Secretory IgA mediates retrotranscytosis of intact gliadin peptides via the transferrin receptor in celiac disease

Celiac disease (CD) is an enteropathy resulting from an abnormal immune response to gluten-derived peptides in genetically susceptible individuals. This immune response is initiated by intestinal transport of intact peptide 31-49 (p31-49) and 33-mer gliadin peptides through an unknown mechanism. We show that the transferrin receptor CD71 is responsible for apical to basal retrotranscytosis of gliadin peptides, a process during which p31-49 and 33-mer peptides are protected from degradation. In patients with active CD, CD71 is overexpressed in the intestinal epithelium and colocalizes with immunoglobulin (Ig) A. Intestinal transport of intact p31-49 and 33-mer peptides was blocked by polymeric and secretory IgA (SIgA) and by soluble CD71 receptors, pointing to a role of SIgA–gliadin complexes in this abnormal intestinal transport. This retrotranscytosis of SIgA–gliadin complexes may promote the entry of harmful gliadin peptides into the intestinal mucosa, thereby triggering an immune response and perpetuating intestinal inflammation. Our findings strongly implicate CD71 in the pathogenesis of CD

T84-intestinal epithelial exosomes bear MHC class II/peptide complexes potentiating antigen presentation by dendritic cells: Function of intestinal epithelial exosomes

International audienceBackground and aims: Intestinal epithelial cells release antigen presenting vesicles (exosomes) bearing MHC class II/peptide complexes stimulating specific immune responses in vivo. To further characterize the role of human epithelial exosomes in antigen presentation, their capacity to load antigenic peptides, to bind immune target cells and to induce T cell activation was analyzed in vitro. Methods: The capacity of exosomes derived from the HLA-DR4 expressing, intestinal epithelial cell line T84, to load the HLA-DR4-specific peptide 3H-HSA 64-76 and to activate a HLA-DR4-restricted T cell hybridoma, was tested in the presence or absence of human monocyte-derived dendritic cells (DCs). Interaction of FITC-labeled exosomes with T cells and DCs was analyzed by flow cytometry and confocal microscopy. Results: T84-derived exosomes, enriched in CD9, CD81, CD82 and A33 antigen, were capable of binding specifically HSA 64-76 peptide on HLA-DR4 molecules and of interacting preferentially with DCs. HSA-loaded exosomes were unable to activate the T cell hybridoma directly, but induced a productive T cell activation through DCs. When HSA peptide was bound to exosomal HLA-DR4 molecules instead of in a soluble form, the threshold of peptide presentation by DCs was markedly decreased (x10-3). Conclusions: Exosomes released by intestinal epithelial cells bear exogenous peptides complexed to MHC class II molecules and interact preferentially with DCs, strongly potentiating peptide presentation to T cells. Epithelial exosomes constitute a powerful link between luminal antigens and local immune cells by mediating the transfer of tiny amounts of luminal antigenic information and facilitating immune surveillance at mucosal surfaces

Mechanisms Involved in Alleviation of Intestinal Inflammation by Bifidobacterium Breve Soluble Factors

Objectives: Soluble factors released by Bifidobacterium breve C50 (Bb) alleviate the secretion of pro-inflammatory cytokines by immune cells, but their effect on intestinal epithelium remains elusive. To decipher the mechanisms accounting for the cross-talk between bacteria/soluble factors and intestinal epithelium, we measured the capacity of the bacteria, its conditioned medium (Bb-CM) and other Gram(+) commensal bacteria to dampen inflammatory chemokine secretion. Methods: TNFa-induced chemokine (CXCL8) secretion and alteration of NF-kB and AP-1 signalling pathways by Bb were studied by EMSA, confocal microscopy and western blotting. Anti-inflammatory capacity was also tested in vivo in a model of TNBS-induced colitis in mice. Results: Bb and Bb-CM, but not other commensal bacteria, induced a time and dose-dependent inhibition of CXCL8 secretion by epithelial cells driven by both AP-1 and NF-kB transcription pathways and implying decreased phosphorylation of p38-MAPK and IkB-a molecules. In TNBS-induced colitis in mice, Bb-CM decreased the colitis score and inflammatory cytokine expression, an effect reproduced by dendritic cell conditioning with Bb-CM. Conclusions: Bb and secreted soluble factors contribute positively to intestinal homeostasis by attenuating chemokine production. The results indicate that Bb down regulate inflammation at the epithelial level by inhibiting phosphorylation

CARD15/NOD2 Is Required for Peyer's Patches Homeostasis in Mice

BACKGROUND: CARD15/NOD2 mutations are associated with susceptibility to Crohn's Disease (CD) and Graft Versus Host Disease (GVHD). CD and GVHD are suspected to be related with the dysfunction of Peyer's patches (PP) and isolated lymphoid follicles (LFs). Using a new mouse model invalidated for Card15/Nod2 (KO), we thus analysed the impact of the gene in these lymphoid formations together with the development of experimental colitis. METHODOLOGY/PRINCIPAL FINDINGS: At weeks 4, 12 and 52, the numbers of PPs and LFs were higher in KO mice while no difference was observed at birth. At weeks 4 and 12, the size and cellular composition of PPs were analysed by flow cytometry and immunohistochemistry. PPs of KO mice were larger with an increased proportion of M cells and CD4(+) T-cells. KO mice were also characterised by higher concentrations of TNFalpha, IFNgamma, IL12 and IL4 measured by ELISA. In contrast, little differences were found in the PP-free ileum and the spleen of KO mice. By using chamber experiments, we found that this PP phenotype is associated with an increased of both paracellular permeability and yeast/bacterial translocation. Finally, KO mice were more susceptible to the colitis induced by TNBS. CONCLUSIONS: Card15/Nod2 deficiency induces an abnormal development and function of the PPs characterised by an exaggerated immune response and an increased permeability. These observations provide a comprehensive link between the molecular defect and the Human CARD15/NOD2 associated disorders: CD and GVHD

Rôle fonctionnel des exosomes epithéliaux dans l immunité intestinale

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Etude in vivo du rétrotransport d'immum-complexes IgA observé dans la maladie coeliaque

La maladie coeliaque (MC) est une entéropathie apparentée aux maladies auto-immunes mais induite par l ingestion de protéines dérivées du blé (gliadines) chez des individus génétiquement prédisposés exprimant les molécules HLA-DQ2/8. Les lésions intestinales sont la conséquence d une activation anormale du système immunitaire intestinal. La présentation élective de certains peptides des gliadines par les molécules HLADQ2/8 induit l activation des lymphocytes T (LT CD4+) intestinaux. D autres peptides semblent capables de renforcer cette réponse immune en favorisant la production d IL15. Enfin, il existe chez les patients une translocation accrue de peptides immunogènes à travers l épithélium intestinal qui enclenche ou entretient cette réaction immunitaire. Le mécanisme de cette translocation reste discuté. Les travaux du laboratoire menés sur des biopsies intestinales suggèrent une transcytose entérocytaire de complexes immuns IgA/peptides assurée par CD71, le récepteur de la transferrine. Ce récepteur est capable de lier les IgA sécrétoires (SIgA). Il est anormalement exprimé à la surface des entérocytes chez les patients actifs, conduisant à un rétrotransport anormal des complexes immuns et à l entrée accrue de peptides immunogènes dans le chorion. L accès aux biopsies intestinales limite les études chez l homme. Mon travail de thèse a eu pour but de mettre au point un modèle murin pour analyser l effet de ce rétrotransport sur la réponse immune intestinale. Les études menées in vitro et ex vivo ont montré que le récepteur CD71 murin ne liait pas les IgA murines mais qu il liait les IgA1 polymériques humaines et des IgA chimériques où la chaîne a1 est humaine. En greffant chez des souris BALB/c un hybridome secrétant des IgA chimériques spécifiques de l ovalbumine (OVA), nous avons induit une production intestinale significative de SIgA humanisées anti-OVA. L expression apicale sur les entérocytes de CD71 a été ensuite induite grâce à un traitement intrapéritonéal par la tyrphostine A8. Chez les souris greffées et traitées, l OVA est transportée à travers l épithélium de fragments intestinaux montés en chambres d Ussing sous forme partiellement intacte (30%). En revanche, l OVA est dégradée (95%) chez les souris témoins non greffées ou non traitées. Ce transport protégé n est pas observé avec une protéine contrôle (HRP) ni chez des souris greffées avec un hybridome secrétant des IgA spécifiques d une autre protéine et traitées par la tyrphostine A8. Ces données indiquent que nous avons obtenu un modèle murin de rétro-transport d OVA dépendant des IgA et de CD71. Nous avons analysé les conséquences in vivo de ce rétro-transport sur la réponse immune chez des souris transgéniques DO11.10 dont les cellules T CD4+ expriment un récepteur T spécifique de l OVA (T-OVA). Chez les souris greffées avec l hybridome sécrétant des IgA anti-OVA et traitées par la tyrphostine A8, l administration orale d OVA induit une expression accrue du marqueur d activation CD69 sur les lymphocytes T-OVA des ganglions mésentériques comparé aux souris contrôles, ainsi qu une production accrue d IFN en réponse à la stimulation in vitro par l OVA. Nos résultats suggèrent que le rétro-transport épithélial des complexes IgA-gliadine par le récepteur CD71 pourrait participer à l activation anormale de la réponse T anti-gliadine chez les malades cœliaquesPARIS-BIUSJ-Biologie recherche (751052107) / SudocSudocFranceF

Probiotiques et fonctions lympho-épithéliales intestinales

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Caractérisation phénotypique et fonctionnelle des exosomes de cellules épithéliales intestinales

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Systèmes de transport membranaires, génétique et nutrition ; l'exemple des anomalies congénitales du transport intestinal chez l'enfant

International audienc

Impact of 10 Days of Intensive Exercise Period (mHealth Grand Tour) on Heart Rate Variability and Links with Glycaemic Excursions in Athletes with Type 1 Diabetes

Meeting Abstract P22Impact of 10 Days of Intensive Exercise Period (mHealth Grand Tour) on Heart Rate Variability and Links with Glycaemic Excursions in Athletes with Type 1 Diabete