Celiac disease (CeD) manifests with autoimmune intestinal inflammation from gluten and genetic predisposition linked to human leukocyte antigen class-II (HLA-II) gene variants. Antigen-presenting cells facilitate gluten exposition through the interaction of their surface major histocompatibility complex (MHC) with the T cell receptor (TCR) on T lymphocytes. This fundamental mechanism of adaptive immunity has broadened upon recognition of extracellular exosomal MHC, raising awareness of an alternative means for antigen presentation. This study demonstrates that conditioned growth media (CGM) previously exposed to monocyte-derived dendritic cells from CeD significantly downregulates the CD3+ lineage marker of control T cells. Such increased activation was reflected in their elevated IL-2 secretion. Exosome localization motif identification and quantification within HLA-DQA1 and HLA-DQB1 transcripts highlighted their significant prevalence within HLA-DQB1 alleles associated with CeD susceptibility. Flow cytometry revealed the strong correlation between HLA-DQ and the CD63 exosomal marker in T cells exposed to CGM from MoDCs sourced from CeD patients. This resulted in lower concentrations of CD25+ CD127- T cells, suggestive of their compromised induction to T-regulatory cells associated with CeD homeostasis. This foremost comparative study deciphered the genomic basis and extracellular exosomal effects of HLA transfer on T lymphocytes in the context of CeD, offering greater insight into this auto-immune disease.

• Keywords
• autoimmunity, exosome, gluten, major histocompatibility complex II, monocyte-derived dendritic cells,
• MeSH
• Alleles MeSH
• Celiac Disease * MeSH
• Glutens genetics   MeSH
• HLA-DQ Antigens genetics   MeSH
• Humans MeSH
• T-Lymphocytes, Regulatory MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• Glutens MeSH
• HLA-DQ Antigens MeSH

The autoimmune condition, Celiac Disease (CeD), displays broad clinical symptoms due to gluten exposure. Its genetic association with DQ variants in the human leukocyte antigen (HLA) system has been recognised. Monocyte-derived mature dendritic cells (MoDCs) present gluten peptides through HLA-DQ and co-stimulatory molecules to T lymphocytes, eliciting a cytokine-rich microenvironment. Having access to CeD associated families prevalent in the Czech Republic, this study utilised an in vitro model to investigate their differential monocyte profile. The higher monocyte yields isolated from PBMCs of CeD patients versus control individuals also reflected the greater proportion of dendritic cells derived from these sources following lipopolysaccharide (LPS)/ peptic-tryptic-gliadin (PTG) fragment stimulation. Cell surface markers of CeD monocytes and MoDCs were subsequently profiled. This foremost study identified a novel bio-profile characterised by elevated CD64 and reduced CD33 levels, unique to CD14++ monocytes of CeD patients. Normalisation to LPS stimulation revealed the increased sensitivity of CeD-MoDCs to PTG, as shown by CD86 and HLA-DQ flow cytometric readouts. Enhanced CD86 and HLA-DQ expression in CeD-MoDCs were revealed by confocal microscopy. Analysis highlighted their dominance at the CeD-MoDC membrane in comparison to controls, reflective of superior antigen presentation ability. In conclusion, this investigative study deciphered the monocytes and MoDCs of CeD patients with the identification of a novel bio-profile marker of potential diagnostic value for clinical interpretation. Herein, the characterisation of CD86 and HLA-DQ as activators to stimulants, along with robust membrane assembly reflective of efficient antigen presentation, offers CeD targeted therapeutic avenues worth further exploration.

• Keywords
• CD33, CD64, CD86, MHCDQ, autoimmunity, major histocompatibility complex II, monocyte, monocyte-derived dendritic cells,
• MeSH
• Autoimmune Diseases immunology   MeSH
• Biomarkers metabolism   MeSH
• Cell Membrane metabolism   MeSH
• Antigens, CD metabolism   MeSH
• Celiac Disease epidemiology   immunology   MeSH
• T-Lymphocytes, Cytotoxic immunology   MeSH
• Dendritic Cells immunology   MeSH
• Adult MeSH
• Gliadin adverse effects   MeSH
• HLA-DQ Antigens metabolism   MeSH
• Middle Aged MeSH
• Humans MeSH
• Lipopolysaccharides MeSH
• Adolescent MeSH
• Young Adult MeSH
• Monocytes metabolism   MeSH
• Disease Susceptibility MeSH
• Antigen Presentation immunology   MeSH
• Family MeSH
• Pedigree MeSH
• Aged MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Adolescent MeSH
• Young Adult MeSH
• Male MeSH
• Aged MeSH
• Female MeSH
• Publication type
• Journal Article MeSH
• Geographicals
• Czech Republic epidemiology   MeSH
• Names of Substances
• Biomarkers MeSH
• Antigens, CD MeSH
• Gliadin MeSH
• HLA-DQ Antigens MeSH
• Lipopolysaccharides MeSH

Tolerogenic dendritic cells (tDCs) may offer an intervention therapy in autoimmune diseases or transplantation. Stable immaturity and tolerogenic function of tDCs after encountering inflammatory environment are prerequisite for positive outcome of immunotherapy. However, the signaling pathways regulating their stable tolerogenic properties are largely unknown. In this study, we demonstrated that human monocyte-derived tDCs established by using paricalcitol (analogue of vitamin D2), dexamethasone and monophosphoryl lipid A exposed for 24h to LPS, cytokine cocktail, polyI:C or CD40L preserved reduced expression of co-stimulatory molecules, increased levels of inhibitory molecules ILT-3, PDL-1 and TIM-3, increased TLR-2, increased secretion of IL-10 and TGF-β, reduced IL-12 and TNF-α secretion and reduced T cell stimulatory capacity. tDCs further induced IL-10-producing T regulatory cells that suppressed the proliferation of responder T cells. In the inflammatory environment, tDCs maintained up-regulated indoleamine 2, 3 dioxygenase but abrogated IκB-α phosphorylation and reduced transcriptional activity of p65/RelA, RelB and c-Rel NF-κB subunits except p50. Mechanistically, p38 MAPK, ERK1/2, mTOR, STAT3 and mTOR-dependent glycolysis regulated expression of ILT-3, PDL-1 and CD86, secretion of IL-10 and T cell stimulatory capacity of tDCs in the inflammatory environment. Stability of tDCs in the inflammatory environment is thus regulated by multiple signaling pathways.

• Keywords
• Immunology and Microbiology Section, activation pathways, glycolysis, immune response, immunity, immunoregulation, stability, tolerogenic DCs,
• MeSH
• Cell Differentiation physiology   MeSH
• Dendritic Cells drug effects   metabolism   MeSH
• Dexamethasone pharmacology   MeSH
• Ergocalciferols pharmacology   MeSH
• Glycolysis drug effects   MeSH
• Cells, Cultured MeSH
• Humans MeSH
• Mitogen-Activated Protein Kinases metabolism   MeSH
• NF-kappa B metabolism   MeSH
• Signal Transduction MeSH
• TOR Serine-Threonine Kinases metabolism   MeSH
• STAT3 Transcription Factor metabolism   MeSH
• Inflammation metabolism   pathology   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Dexamethasone MeSH
• Ergocalciferols MeSH
• Mitogen-Activated Protein Kinases MeSH
• MTOR protein, human MeSH Browser
• NF-kappa B MeSH
• paricalcitol MeSH Browser
• STAT3 protein, human MeSH Browser
• TOR Serine-Threonine Kinases MeSH
• STAT3 Transcription Factor MeSH

Induction of long-term tolerance to β-cell autoantigens has been investigated both in animal models and in human type 1 diabetes (T1D) in order to prevent the disease. As regards external compounds, the dietary plant protein fraction has been associated with high penetrance of the disease, whereas gluten-free diets prevent T1D in animal models. Herewith we investigated whether intranasal (i.n.) administration of gliadin or gluten may arrest the diabetogenic process. I.n. administration of gliadin to 4-week-old NOD mice significantly reduced the diabetes incidence. Similarly, the insulitis was lowered. Intranasal gliadin also rescued a fraction of prediabetic 13-week-old NOD mice from progressing to clinical onset of diabetes compared to OVA-treated controls. Vaccination with i.n. gliadin led to an induction of CD4(+)Foxp3(+) T cells and even more significant induction of γδ T cells in mucosal, but not in non-mucosal lymphoid compartments. This prevention strategy was characterized by an increased proportion of IL-10 and a decreased proportion of IL-2, IL-4 and IFN-γ-positive CD4(+)Foxp3(+) T cells, and IFN-γ-positive γδ T cells, preferentially in mucosal lymphoid organs. In conclusion, i.n. vaccination with gliadin, an environmental antigen with possible etiological influence in T1D, may represent a novel, safer strategy for prevention or even early cure of T1D.

• MeSH
• Administration, Intranasal MeSH
• CD4-Positive T-Lymphocytes immunology   MeSH
• Cytokines metabolism   MeSH
• Diabetes Mellitus, Type 1 drug therapy   immunology   prevention & control   MeSH
• Forkhead Transcription Factors metabolism   MeSH
• Gliadin administration & dosage   therapeutic use   MeSH
• Glutens administration & dosage   MeSH
• Humans MeSH
• Lymphoid Tissue immunology   pathology   MeSH
• Mice, Inbred NOD MeSH
• Lymphocyte Count MeSH
• Immunity, Mucosal MeSH
• Animals MeSH
• Check Tag
• Humans MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Cytokines MeSH
• Forkhead Transcription Factors MeSH
• FOXP3 protein, human MeSH Browser
• Gliadin MeSH
• Glutens MeSH

Celiac disease (CD) is a gluten-responsive, chronic inflammatory enteropathy. IL-1 cytokine family members IL-1β and IL-18 have been associated with the inflammatory conditions in CD patients. However, the mechanisms of IL-1 molecule activation in CD have not yet been elucidated. We show in this study that peripheral blood mononuclear cells (PBMC) and monocytes from celiac patients responded to pepsin digest of wheat gliadin fraction (PDWGF) by a robust secretion of IL-1β and IL-1α and a slightly elevated production of IL-18. The analysis of the upstream mechanisms underlying PDWGF-induced IL-1β production in celiac PBMC show that PDWGF-induced de novo pro-IL-1β synthesis, followed by a caspase-1 dependent processing and the secretion of mature IL-1β. This was promoted by K+ efflux and oxidative stress, and was independent of P2X7 receptor signaling. The PDWGF-induced IL-1β release was dependent on Nod-like receptor family containing pyrin domain 3 (NLRP3) and apoptosis-associated speck like protein (ASC) as shown by stimulation of bone marrow derived dendritic cells (BMDC) from NLRP3(-/-) and ASC(-/-) knockout mice. Moreover, treatment of human PBMC as well as MyD88(-/-) and Toll-interleukin-1 receptor domain-containing adaptor-inducing interferon-β (TRIF)(-/-) BMDC illustrated that prior to the activation of caspase-1, the PDWGF-triggered signal constitutes the activation of the MyD88/TRIF/MAPK/NF-κB pathway. Moreover, our results indicate that the combined action of TLR2 and TLR4 may be required for optimal induction of IL-1β in response to PDWGF. Thus, innate immune pathways, such as TLR2/4/MyD88/TRIF/MAPK/NF-κB and an NLRP3 inflammasome activation are involved in wheat proteins signaling and may play an important role in the pathogenesis of CD.

• MeSH
• Adaptor Proteins, Vesicular Transport genetics   immunology   MeSH
• Celiac Disease MeSH
• Adult MeSH
• Gliadin chemistry   immunology   MeSH
• Inflammasomes drug effects   genetics   immunology   MeSH
• Interleukin-1beta genetics   immunology   MeSH
• Leukocytes, Mononuclear drug effects   immunology   pathology   MeSH
• Middle Aged MeSH
• Humans MeSH
• Mitogen-Activated Protein Kinases genetics   immunology   MeSH
• Myeloid Differentiation Factor 88 genetics   immunology   MeSH
• Mice, Knockout MeSH
• Mice MeSH
• Pepsin A MeSH
• Peptide Fragments pharmacology   MeSH
• Primary Cell Culture MeSH
• NLR Family, Pyrin Domain-Containing 3 Protein MeSH
• Gene Expression Regulation drug effects   immunology   MeSH
• Signal Transduction drug effects   genetics   immunology   MeSH
• Toll-Like Receptor 2 genetics   immunology   MeSH
• Toll-Like Receptor 4 genetics   immunology   MeSH
• Carrier Proteins genetics   immunology   MeSH
• Animals MeSH
• Check Tag
• Adult MeSH
• Middle Aged MeSH
• Humans MeSH
• Male MeSH
• Mice MeSH
• Female MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Adaptor Proteins, Vesicular Transport MeSH
• Gliadin MeSH
• Inflammasomes MeSH
• Interleukin-1beta MeSH
• Mitogen-Activated Protein Kinases MeSH
• MYD88 protein, human MeSH Browser
• Myeloid Differentiation Factor 88 MeSH
• NLRP3 protein, human MeSH Browser
• Pepsin A MeSH
• Peptide Fragments MeSH
• NLR Family, Pyrin Domain-Containing 3 Protein MeSH
• TICAM1 protein, human MeSH Browser
• TLR2 protein, human MeSH Browser
• TLR4 protein, human MeSH Browser
• Toll-Like Receptor 2 MeSH
• Toll-Like Receptor 4 MeSH
• Carrier Proteins MeSH

In genetically predisposed individuals, ingestion of wheat gliadin provokes a T-cell-mediated enteropathy, celiac disease. Gliadin fragments were previously reported to induce phenotypic maturation and Th1 cytokine production by human dendritic cells (DCs) and to boost their capacity to stimulate allogeneic T cells. Here, we monitor the effects of gliadin on migratory capacities of DCs. Using transwell assays, we show that gliadin peptic digest stimulates migration of human DCs and their chemotactic responsiveness to the lymph node-homing chemokines CCL19 and CCL21. The gliadin-induced migration is accompanied by extensive alterations of the cytoskeletal organization, with dissolution of adhesion structures, podosomes, as well as up-regulation of the CC chemokine receptor (CCR) 7 on cell surface and induction of cyclooxygenase (COX)-2 enzyme that mediates prostaglandin E2 (PGE₂) production. Blocking experiments confirmed that gliadin-induced migration is independent of the TLR4 signalling. Moreover, we showed that the α-gliadin-derived 31-43 peptide is an active migration-inducing component of the digest. The migration promoted by gliadin fragments or the 31-43 peptide required activation of p38 mitogen-activated protein kinase (MAPK). As revealed using p38 MAPK inhibitor SB203580, this was responsible for DC cytoskeletal transition, CCR7 up-regulation and PGE₂ production in particular. Taken together, this study provides a new insight into pathogenic features of gliadin fragments by demonstrating their ability to promote DC migration, which is a prerequisite for efficient priming of naive T cells, contributing to celiac disease pathology.

• MeSH
• Enzyme Activation drug effects   MeSH
• Models, Biological MeSH
• Chemokine CCL19 pharmacology   MeSH
• Chemokine CCL21 pharmacology   MeSH
• Chemotaxis drug effects   MeSH
• Cyclooxygenase 2 metabolism   MeSH
• Cytoskeleton drug effects   metabolism   MeSH
• Dendritic Cells cytology   drug effects   enzymology   MeSH
• Dinoprostone biosynthesis   MeSH
• Gliadin pharmacology   MeSH
• Humans MeSH
• MAP Kinase Signaling System drug effects   MeSH
• p38 Mitogen-Activated Protein Kinases metabolism   MeSH
• Peptide Fragments pharmacology   MeSH
• Receptors, CCR7 metabolism   MeSH
• Up-Regulation drug effects   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• CCR7 protein, human MeSH Browser
• Chemokine CCL19 MeSH
• Chemokine CCL21 MeSH
• Cyclooxygenase 2 MeSH
• Dinoprostone MeSH
• Gliadin MeSH
• p38 Mitogen-Activated Protein Kinases MeSH
• Peptide Fragments MeSH
• PTGS2 protein, human MeSH Browser
• Receptors, CCR7 MeSH

BACKGROUND AND AIMS: Celiac disease (CD) is a chronic inflammatory disorder of the small intestine that is induced by dietary wheat gluten proteins (gliadins) in genetically predisposed individuals. The overgrowth of potentially pathogenic bacteria and infections has been suggested to contribute to CD pathogenesis. We aimed to study the effects of gliadin and various intestinal bacterial strains on mucosal barrier integrity, gliadin translocation, and cytokine production. METHODOLOGY/PRINCIPAL FINDINGS: Changes in gut mucosa were assessed in the intestinal loops of inbred Wistar-AVN rats that were reared under germ-free conditions in the presence of various intestinal bacteria (enterobacteria and bifidobacteria isolated from CD patients and healthy children, respectively) and CD-triggering agents (gliadin and IFN-γ) by histology, scanning electron microscopy, immunofluorescence, and a rat cytokine antibody array. Adhesion of the bacterial strains to the IEC-6 rat cell line was evaluated in vitro. Gliadin fragments alone or together with the proinflammatory cytokine interferon (IFN)-γ significantly decreased the number of goblet cells in the small intestine; this effect was more pronounced in the presence of Escherichia coli CBL2 and Shigella CBD8. Shigella CBD8 and IFN-γ induced the highest mucin secretion and greatest impairment in tight junctions and, consequently, translocation of gliadin fragments into the lamina propria. Shigella CBD8 and E. coli CBL2 strongly adhered to IEC-6 epithelial cells. The number of goblet cells in small intestine increased by the simultaneous incubation of Bifidobacterium bifidum IATA-ES2 with gliadin, IFN-γ and enterobacteria. B. bifidum IATA-ES2 also enhanced the production of chemotactic factors and inhibitors of metalloproteinases, which can contribute to gut mucosal protection. CONCLUSIONS: Our results suggest that the composition of the intestinal microbiota affects the permeability of the intestinal mucosa and, consequently, could be involved in the early stages of CD pathogenesis.

• MeSH
• Bacteria pathogenicity   MeSH
• Bacterial Toxins pharmacology   MeSH
• Bifidobacterium pathogenicity   MeSH
• Celiac Disease etiology   MeSH
• Cytokines biosynthesis   MeSH
• Enterobacteriaceae pathogenicity   MeSH
• Gliadin pharmacokinetics   pharmacology   MeSH
• Germ-Free Life MeSH
• Host-Pathogen Interactions drug effects   MeSH
• Interferon-gamma pharmacology   MeSH
• Rats MeSH
• Permeability MeSH
• Goblet Cells pathology   MeSH
• Intestines microbiology   pathology   MeSH
• Intestinal Mucosa drug effects   metabolism   microbiology   MeSH
• Animals MeSH
• Check Tag
• Rats MeSH
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• Bacterial Toxins MeSH
• Cytokines MeSH
• Gliadin MeSH
• Interferon-gamma MeSH

To elucidate the role of innate immune responses in celiac disease, we investigated the effect of gliadin on blood monocytes from patients with celiac disease. Gliadin induced substantial TNF-alpha and IL-8 production by monocytes from patients with active celiac disease, lower levels by monocytes from patients with inactive celiac disease, and even lower levels by monocytes from healthy donors. In healthy donor monocytes gliadin induced IL-8 from monocytes expressing HLA-DQ2 and increased monocyte expression of the costimulatory molecules CD80 and CD86, the dendritic cell marker CD83, and the activation marker CD40. Gliadin also increased DNA binding activity of NF-kappaB p50 and p65 subunits in monocytes from celiac patients, and NF-kappaB inhibitors reduced both DNA binding activity and cytokine production. Thus, gliadin activation of HLA-DQ2(+) monocytes leading to chemokine and proinflammatory cytokine production may contribute to the host innate immune response in celiac disease.

• MeSH
• Macrophage Activation immunology   MeSH
• CD83 Antigen MeSH
• CD40 Antigens metabolism   MeSH
• B7-1 Antigen metabolism   MeSH
• B7-2 Antigen metabolism   MeSH
• Antigens, CD metabolism   MeSH
• Celiac Disease immunology   metabolism   MeSH
• Cytokines biosynthesis   MeSH
• Gliadin metabolism   MeSH
• HLA-DQ Antigens metabolism   MeSH
• Immunoglobulins metabolism   MeSH
• Interleukin-8 biosynthesis   MeSH
• Humans MeSH
• Membrane Glycoproteins metabolism   MeSH
• Monocytes metabolism   MeSH
• NF-kappa B metabolism   MeSH
• Peptide Fragments MeSH
• Flow Cytometry MeSH
• Tumor Necrosis Factor-alpha biosynthesis   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Names of Substances
• CD40 Antigens MeSH
• B7-1 Antigen MeSH
• B7-2 Antigen MeSH
• Antigens, CD MeSH
• Cytokines MeSH
• Gliadin MeSH
• HLA-DQ Antigens MeSH
• HLA-DQ2 antigen MeSH Browser
• Immunoglobulins MeSH
• Interleukin-8 MeSH
• Membrane Glycoproteins MeSH
• NF-kappa B MeSH
• Peptide Fragments MeSH
• Tumor Necrosis Factor-alpha MeSH