cell-based therapies in the management of severe COVID-19

Open accessInternational audienceIn view of dysregulated immune response, cytokine storm and inflammation-induced severe lung damage in severely ill COVID-19 patients, we propose that CD4+CD25+FoxP3+ regulatory T cell-based therapies could be considered for the patient managemen

Human basophils may not undergo modulation by DC-SIGN and mannose receptor–targeting immunotherapies due to absence of receptors

Comment inReply. [J Allergy Clin Immunol. 2017]Comment onNovel vaccines targeting dendritic cells by coupling allergoids to nonoxidized mannan enhance allergen uptake and induce functional regulatory T cells through programmed death ligand 1. [J Allergy Clin Immunol. 2016]International audienceSirvent et al1 recently showed that novel vaccines targeting dendritic cells (DCs) by coupling glutaraldehyde-polymerized grass pollen allergoids to nonoxidized mannan enhance allergen uptake and induce functional regulatory T cells through programmed death ligand 1. Mechanistically, they found that nonoxidized mannan-coupled glutaraldehyde-polymerized grass pollen allergoids are captured and internalized by 2 lectin receptors on DCs: mannose receptor (CD206) and DC-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN or CD209). These data thus indicated that DCs could be targeted by C-type lectin receptors for efficient allergen immunotherapy

Wnt-β-Catenin Signaling in Human Dendritic Cells Mediates Regulatory T-Cell Responses to Fungi via the PD-L1 Pathway

International audienceThe signaling pathways activated following interaction between dendritic cells (DCs) and a pathogen determine the polarization of effector T-cell and regulatory T-cell (Treg) responses to the infection. Several recent studies, mostly in the context of bacterial infections, have shown that the Wnt/b-catenin pathway plays a major role in imparting tolerogenic features in DCs and in promotion of Treg responses. However, the significance of the Wnt/b-catenin pathway's involvement in regulating the immune response to the fungal species is not known. Using Aspergillus fumigatus, a ubiquitous airborne opportunistic fungal species, we show here that fungi activate the Wnt/b-catenin pathway in human DCs and are critical for mediating the immunosuppressive Treg responses. Pharmacological inhibition of this pathway in DCs led to inhibition of maturation-associated molecules and interleukin 10 (IL-10) secretion without affecting the majority of the inflammatory cytokines. Furthermore, blockade of Wnt signaling in DCs suppressed DC-mediated Treg responses in CD4 1 T cells and downregulated both tumor necrosis factor alpha (TNF-a) and IL-10 responses in CD8 1 T cells. Mechanistically, induction of b-catenin pathway by A. fumigatus required C-type lectin receptors and promoted Treg polarization via the induction of programmed death-ligand 1 on DCs. Further investigation on the identity of fungal molecular patterns has revealed that the cell wall polysaccharides b-(1, 3)-glucan and a-(1, 3)-glucan, but not chitin, possess the capacity to activate the b-catenin pathway. Our data suggest that the Wnt/b-catenin pathway is a potential therapeutic target to selectively suppress the Treg response and to sustain the protective Th1 response in the context of invasive aspergillosis caused by A. fumigatus

Regulatory T cells induce activation rather than suppression of human basophils

International audienceBasophils are a rare granulocyte population that have been associated with allergic and inflammatory responses. It is essential to understand the regulatory mechanisms by which basophils are kept in check, considering the impact of dysregulated basophil function on immune responses under different pathological conditions. Among immunoregulatory cells, CD4+CD25+FoxP3+ regulatory T cells (Tregs) are the key players that maintain immune tolerance. The mechanisms by which Tregs regulate and suppress diverse immune cell subsets have been studied extensively, but the impact of Tregs on basophil functions is not well understood. We report that human basophils are refractory to Treg-mediated suppression and found that Tregs stimulate resting basophils to induce the expression of activation markers including CD69, CD203c and CD13, and the release of basophil cytokines including IL-13, IL-8 and IL-4. Mechanistically, Tregs could induce human basophil activation via IL-3 and STAT5 activation, whereas cellular contact was dispensable. Inhibition of either IL-3-IL-3 receptor interactions or STAT5 phosphorylation abrogated Treg-mediated activation of basophils. These results provide evidence of direct positive effects that human Tregs have on basophil activation and reveal a previously unrecognized feature of this cell subset well known for immunosuppressive functions

Therapeutic normal IgG intravenous immunoglobulin activates Wnt-β-catenin pathway in dendritic cells

International audienceTherapeutic normal IgG intravenous immunoglobulin (IVIG) is a well-established first-line immunotherapy for many autoimmune and inflammatory diseases. Though several mechanisms have been proposed for the anti-inflammatory actions of IVIG, associated signaling pathways are not well studied. As β-catenin, the central component of the canonical Wnt pathway, plays an important role in imparting tolerogenic properties to dendritic cells (DCs) and in reducing inflammation, we explored whether IVIG induces the β-catenin pathway to exert anti-inflammatory effects. We show that IVIG in an IgG-sialylation independent manner activates β-catenin in human DCs along with upregulation of Wnt5a secretion. Mechanistically, β-catenin activation by IVIG requires intact IgG and LRP5/6 co-receptors, but FcγRIIA and Syk are not implicated. Despite induction of β-catenin, this pathway is dispensable for anti-inflammatory actions of IVIG in vitro and for mediating the protection against experimental autoimmune encephalomyelitis in vivo in mice, and reciprocal regulation of effector Th17/Th1 and regulatory T cells