Mast Cell Degranulation Exacerbates Skin Rejection by Enhancing Neutrophil Recruitment

Recent evidences indicate an important role of tissue inflammatory responses by innate immune cells in allograft acceptance and survival. Here we investigated the role of mast cells (MC) in an acute male to female skin allograft rejection model using red MC and basophil (RMB) mice enabling conditional MC depletion. Kinetic analysis showed that MCs markedly accelerate skin rejection. They induced an early inflammatory response through degranulation and boosted local synthesis of KC, MIP-2, and TNF. This enhanced early neutrophil infiltration compared to a female-female graft-associated repair response. The uncontrolled neutrophil influx accelerated rejection as antibody-mediated depletion of neutrophils delayed skin rejection. Administration of cromolyn, a MC stabilizer and to a lesser extent ketotifen, a histamine type I receptor antagonist, and absence of MCPT4 chymase also delayed graft rejection. Together our data indicate that mediators contained in secretory granules of MC promote an inflammatory response with enhanced neutrophil infiltration that accelerate graft rejection

Context-dependent effects of IL-2 rewire immunity into distinct cellular circuits

Interleukin 2 (IL-2) is a key homeostatic cytokine, with therapeutic applications in both immunogenic and tolerogenic immune modulation. Clinical use has been hampered by pleiotropic functionality and widespread receptor expression, with unexpected adverse events. Here, we developed a novel mouse strain to divert IL-2 production, allowing identification of contextual outcomes. Network analysis identified priority access for Tregs and a competitive fitness cost of IL-2 production among both Tregs and conventional CD4 T cells. CD8 T and NK cells, by contrast, exhibited a preference for autocrine IL-2 production. IL-2 sourced from dendritic cells amplified Tregs, whereas IL-2 produced by B cells induced two context-dependent circuits: dramatic expansion of CD8+ Tregs and ILC2 cells, the latter driving a downstream, IL-5–mediated, eosinophilic circuit. The source-specific effects demonstrate the contextual influence of IL-2 function and potentially explain adverse effects observed during clinical trials. Targeted IL-2 production therefore has the potential to amplify or quench particular circuits in the IL-2 network, based on clinical desirability

The role of the endosomal system in the function of Fc immunoglobulin receptors

Les Récepteurs aux fragments Fcs des immunoglobulines (RFcs) sont des récepteurs majeurs du système immunitaire, présents à la surface des cellules immunes et faisant l’interface entre l’immunité innée et l’immunité adaptative. Ils reconnaissent la partie Fc des immunoglobulines des complexes immuns (IC) constitués d’un antigène et d’une immunoglobuline spécifique déclenchant des voies de signalisation qui déterminent la production de médiateurs anti- ou pro-inflammatoires ainsi que d’autres réactions permettant l’élimination de l’infection. Après la liaison de la partie Fc des ICs, le récepteur et les ICs sont internalisés. Il est connu que cette internalisation permet la présentation de l’antigène contenu dans les ICs et l’élimination de pathogènes internalisés, mais le rôle de l’internalisation dans la transduction du signal via les RFcs n’avait pas été bien étudié auparavant. Nous avons étudié le rôle du système endosomal dans la fonction des RFcs et montré que le RFcγI, récepteur de haute affinité aux IgG, continue à signaler via des plates-formes de signalisation endosomale, ce qui serait un mécanisme essentiel pour l'activation complète des fonctions clés des RFcs. Les endosomes caractérisés par l’aminopeptidase insulinodépendante IRAP constituerait une plateforme de signalisation pour le RFcγI. De manière similaire, nous avons mis en évidence que la sous unité CD3ζ du TCR, récepteur au lymphocyte T, forme un pool intracellulaire dans les endosomes contenant IRAP et la syntaxine STX6. Nos résultats montrent que le TCR continue de signaler après l'endocytose et que cette signalisation intracellulaire est particulièrement importante pour l’activation de la cellule T par des complexes peptide-CMH de faible affinité.Immunoglobulin Fc receptors (FcR) are cell surface immune receptors at the interface between innate and adaptive immunity. They recognize the Fc part of the immunoglobulin of the immune complexes consisting of an antigen and a specific immunoglobulin. They trigger signaling pathways that determine the production of anti- or pro-inflammatory mediators and other reactions allowing the elimination of infection. After the binding of ICs, the receptor and the ICs are internalized. It is known that this internalization allows the presentation of the antigen contained in the ICs and the elimination of internalized pathogens, but the role of internalization in signal transduction via RFCs has not been well studied before.We investigated the role of the endosomal system in FcR function and we showed that the FcγR1, a high affinity IgG receptor, continues to signal via endosomal signaling platfrorms, which would be essential for the complete activation of the receptor functions. The endosomes characterized by insulin-dependent aminopeptidase IRAP constituted a signaling platform for RFcγI. Similarly, we have demonstrated that the CD3ζ subunit of the TCR, the lymphocyte T receptor, form an intracellular pool in the endosomes containing IRAP and the syntaxine STX6. Our results demonstrate that the TCR continues to signal after endocytosis and that this intracellular signaling is particularly important for T cell activation by low affinity peptide-MHC complexes

Development of bioartificial myocardium by electrostimulation of 3D collagen scaffolds seeded with stem cells

Electrostimulation (ES) can be defined as a safe physical method to induce stem cell differentiation. The aim of this study is to evaluate the effectiveness of ES on bone marrow mesenchymal stem cells (BMSCs) seeded in collagen scaffolds in terms of proliferation and differentiation into cardiomyocytes. BMSCs were isolated from Wistar rats and seeded into 3D collagen type 1 templates measuring 25 x 25 x 6 mm. Bipolar <em>in vitro</em> ES was performed during 21 days. Electrical impedance and cell proliferation were measured. Expression of cardiac markers was assessed by immunocytochemistry. Viscoelasticity of collagen matrix was evaluated. Electrical impedance assessments showed a low resistance of 234±41 Ohms which indicates good electrical conductivity of collagen matrix. Cell proliferation at 570 nm as significantly increased in ES groups after seven day (ES 0.129±0.03 <em>vs</em> non-stimulated control matrix 0.06±0.01, P=0.002) and after 21 days, (ES 0.22±0.04 <em>vs</em> control 0.13±0.01, P=0.01). Immunocytochemistry of BMSCs after 21 days ES showed positive staining of cardiac markers, troponin I, connexin 43, sarcomeric alpha-actinin, slow myosin, fast myosin and desmin. Staining for BMSCs marker CD29 after 21 days was negative. Electrostimulation of cell-seeded collagen matrix changed stem cell morphology and bio- chemical characteristics, increasing the expression of cardiac markers. Thus, MSC-derived differentiated cells by electrostimulation grafted in biological scaffolds might result in a convenient tissue engineering source for myocardial diseases

The In Vitro Contribution of Autolysins to Bacterial Killing Elicited by Amoxicillin Increases with Inoculum Size in Enterococcus faecalis▿ †

The mechanisms of antibiotic-induced cell death are poorly understood despite the critical role of the bactericidal activities of antibiotics for successful treatment of severe infections. These mechanisms include irreversible damaging of macromolecules by reactive oxygen species and bacteriolysis mediated by peptidoglycan hydrolases (autolysins). We have assessed the contribution of the second mechanism by using an autolysin-deficient mutant of Enterococcus faecalis and shown that it contributes to amoxicillin-induced cell lysis only at a high bacterial density

P-selectin targeting polysaccharide-based nanogels for miRNA delivery

International audienc

Activating FcγR function depends on endosomal-signaling platforms

Summary: Cell surface receptor internalization can either terminate signaling or activate alternative endosomal signaling pathways. We investigated here whether endosomal signaling is involved in the function of the human receptors for Fc immunoglobulin fragments (FcRs): FcαRI, FcγRIIA, and FcγRI. All these receptors were internalized after their cross-linking with receptor-specific antibodies, but their intracellular trafficking was different. FcαRI was targeted directly to lysosomes, while FcγRIIA and FcγRI were internalized in particular endosomal compartments described by the insulin responsive aminopeptidase (IRAP), where they recruited signaling molecules, such as the active form of the kinase Syk, PLCγ and the adaptor LAT. Destabilization of FcγR endosomal signaling in the absence of IRAP compromised cytokine secretion downstream FcγR activation and macrophage ability to kill tumor cells by antibody-dependent cell-mediated cytotoxicity (ADCC). Our results indicate that FcγR endosomal signaling is required for the FcγR-driven inflammatory reaction and possibly for the therapeutic action of monoclonal antibodies

Mast Cell Degranulation Exacerbates Skin Rejection by Enhancing Neutrophil Recruitment

Recent evidences indicate an important role of tissue inflammatory responses by innate immune cells in allograft acceptance and survival. Here we investigated the role of mast cells (MC) in an acute male to female skin allograft rejection model using red MC and basophil (RMB) mice enabling conditional MC depletion. Kinetic analysis showed that MCs markedly accelerate skin rejection. They induced an early inflammatory response through degranulation and boosted local synthesis of KC, MIP-2, and TNF. This enhanced early neutrophil infiltration compared to a female-female graft-associated repair response. The uncontrolled neutrophil influx accelerated rejection as antibody-mediated depletion of neutrophils delayed skin rejection. Administration of cromolyn, a MC stabilizer and to a lesser extent ketotifen, a histamine type I receptor antagonist, and absence of MCPT4 chymase also delayed graft rejection. Together our data indicate that mediators contained in secretory granules of MC promote an inflammatory response with enhanced neutrophil infiltration that accelerate graft rejection

CD31 signaling promotes the detachment at the uropod of extravasating neutrophils allowing their migration to sites of inflammation

Effective neutrophil migration to sites of inflammation is crucial for host immunity. A coordinated cascade of steps allows intravascular leukocytes to counteract the shear stress, transmigrate through the endothelial layer, and move toward the extravascular, static environment. Those events are tightly orchestrated by integrins, but, while the molecular mechanisms leading to their activation have been characterized, the regulatory pathways promoting their detachment remain elusive. In light of this, it has long been known that platelet-endothelial cell adhesion molecule (Pecam1, also known as CD31) deficiency blocks leukocyte transmigration at the level of the outer vessel wall, yet the associated cellular defects are controversial. In this study, we combined an unbiased proteomic study with in vitro and in vivo single-cell tracking in mice to study the dynamics and role of CD31 during neutrophil migration. We found that CD31 localizes to the uropod of migrating neutrophils along with closed β2-integrin and is required for essential neutrophil actin/integrin polarization. Accordingly, the uropod of Pecam1-/- neutrophils is unable to detach from the extracellular matrix, while antagonizing integrin binding to extracellular matrix components rescues this in vivo migratory defect. Conversely, we showed that sustaining CD31 co-signaling actively favors uropod detachment and effective migration of extravasated neutrophils to sites of inflammation in vivo. Altogether, our results suggest that CD31 acts as a molecular rheostat controlling integrin-mediated adhesion at the uropod of egressed neutrophils, thereby triggering their detachment from the outer vessel wall to reach the inflammatory sites