Tetramers of MHC-peptide complexes are used for detection and characterization of antigen-specific T cell responses, but they require knowledge about both antigenic peptide and the MHC restriction element. The successful application of these reagents in human diseases involving CD4 ؉ T cells is limited. Celiac disease, an intestinal inflammation driven by mucosal CD4 ؉ T cells recognizing wheat gluten peptides in the context of disease-associated HLA-DQ molecules, is an ideal model to test the potential clinical use of these reagents. We investigated whether gluten-specific T cells can be detected in the peripheral blood of celiac disease patients using DQ2 tetramers. Nine DQ2 ؉ patients and six control individuals on a gluten-free diet were recruited to the study. Participants consumed 160 g of gluten-containing bread daily for 3 days. After bread-challenge, gluten-specific T cells were detectable in the peripheral blood of celiac patients but not controls both directly by tetramer staining and indirectly by enzyme-linked immunospot. These T cells expressed the ␤7 integrin indicative of gut-homing properties. Most of the cells had a memory phenotype, but many other phenotypic markers showed a heterogeneous pattern. Tetramer staining of gluten-specific T cells has the potential to be used for diagnosis of celiac disease. tetramers T he development of multimeric MHC-peptide complexes has revolutionized the analysis of antigen-specific T cell responses. Tetramers are such reagents consisting of four soluble recombinant MHC molecules, each loaded with a single peptide and bound to a streptavidin molecule that is coupled with a fluorogenic marker (1). Multivalent engagement of the MHCpeptide complexes leads to a stable binding of the tetramer to T cell receptors on the T cell surface, allowing direct visualization of T cells with a defined specificity.MHC class I tetramer technology has greatly facilitated our understanding of CD8 ϩ T cell responses in viral infections and cancer (2). The benefit of tetramers for characterization and diagnosis of human autoimmune and infectious diseases has, however, been modest (3-6). This particularly relates to MHC class II tetramers used for the characterization of antigen-specific CD4 ϩ T cells. Only a few studies of relevance to autoimmunity exist (7-9). MHC class II tetramers are more difficult to produce than MCH class I tetramers (10, 11), and CD4 ϩ T cells of a given specificity appear to be present at much lower frequencies than their CD8 ϩ counterparts (12, 13). Several criteria have to be met to be able to detect antigen-reactive T cells with MHC II tetramers: both the peptide epitope and HLA-restriction element have to be identified, and sufficient frequency and relative high avidity of reactive T cells are needed (14).Celiac disease, a chronic inf lammatory disorder of the small intestine precipitated by ingestion of cereal gluten proteins, presents as an ideal model to test the potential clinical use of MHC class II tetramers. The disorder is driven by intestinal glu...

CD patients mounted a concomitant innate and adaptive immune response to gluten challenge. NCGS patients had increased density of intraepithelial CD3(+) T cells before challenge compared with disease controls and increased IFN-γ mRNA after challenge. Our results warrant further search for the pathogenic mechanisms for NCGS.

Tetramer staining for gluten-specific T cells is a sensitive method in detecting an immune response in CD patients after a short gluten challenge. The prevalence of CD in the group with self-prescribed GFD was about 10%.

BackgroundOf antigen-presenting cells (APCs) expressing HLA-DQ molecules in the celiac disease (CD) lesion, CD11c+ dendritic cells (DCs) co-expressing the monocyte marker CD14 are increased, whereas other DC subsets (CD1c+ or CD103+) and CD163+CD11c− macrophages are all decreased. It is unclear whether these changes result from chronic inflammation or whether they represent early events in the gluten response. We have addressed this in a model of in vivo gluten challenge.MethodsTreated HLA-DQ2+ CD patients (n = 12) and HLA-DQ2+ gluten-sensitive control subjects (n = 12) on a gluten-free diet (GFD) were orally challenged with gluten for three days. Duodenal biopsies obtained before and after gluten challenge were subjected to immunohistochemistry. Single cell digests of duodenal biopsies from healthy controls (n = 4), treated CD (n = 3) and untreated CD (n = 3) patients were analyzed by flow cytometry.ResultsIn treated CD patients, the gluten challenge increased the density of CD14+CD11c+ DCs, whereas the density of CD103+CD11c+ DCs and CD163+CD11c− macrophages decreased, and the density of CD1c+CD11c+ DCs remained unchanged. Most CD14+CD11c+ DCs co-expressed CCR2. The density of neutrophils also increased in the challenged mucosa, but in most patients no architectural changes or increase of CD3+ intraepithelial lymphocytes (IELs) were found. In control tissue no significant changes were observed.ConclusionsRapid accumulation of CD14+CD11c+ DCs is specific to CD and precedes changes in mucosal architecture, indicating that this DC subset may be directly involved in the immunopathology of the disease. The expression of CCR2 and CD14 on the accumulating CD11c+ DCs indicates that these cells are newly recruited monocytes.