A man in his seventies with fatigue and renal failure. / En mann i 70-årene med utmattelse og nyresvikt.

A man in his seventies underwent routine heart examinations as part of workup for kidney transplantation. Unexpected findings led to more extensive investigations and revealed two rare systemic diseases as causes of his heart failure.

Case Report: Let Us Not Forget the Treatment That Some Patients Have Received-The Brief 50-Year History of a Kidney Transplant Survivor.

Background: There has been a considerable improvement in post-transplant care since the early 1960s. Some patients we meet in the clinic have personally experienced this progress and have histories to tell that one must not forget. This is the brief history of a long-time "transplant survivor." Case Presentation: In 1970, a young woman developed acute oedema, proteinuria, hypertension and oliguria during pregnancy. Labor was induced, but neither the child nor the kidney function could be saved. Our patient started dialysis, and 4 years later received a kidney transplant donated by her father (then 55 years of age). Maintenance immunosuppression consisted of prednisolone and azathioprine until 2011, when azathioprine was switched to everolimus due to skin cancer. Before this, our patient was highly satisfied with prednisolone/azathioprine, despite discussions regarding newer immunosuppressive drugs, and always reminded the treating physician that one should "never change a winning team." Retrospectively, the avoidance of calcineurin inhibitors might have been beneficial for this patient who still has preserved an excellent renal function with s-creatinine levels around 100 µmol/L and just had sparse fibrosis detected in a recently performed transplant biopsy. The transplanted kidney is now 101 years old and is still working 24/7. Conclusions: Our patient received a kidney transplant for 46 years ago and still has a remarkably stable transplant function with s-creatinine levels around 100 µmol/L. This case report illustrates the potential endurance of the kidneys and is a reminder to keep taking individualized treatment decisions even though new treatment alternatives promise superiority.

Glomerulonephritis after contact with a horse. / Glomerulonefritt etter kontakt med hest.

BACKGROUND: Zoonoses are important to consider when humans become ill after being in contact with animals. In such cases thorough patient history is crucial, especially when infections have an unclear cause. We present a patient with infection-associated glomerulonephritis, where a horse was the probable source of infection. CASE PRESENTATION: A young woman was admitted to the district general hospital in Vestfold, Norway, with infection and acute kidney failure. Renal biopsy suggested glomerulonephritis, and nasopharyngeal culture taken at admission detected Streptococcus equi. It emerged that the patient had daily contact with horses. INTERPRETATION: As Streptococcus equi is not part of normal human flora and the clinical signs were compatible with infection-associated glomerulonephritis, it was considered a probable causal link between the microbial finding and diagnosis. The source of infection was one of the horses.

Similar Responses of Intestinal T Cells From Untreated Children and Adults With Celiac Disease to Deamidated Gluten Epitopes.

BACKGROUND & AIMS: Celiac disease is a chronic small intestinal inflammatory disorder mediated by an immune response to gluten peptides in genetically susceptible individuals. Celiac disease is often diagnosed in early childhood, but some patients receive a diagnosis late in life. It is uncertain whether pediatric celiac disease is distinct from adult celiac disease. It has been proposed that gluten-reactive T cells in children recognize deamidated and native gluten epitopes, whereas T cells from adults only recognize deamidated gluten peptides. We studied the repertoire of gluten epitopes recognized by T cells from children and adults. METHODS: We examined T-cell responses against gluten by generating T-cell lines and T-cell clones from intestinal biopsies of adults and children and tested proliferative response to various gluten peptides. We analyzed T cells from 14 children (2-5 years old) at high risk for celiac disease who were followed for celiac disease development. We also analyzed T cells from 6 adults (26-55 years old) with untreated celiac disease. All children and adults were positive for HLA-DQ2.5. Biopsies were incubated with gluten digested with chymotrypsin (modified or unmodified by the enzyme transglutaminase 2) or the peptic-tryptic digest of gliadin (in native and deamidated forms) before T-cell collection. RESULTS: Levels of T-cell responses were higher to deamidated gluten than to native gluten in children and adults. T cells from children and adults each reacted to multiple gluten epitopes. Several T-cell clones were cross-reactive, especially clones that recognized epitopes from γ-and ω-gliadin. About half of the generated T-cell clones from children and adults reacted to unknown epitopes. CONCLUSIONS: T-cell responses to different gluten peptides appear to be similar between adults and children at the time of diagnosis of celiac disease.

Long-term mucosal recovery and healing in celiac disease is the rule - not the exception.

OBJECTIVE: The prevalence of persistent villous atrophy (VA) in patients with celiac disease (CD) on a gluten-free diet (GFD) varies greatly between studies. Most studies show a relatively high prevalence of mucosal atrophy and inflammation in treated patients, a finding which have led to a concept of non-responsive CD. Few studies have examined the prevalence of long-term mucosal healing. Our study aimed to determine the extent of mucosal healing in a cohort of Norwegian patients with CD treated with GFD for several years. MATERIALS AND METHODS: Adult patients diagnosed with VA between 1989 and 2009 were included. We performed a follow-up gastroscopy with duodenal biopsies. Two pathologists evaluated the biopsies according to the Marsh-Oberhuber classification. Mucosal healing was defined as Marsh 0 while mucosal recovery was defined as Marsh 0-2. RESULTS: Duodenal biopsies were obtained from 127 adult patients with established CD. After a follow-up time of 8.1 years (median, range 2.3-22.3), 103 (81%) of the patients showed mucosal healing, 120 patients (94%) showed mucosal recovery, and 7 patients (6%) showed persistent VA. In addition, 103 of the 127 patients (81%) had undergone a routine follow-up biopsy 12.6 months (median, range 5.2-28.8) after diagnosis. At the time of the routine follow-up, only 52 of these patients (50.5%) had achieved mucosal recovery. CONCLUSIONS: Although half of the patients had persistent VA at the time of routine follow-up, both long-term mucosal recovery and healing is possible for the vast majority of adult patients with CD.

Tetramer-visualized gluten-specific CD4+ T cells in blood as a potential diagnostic marker for coeliac disease without oral gluten challenge.

BACKGROUND: Diagnosing coeliac disease (CD) can be challenging, despite highly specific autoantibodies and typical mucosal changes in the small intestine. The T-cell response to gluten is a hallmark of the disease that has been hitherto unexploited in clinical work-up. OBJECTIVES: We aimed to develop a new method that directly visualizes and characterizes gluten-reactive CD4+ T cells in blood, independently of gluten challenge, and to explore its diagnostic potential. METHODS: We performed bead-enrichment of DQ2.5-glia-α1a and DQ2.5-glia-α2 tetramer+ cells in the blood of control individuals, treated (TCD) and untreated patients (UCD). We visualized these cells by flow cytometry, sorted them and cloned them. We assessed their specificity by antigen stimulation and re-staining with tetramers. RESULTS: We detected significantly more gliadin-tetramer+ CD4+ effector memory T cells (TEM) in UCD and TCD patients, compared to controls. Significantly more gliadin-tetramer+ TEM in the CD patients than in controls expressed the gut-homing marker integrin-ß7. CONCLUSION: Quantification of gut-homing, gluten-specific TEM in peripheral blood, visualized with human leukocyte antigen (HLA) -tetramers, may be used to distinguish CD patients from healthy individuals. Easy access to gluten-reactive blood T cells from diseased and healthy individuals may lead to new insights on the disease-driving CD4+ T cells in CD.

Direct cloning and tetramer staining to measure the frequency of intestinal gluten-reactive T cells in celiac disease.

Knowledge of the frequency of disease-driving CD4⁺ T cells in lesions of chronic human inflammatory diseases is limited. In celiac disease (CD), intestinal gluten-reactive CD4⁺ T cells, which recognize gluten peptides only in the context of the disease-associated HLA-DQ molecules, are key pathogenic players. By cloning CD4⁺ T cells directly from intestinal biopsies of CD patients, we found that 0.5-1.8% of CD4⁺ T cells were gluten reactive. About half of the gluten-reactive T cells were specific for either the immuno-dominant DQ2.5-glia-α1a or DQ2.5-glia-α2 epitopes, suggesting that direct visualization of gluten-specific T cells could be possible. Assessed by flow cytometry, tetramer-positive T cells were present in 10/10 untreated CD patients with a frequency of 0.1-1.2% of CD4⁺ T cells. Gluten-specific T cells were also detectable in most treated CD patients (7/10). Moreover, the frequency of gluten-specific T cells correlated with the degree of histological damage in the gut mucosa as scored by Marsh-grading, and also with serum IgA anti-transglutaminase 2 antibody levels. Tetramer staining of gluten-reactive T cells in biopsy material is a useful tool for future studies of such cells in CD and could also potentially serve as a diagnostic supplement in selected cases.

The adaptive immune response in celiac disease.

Compared to other human leukocyte antigen (HLA)-associated diseases such as type 1 diabetes, multiple sclerosis, and rheumatoid arthritis, fundamental aspects of the pathogenesis in celiac disease are relatively well understood. This is mostly because the causative antigen in celiac disease-cereal gluten proteins-is known and the culprit HLA molecules are well defined. This has facilitated the dissection of the disease-relevant CD4+ T cells interacting with the disease-associated HLA molecules. In addition, celiac disease has distinct antibody responses to gluten and the autoantigen transglutaminase 2, which give strong handles to understand all sides of the adaptive immune response leading to disease. Here we review recent developments in the understanding of the role of T cells, B cells, and antigen-presenting cells in the pathogenic immune response of this instructive disorder.

Rapid accumulation of CD14+CD11c+ dendritic cells in gut mucosa of celiac disease after in vivo gluten challenge.

BACKGROUND: Of 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. METHODS: Treated 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. RESULTS: In 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. CONCLUSIONS: Rapid 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.

Interaction between granulosa-lutein cells and monocytes regulates secretion of angiogenic factors in vitro.

BACKGROUND: Leukocyte infiltration and angiogenesis in the forming corpus luteum are prerequisites for normal ovarian function and may also underlie disorders like ovarian hyperstimulation syndrome. We examined whether ovarian angiogenesis could be affected by an interaction between granulosa-lutein (GL) cells and leukocytes. METHODS AND RESULTS: We found that GL cells isolated from the follicular fluid synthesize and secrete the chemokine interleukin-8 (IL-8), which activates IL-8-receptor-specific Ca(2+) and p38 mitogen-activated protein kinase signalling in monocytes and induces a directed migration of these cells towards the chemical gradient. Monocytes were found to further enhance IL-8 release, which suggests that these cells promote a massive leukocyte infiltration of the forming corpus luteum. A possible utility of leukocyte infiltration is the modulation of angiogenesis. We found that GL cells induce migration and capillary tube formation by endothelial cells in vitro. Furthermore, monocytes altered the profile of angiogenic factors released by GL cells, which supports the theory that an interaction between GL cells and leukocytes regulates ovarian angiogenesis. In addition, we found a correlation between increased secretion of pro-angiogenic cytokines and number of oocytes collected during IVF, which suggests that ovarian angiogenesis is related to the clinical response during ovarian stimulation. CONCLUSIONS: An intricate communication may exist between infiltrating leukocytes and ovarian GL cells during the formation of corpus luteum, affecting neo-vascularization of the luteal tissue.

Posttranslational modification of gluten shapes TCR usage in celiac disease.

Posttranslational modification of Ag is implicated in several autoimmune diseases. In celiac disease, a cereal gluten-induced enteropathy with several autoimmune features, T cell recognition of the gluten Ag is heavily dependent on the posttranslational conversion of Gln to Glu residues. Evidence suggests that the enhanced recognition of deamidated gluten peptides results from improved peptide binding to the MHC and TCR interaction with the peptide-MHC complex. In this study, we report that there is a biased usage of TCR Vß6.7 chain among TCRs reactive to the immunodominant DQ2-α-II gliadin epitope. We isolated Vß6.7 and DQ2-αII tetramer-positive CD4(+) T cells from peripheral blood of gluten-challenged celiac patients and sequenced the TCRs of a large number of single T cells. TCR sequence analysis revealed in vivo clonal expansion, convergent recombination, semipublic response, and the notable conservation of a non-germline-encoded Arg residue in the CDR3ß loop. Functional testing of a prototype DQ2-α-II-reactive TCR by analysis of TCR transfectants and soluble single-chain TCRs indicate that the deamidated residue in the DQ2-α-II peptide poses constraints on the TCR structure in which the conserved Arg residue is a critical element. The findings have implications for understanding T cell responses to posttranslationally modified Ags.

CD62L(neg)CD38⁺ expression on circulating CD4⁺ T cells identifies mucosally differentiated cells in protein fed mice and in human celiac disease patients and controls.

OBJECTIVES: The aim of this study was to identify new markers of mucosal T cells to monitor ongoing intestinal immune responses in peripheral blood. METHODS: Expression of cell-surface markers was studied in mice on ovalbumin (OVA)-specific T cells in the gut-draining mesenteric lymph nodes (MLN) after OVA feed. The effect of the local mucosal mediators retinoic acid (RA) and transforming growth factor-ß (TGF-ß) on the induction of a mucosal phenotype was determined in in vitro T-cell differentiation assays with murine and human T cells. Tetramer stainings were performed to study gluten-specific T cells in the circulation of patients with celiac disease, a chronic small-intestinal inflammation. RESULTS: In mice, proliferating T cells in MLN were CD62L(neg)CD38(+) during both tolerance induction and abrogation of intestinal homeostasis. This mucosal CD62L(neg)CD38(+) T-cell phenotype was efficiently induced by RA and TGF-ß in mice, whereas for human CD4(+) T cells RA alone was sufficient. The CD4(+)CD62L(neg)CD38(+) T-cell phenotype could be used to identify T cells with mucosal origin in human peripheral blood, as expression of the gut-homing chemokine receptor CCR9 and ß(7) integrin were highly enriched in this subset whereas expression of cutaneous leukocyte-associated antigen was almost absent. Tetramer staining revealed that gluten-specific T cells appearing in blood of treated celiac disease patients after oral gluten challenge were predominantly CD4(+)CD62L(neg)CD38(+). The total percentage of circulating CD62L(neg)CD38(+) of CD4 T cells was not an indicator of intestinal inflammation as percentages did not differ between pediatric celiac disease patients, inflammatory bowel disease patients and respective controls. However, the phenotypic selection of mucosal T cells allowed cytokine profiling as upon restimulation of CD62L(neg)CD38(+) cells interleukin-10 (IL-10) and interferon-γ (IFN-γ) transcripts were readily detected in circulating mucosal T cells. CONCLUSIONS: By selecting for CD62L(neg)CD38(+) expression that comprises 5-10% of the cells within the total CD4(+) T-cell pool we are able to highly enrich for effector T cells with specificity for mucosal antigens. This is of pivotal importance for functional studies as this purification enhances the sensitivity of cytokine detection and cellular activation.

Assessing possible celiac disease by an HLA-DQ2-gliadin Tetramer Test.

OBJECTIVES: Investigation of uncertain celiac disease (CD) in patients already on a gluten-free diet (GFD) is difficult. We evaluated HLA-DQ2-gliadin tetramers for detection of gluten-specific T cells in peripheral blood and histological changes in the duodenum after a short gluten challenge as a diagnostic tool. METHODS: HLA-DQ2+ individuals on a GFD for at least 4 weeks were investigated; 35 with uncertain diagnosis, 13 CD patients, and 2 disease controls. All participants had a challenge with four slices of gluten-containing white bread, daily for 3 days (d1-d3). An esophagogastroduodenoscopy with biopsy sampling was done on d0 and d4. Biopsies were scored according to revised Marsh criteria. Peripheral blood CD4+ T cells were isolated, stained with HLA-DQ2-gliadin peptide tetramers, and analyzed by flow cytometry on d0 and d6. RESULTS: After challenge, a positive tetramer test was seen in 11/13 CD patients. Four of these subjects also showed typical histological changes on challenge. Of the 35 patients with uncertain diagnosis, 3 were diagnosed with CD. Two of these three patients had both positive tetramer staining and histological changes in biopsies after challenge. CONCLUSIONS: 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%.

Rapid generation of rotavirus-specific human monoclonal antibodies from small-intestinal mucosa.

The gut mucosal surface is efficiently protected by Abs, and this site represents one of the richest compartments of Ab-secreting cells in the body. A simple and effective method to generate Ag-specific human monoclonal Abs (hmAbs) from such cells is lacking. In this paper, we describe a method to generate hmAbs from single Ag-specific IgA- or IgM-secreting cells of the intestinal mucosa. We found that CD138-positive plasma cells from the duodenum expressed surface IgA or IgM. Using eGFP-labeled virus-like particles, we harnessed the surface Ig expression to detect rotavirus-specific plasma cells at low frequency (0.03-0.35%) in 9 of 10 adult subjects. Single cells were isolated by FACS, and as they were viable, further testing of secreted Abs by ELISPOT and ELISA indicated a highly specific selection procedure. Ab genes from single cells of three donors were cloned, sequenced, and expressed as recombinant hmAbs. Of 26 cloned H chain Ab genes, 22 were IgA and 4 were IgM. The genes were highly mutated, and there was an overrepresentation of the VH4 family. Of 10 expressed hmAbs, 8 were rotavirus-reactive (6 with K(d) < 1 × 10(-10)). Importantly, our method allows generation of hmAbs from cells implicated in the protection of mucosal surfaces, and it can potentially be used in passive vaccination efforts and for discovery of epitopes directly relevant to human immunity.

Differential release of matrix metalloproteinases and tissue inhibitors of metalloproteinases by human granulosa-lutein cells and ovarian leukocytes.

Tissue reorganization during ovulation and corpus luteum formation involves a coordinated action of matrix metalloproteinases (MMPs) and tissue MMP inhibitors (TIMPs). In this study we investigated the cellular source of ovarian MMPs and TIMPs. Cells isolated from the preovulatory human follicle were cultured after immunobead depletion of CD45-expressing cells, which allowed differential assessment of leukocyte and granulosa-lutein cell fractions. Secretion of MMP-9 by follicular fluid-derived cells was associated with the presence of leukocytes. Granulosa-lutein cells synthesized low levels of MMP-9 but failed to secrete this enzyme that presumably accumulated in the cytoplasm, indicated by an increased MMP-9 expression of luteinized cells in sectioned midluteal phase corpora lutea. Synthesis and secretion of TIMP by follicular fluid-derived cells was associated with granulosa-lutein cells. TIMPs derived by granulosa-lutein cells failed to inhibit MMP-related pericellular proteolysis. The findings support a two-cell model of periovulatory MMP/TIMP release, in which leukocytes secrete MMPs and granulosa-lutein cells release TIMP, suggesting that there exists an intriguing interaction among cells that intertwingle during ovulation and corpus luteum formation.

Tetramer visualization of gut-homing gluten-specific T cells in the peripheral blood of celiac disease patients.

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 beta(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.

Characterization and depletion of leukocytes from cells isolated from the pre-ovulatory ovarian follicle.

BACKGROUND: Cells isolated from the periovulatory ovarian follicle are often used as a model of ovarian steroidogenesis and corpus luteum formation. The follicular fluid-derived cell (FFDC) population is, however, heterogeneous and in addition to granulosa-lutein cells, non-steroidogenic cells are also present. These non-steroidogenic cells, especially the immune cells, may have important biological functions in this model. Here, we describe a method to isolate FFDC, characterize the phenotype of the immune cells and deplete immune cells from FFDC. METHODS AND RESULTS: Follicular fluid aspirated transvaginally during IVF was clarified by centrifugation and enzymatic dispersion, labelled for leukocyte-specific markers and analysed by flow cytometry. Leukocytes constituted 22% of FFDC and expressed macrophage/dendritic cell, monocyte and lymphocyte markers. Leukocytes were depleted with anti-CD45-conjugated immunobeads, resulting in an FFDC population with <1.9% leukocytes. Leukocyte-containing FFDC secreted more interleukin-8 in culture than leukocyte-depleted FFDC. CONCLUSION: Leukocyte-depleted FFDC may serve as a useful model to study the interaction of immune cells and luteinizing cells during corpus luteum formation.

A unique dendritic cell subset accumulates in the celiac lesion and efficiently activates gluten-reactive T cells.

BACKGROUND & AIMS: Celiac disease is a chronic inflammation of the duodenal mucosa driven by gluten-reactive T cells restricted by the disease-associated HLA-DQ2 molecule. The mechanisms that regulate the activation of mucosal T cells are, however, understood poorly. The aim of this study was to identify the antigen-presenting cells that are responsible for the activation of gluten-reactive T cells in the celiac lesion. METHODS: Intestinal biopsy specimens obtained from untreated and treated celiac patients and normal controls were either snap-frozen directly or incubated for 24 hours with or without gluten peptides. Cryosections were subjected to multicolor immunofluorescence applying monoclonal antibodies to a range of antigen-presenting cell markers. Macrophages and dendritic cells were isolated from enzymatically digested small intestinal biopsies of untreated patients and incubated with gluten-reactive T-cell clones to measure their antigen-presenting capacity. RESULTS: HLA-DQ2+ cells in the normal duodenal mucosa consisted of 2 distinct cell populations: about 80% were CD68+ DC-lysosome intercellular adhesion molecule-3-grabbing nonintegrin+ macrophages and 20% were CD11c+ dendritic cells. Importantly, the CD11c+ dendritic cells accumulated in the celiac lesion and revealed an activated phenotype expressing CD86 and DC-specific-associated membrane protein. Moreover, when isolated from challenged biopsy specimens, the CD11c+ dendritic cells efficiently activated gluten-reactive T cells. CONCLUSIONS: Our results suggest that a unique subset of dendritic cells are responsible for local activation of gluten-reactive T cells in the celiac lesion.

HLA-DQ2 and -DQ8 signatures of gluten T cell epitopes in celiac disease.

Celiac disease is associated with HLA-DQ2 and, to a lesser extent, HLA-DQ8. Type 1 diabetes is associated with the same DQ molecules in the opposite order and with possible involvement of trans-encoded DQ heterodimers. T cells that are reactive with gluten peptides deamidated by transglutaminase 2 and invariably restricted by DQ2 or DQ8 can be isolated from celiac lesions. We used intestinal T cells from celiac patients to map DQ2 and DQ8 epitopes within 2 representative gluten proteins, alpha-gliadin AJ133612 and gamma-gliadin M36999. For alpha-gliadin, DQ2- and DQ8-restricted T cells recognized deamidated peptides of 2 separate regions. For gamma-gliadin, DQ2- and DQ8-restricted T cells recognized deamidated peptides of the same region. Some gamma-gliadin peptides were recognized by T cells in the context of DQ2 or DQ8 when bound in exactly the same registers, but with different requirements for deamidation; deamidation at peptide position 4 (P4) was important for DQ2-restricted T cells, whereas deamidation at P1 and/or P9 was important for DQ8-restricted T cells. Peptides combining the DQ2 and DQ8 signatures could be presented by DQ2, DQ8, and trans-encoded DQ heterodimers. Our findings shed light on the basis for the HLA associations in celiac disease and type 1 diabetes.

Mapping of gluten T-cell epitopes in the bread wheat ancestors: implications for celiac disease.

BACKGROUND AND AIMS: Celiac disease is a prevalent disorder characterized by a chronic intestinal inflammation driven by HLA-DQ2 or -DQ8-restricted T cells specific for ingested wheat gluten peptides. The dominant T-cell responses are to epitopes that cluster within a stable 33mer fragment formed by physiologic digestion of distinct alpha-gliadins. Celiac disease is treated by excluding all gluten proteins from the diet. Conceivably, a diet based on baking-quality gluten from a wheat species that expresses no or few T-cell stimulatory gluten peptides should be equally well tolerated by the celiac patients and, importantly, also be beneficial for disease prevention. METHODS: To identify baking quality, harmless wheat, we followed the evolution of the wheat back to the species that most likely have contributed the AA, BB, and DD genomes to the bread wheat. Gluten were extracted from a large collection of these ancient wheat species and screened for T-cell stimulatory gluten peptides. RESULTS: Distinct differences in the intestinal T-cell responses to the diploid species were identified. Interestingly, we found that the fragments identical or equivalent to the immunodominant 33mer fragment are encoded by alpha-gliadin genes on the wheat chromosome 6D and thus absent from gluten of diploid einkorn (AA) and even certain cultivars of the tetraploid (AABB) pasta wheat. CONCLUSIONS: These findings have implications for celiac disease because they raise the prospect of identifying or producing by breeding wheat species with low or absent levels of harmful gluten proteins.