Celiac disease: how complicated can it get?

In the small intestine of celiac disease patients, dietary wheat gluten and similar proteins in barley and rye trigger an inflammatory response. While strict adherence to a gluten-free diet induces full recovery in most patients, a small percentage of patients fail to recover. In a subset of these refractory celiac disease patients, an (aberrant) oligoclonal intraepithelial lymphocyte population develops into overt lymphoma. Celiac disease is strongly associated with HLA-DQ2 and/or HLA-DQ8, as both genotypes predispose for disease development. This association can be explained by the fact that gluten peptides can be presented in HLA-DQ2 and HLA-DQ8 molecules on antigen presenting cells. Gluten-specific CD4⁺ T cells in the lamina propria respond to these peptides, and this likely enhances cytotoxicity of intraepithelial lymphocytes against the intestinal epithelium. We propose a threshold model for the development of celiac disease, in which the efficiency of gluten presentation to CD4⁺ T cells determines the likelihood of developing celiac disease and its complications. Key factors that influence the efficiency of gluten presentation include: (1) the level of gluten intake, (2) the enzyme tissue transglutaminase 2 which modifies gluten into high affinity binding peptides for HLA-DQ2 and HLA-DQ8, (3) the HLA-DQ type, as HLA-DQ2 binds a wider range of gluten peptides than HLA-DQ8, (4) the gene dose of HLA-DQ2 and HLA-DQ8, and finally,(5) additional genetic polymorphisms that may influence T cell reactivity. This threshold model might also help to understand the development of refractory celiac disease and lymphoma.     

Detection of Specific IgA Antibodies against a Novel Deamidated 8-Mer Gliadin Peptide in Blood Plasma Samples from Celiac Patients

We studied whether celiac disease (CD) patients produce antibodies against a novel gliadin peptide specifically generated in the duodenum of CD patients by a previously described pattern of CD-specific duodenal proteases. Fingerprinting and ion-trap mass spectrometry of CD-specific duodenal gliadin-degrading protease pattern revealed a new 8-mer gliadin-derived peptide. An ELISA against synthetic deamidated 8-mer peptides (DGP 8-mer) was used to study the presence of IgA anti-DGP 8-mer antibodies in plasma samples from 81 children (31 active CD patients (aCD), 17 CD patients on a gluten-free diet (GFD), 10 healthy controls (C) and 23 patients with other gastrointestinal pathology (GP)) and 101 adults (16 aCD, 12 GFD, 27 C and 46 GP-patients). Deamidation of the 8-mer peptide significantly increased the reactivity of the IgA antibodies from CD patients against the peptide. Significant IgA anti-DGP 8-mer antibodies levels were detected in 93.5% of aCD-, 11.8% of GFD- and 4.3% of GP-patients in children. In adults, antibodies were detected in 81.3% of aCD-patients and 8.3% of GFD-patients while were absent in 100% of C- and GP-patients. Duodenal CD-specific gliadin degrading proteases release an 8-mer gliadin peptide that once deamidated is an antigen for specific IgA antibodies in CD patients which may provide a new accurate diagnostic tool in CD.     

Impact of dietary gluten on regulatory T cells and Th17 cells in BALB/c mice

Dietary gluten influences the development of type 1 diabetes (T1D) and a gluten-free (GF) diet has a protective effect on the development of T1D. Gluten may influence T1D due to its direct effect on intestinal immunity; however, these mechanisms have not been adequately studied. We studied the effect of a GF diet compared to a gluten-containing standard (STD) diet on selected T cell subsets, associated with regulatory functions as well as proinflammatory Th17 cells, in BALB/c mice. Furthermore, we assessed diet-induced changes in the expression of various T cell markers, and determined if changes were confined to intestinal or non-intestinal lymphoid compartments. The gluten-containing STD diet led to a significantly decreased proportion of γδ T cells in all lymphoid compartments studied, although an increase was detected in some γδ T cell subsets (CD8(+), CD103(+)). Further, it decreased the proportion of CD4(+)CD62L(+) T cells in Peyer's patches. Interestingly, no diet-induced changes were found among CD4(+)Foxp3(+) T cells or CD3(+)CD49b(+)cells (NKT cells) and CD3(-)CD49b(+) (NK) cells. Mice fed the STD diet showed increased proportions of CD4(+)CD45RB(high+) and CD103(+) T cells and a lower proportion of CD4(+)CD45RB(low+) T cells in both mucosal and non-mucosal compartments. The Th17 cell population, associated with the development of autoimmunity, was substantially increased in pancreatic lymph nodes of mice fed the STD diet. Collectively, our data indicate that dietary gluten influences multiple regulatory T cell subsets as well as Th17 cells in mucosal lymphoid tissue while fewer differences were observed in non-mucosal lymphoid compartments.     

Tissue transglutaminase in celiac disease: role of autoantibodies

In celiac disease (CD), gluten, the disease-inducing toxic component in wheat, induces the secretion of IgA-class autoantibodies which target tissue transglutaminase (tTG). These autoantibodies are produced in the small-intestinal mucosa, and, during gluten consumption, they can also be detected in patients' serum but disappear slowly from the circulation on a gluten-free diet. Interestingly, after adoption of a gluten-free diet the serum autoantibodies disappear from the circulation more rapidly than the small-intestinal mucosal autoantibody deposits. The finding of IgA deposits on extracellular tTG in the liver, kidney, lymph nodes and muscles of patients with CD indicates that tTG is accessible to the gut-derived autoantibodies. Although the specific autoantibody response directed against tTG is very characteristic in celiac patients, their role in the immunopathology of the celiac mucosal lesion is a matter of debate. Here we report a brief summary of anti-tTG antibody effects demonstrating that these antibodies are functional and not mere bystanders in the disease pathogenesis. In fact, they inhibit intestinal epithelial cell differentiation, induce intestinal epithelial cell proliferation, increase epithelial permeability and activate monocytes and disturb angiogenesis.     

DNAM-1 mediates epithelial cell-specific cytotoxicity of aberrant intraepithelial lymphocyte lines from refractory celiac disease type II patients

In refractory celiac disease (RCD), intestinal epithelial damage persists despite a gluten-free diet. Characteristic for RCD type II (RCD II) is the presence of aberrant surface TCR-CD3(-) intraepithelial lymphocytes (IELs) that can progressively replace normal IELs and eventually give rise to overt lymphoma. Therefore, RCD II is considered a malignant condition that forms an intermediate stage between celiac disease (CD) and overt lymphoma. We demonstrate in this study that surface TCR-CD3(-) IEL lines isolated from three RCD II patients preferentially lyse epithelial cell lines. FACS analysis revealed that DNAM-1 was strongly expressed on the three RCD cell lines, whereas other activating NK cell receptors were not expressed on all three RCD cell lines. Consistent with this finding, cytotoxicity of the RCD cell lines was mediated mainly by DNAM-1 with only a minor role for other activating NK cell receptors. Furthermore, enterocytes isolated from duodenal biopsies expressed DNAM-1 ligands and were lysed by the RCD cell lines ex vivo. Although DNAM-1 on CD8(+) T cells and NK cells is known to mediate lysis of tumor cells, this study provides, to our knowledge, the first evidence that (pre)malignant cells themselves can acquire the ability to lyse epithelial cells via DNAM-1. This study confirms previous work on epithelial lysis by RCD cell lines and identifies a novel mechanism that potentially contributes to the gluten-independent tissue damage in RCD II and RCD-associated lymphoma.     

Iron,vitamins and minerals status in pediatric patients with celiac diseaseand non celiac gluten sensitivity prior to diagnosis and three months later

AimPathologies associated with gluten intake are increasingly prevalent. Diagnosis of celiac disease (CD) or non-celiac gluten sensitivity (NCGS) is based on compatible clinical alterations and in case of CD on compatible serology or intestinal biopsy. The aim was to determine the values of iron, vitamins and minerals prior to diagnosis and to verify whether a gluten-free diet treatment can cause the normalization of these parameters both in patients diagnosed with celiac disease and gluten sensitivity.Methods^{Retrospective} observational study from November 2016 to November 2021. 101 celiac patients and 26 with NCGS were included, all under 18 years of age. Levels of Fe, Na+ , K+ , Cl-, Ferritin, Ca2 + , P, Vitamin B12, Vitamin D and Transferrin were determined, following the quality standards of the Hospital Clínico San Carlos laboratory. Statistical software IBM SPSS Statistics v.26 was used.ResultsCalcium levels in celiac patients follow a positive trend after 3 months of gluten-free diet. Ferritin levels in patients with NCGS increased in a statistically significant way (p < 0.017).ConclusionCalcium in the CD group increases its values after the establishment of a gluten-free diet as treatment, as well as ferritin in patients with NCGS. No significant changes were found in the rest of the analyzed parameters. This could be due to the precocity of the diagnosis thanks to a rapid clinical suspicion that determines few analytical alterations.     

Celiac disease: risk assessment, diagnosis, and monitoring

Celiac disease is an autoimmune disorder occurring in genetically susceptible individuals, triggered by gluten and related prolamins. Well identified haplotypes in the human leukocyte antigen (HLA) class II region (either DQ2 [DQA*0501-DQB*0201] or DQ8 [DQA*0301-DQB1*0302]) confer a large part of the genetic susceptibility to celiac disease.Celiac disease originates as a result of a combined action involving both adaptive and innate immunity. The adaptive immune response to gluten has been well described, with the identification of specific peptide sequences demonstrating HLA-DQ2 or -DQ8 restrictive binding motifs across various gluten proteins. As for innate immunity, through specific natural killer receptors expressed on their surface, intra-epithelial lymphocytes recognize nonclassical major histocompatibility complex (MHC)-I molecules such as MICA, which are induced on the surface of enterocytes by stress and inflammation, and this interaction leads to their activation to become lymphokine-activated killing cells. Four possible presentations of celiac disease are recognized: (i) typical, characterized mostly by gastrointestinal signs and symptoms; (ii) atypical or extraintestinal, where gastrointestinal signs/symptoms are minimal or absent and a number of other manifestations are present; (iii) silent, where the small intestinal mucosa is damaged and celiac disease autoimmunity can be detected by serology, but there are no symptoms; and, finally, (iv) latent, where individuals possess genetic compatibility with celiac disease and may also show positive autoimmune serology, that have a normal mucosa morphology and may or may not be symptomatic.The diagnosis of celiac disease still rests on the demonstration of changes in the histology of the small intestinal mucosa. The classic celiac lesion occurs in the proximal small intestine with histologic changes of villous atrophy, crypt hyperplasia, and increased intraepithelial lymphocytosis. Currently, serological screening tests are utilized primarily to identify those individuals in need of a diagnostic endoscopic biopsy. The serum levels of immunoglobulin (Ig)A anti-tissue transglutaminase (or TG2) are the first choice in screening for celiac disease, displaying the highest levels of sensitivity (up to 98%) and specificity (around 96%). Anti-endomysium antibodies-IgA (EMA), on the other hand, have close to 100% specificity and a sensitivity of greater than 90%. The interplay between gliadin peptides and TG2 is responsible for the generation of novel antigenic epitopes, the TG2-generated deamidated gliadin peptides. Such peptides represent much more celiac disease-specific epitopes than native peptides, and deamidated gliadin antibodies (DGP) have shown promising results as serological markers for celiac disease. Serology has also been employed in monitoring the response to a gluten-free diet.Despite the gluten-free diet being so effective, there is a growing demand for alternative treatment options. In the future, new forms of treatment may include the use of gluten-degrading enzymes to be ingested with meals, the development of alternative, gluten-free grains by genetic modification, the use of substrates regulating intestinal permeability to prevent gluten entry across the epithelium, and, finally, the availability of different forms of immunotherapy.     

In vitro cytotoxic effect of wheat gliadin-derived peptides on the Caco-2 intestinal cell line is associated with intracellular oxidative imbalance: implications for coeliac disease

Coeliac disease (CD) is an inflammatory disorder of the upper small intestine in which gluten acts as an essential factor in its pathogenesis. Although it is generally accepted that cereal protein activation of the immune system is involved in CD progression, a non-immunomediated cytotoxic activity of gliadin-derived peptides on the jejunal/duodenal tract cannot be excluded. In this work, considering that (a) little has been reported about the intracellular metabolic events associated with gliadin toxicity, and (b) an important role for free radicals in a number of gastrointestinal disease has been demonstrated, we investigated the in vitro effects of gliadin-derived peptides on redox metabolism of Caco-2 intestinal cells during a kinetic study in which cells were exposed to peptic–tryptic digest of bread wheat up to 48 h. We found that the antiproliferative effects displayed by gliadin exposure was associated with intracellular oxidative imbalance, characterised by an increased presence of lipid peroxides, an augmented oxidised (GSSG)/reduced (GSH) glutathione ratio and a loss in protein-bound sulfhydryl groups. Significant structural perturbations of the cell plasma membrane were also detected. Additional experiments performed by using the specific GSH-depleting agent buthionine sulfoximine provide evidence that the extent of gliadin-induced cell growth arrest critically depends upon the ‘basal’ redox profile of the enterocytes. On the whole, these findings seem to suggest that, besides the adoption of a strictly gluten-free diet, the possibility for an adjuvant therapy with antioxidants may be considered for CD patients.     

Split T cell tolerance against a self/tumor antigen: spontaneous CD4+ but not CD8+ T cell responses against p53 in cancer patients and healthy donors

Analyses of NY-ESO-1-specific spontaneous immune responses in cancer patients revealed that antibody and both CD4(+) and CD8(+) T cell responses were induced together in cancer patients. To explore whether such integrated immune responses are also spontaneously induced for other tumor antigens, we have evaluated antibody and T cell responses against self/tumor antigen p53 in ovarian cancer patients and healthy individuals. We found that 21% (64/298) of ovarian cancer patients but no healthy donors showed specific IgG responses against wild-type p53 protein. While none of 12 patients with high titer p53 antibody showed spontaneous p53-specific CD8(+) T cell responses following a single in vitro sensitization, significant p53-specific IFN-γ producing CD4(+) T cells were detected in 6 patients. Surprisingly, similar levels of p53-specific CD4(+) T cells but not CD8(+) T cells were also detected in 5/10 seronegative cancer patients and 9/12 healthy donors. Importantly, p53-specific CD4(+) T cells in healthy donors originated from a CD45RA(-) antigen-experienced T cell population and recognized naturally processed wild-type p53 protein. These results raise the possibility that p53-specific CD4(+) T cells reflect abnormalities in p53 occurring in normal individuals and that they may play a role in processes of immunosurveillance or immunoregulation of p53-related neoplastic events.     

Celiac sprue: correlation with murine T cell responses to wheat gliadin components

Celiac sprue is a disease in humans that is characterized by small intestinal mucosal injury and malabsorption. Dietary exposure to gliadin and similar proteins in rye and barley activates the disease in susceptible individuals. Celiac sprue appears to be the only disease with a marked HLA-association in which the proteins that activate the disease currently are well known. However, bread wheat gliadins are a complex mixture of proteins that contain at least 40 different components. In the present study we have purified the major gliadin components of Scout 66 wheat and used these proteins to examine murine T cell proliferative responses to gliadin. Differences in T cell proliferation stimulated by alpha-, beta-, gamma-, and omega-gliadins paralleled the known structural differences among these proteins. After priming with whole gliadin, the components that stimulated T cell proliferation were the same as those recognized to activate celiac sprue in humans. Studies with reduced and alkylated A-gliadin (i.e., S-methyl A-gliadin) suggested that epitopes determined by the native conformation of A-gliadin may be important in its interaction with T cells. By using three different A-gliadin peptides that span the entire molecule, T cell proliferative responses were shown to be stimulated predominantly by antigenic determinants on the NH2-terminal peptide.     

Intestinal epithelial antigen induces mucosal CD8 T cell tolerance, activation, and inflammatory response

Intestinal autoimmune diseases are thought to be associated with a breakdown in tolerance, leading to mucosal lymphocyte activation perhaps as a result of encounter with bacterium-derived Ag. To study mucosal CD8(+) T cell activation, tolerance, and polarization of autoimmune reactivity to self-Ag, we developed a novel (Fabpl(4x at -132)-OVA) transgenic mouse model expressing a truncated form of OVA in intestinal epithelia of the terminal ileum and colon. We found that OVA-specific CD8(+) T cells were partially tolerant to intestinal epithelium-derived OVA, because oral infection with Listeria monocytogenes-encoding OVA did not elicit an endogenous OVA-specific MHC class I tetramer(+)CD8(+) T cell response and IFN-gamma-, IL-4-, and IL-5-secreting T cells were decreased in the Peyer's patches, mesenteric lymph nodes, and intestinal mucosa of transgenic mice. Adoptive transfer of OVA-specific CD8(+) (OT-I) T cells resulted in their preferential expansion in the Peyer's patches and mesenteric lymph nodes and subsequently in the epithelia and lamina propria but failed to cause mucosal inflammation. Thus, CFSE-labeled OT-I cells greatly proliferated in these tissues by 5 days posttransfer. Strikingly, OT-I cell-transferred Fabpl(4x at -132)-OVA transgenic mice underwent a transient weight loss and developed a CD8(+) T cell-mediated acute enterocolitis 5 days after oral L. monocytogenes-encoding OVA infection. These findings indicate that intestinal epithelium-derived "self-Ag" gains access to the mucosal immune system, leading to Ag-specific T cell activation and clonal deletion. However, when Ag is presented in the context of bacterial infection, the associated inflammatory signals drive Ag-specific CD8(+) T cells to mediate intestinal immunopathology.     

Intraepithelial lymphocyte immunophenotype: a useful tool in the diagnosis of celiac disease

According to new ESPGHAN guidelines, gluten challenge is considered necessary when there is doubt about the initial diagnosis of celiac disease (CD). The main aim of this study was to quantify intraepithelial lymphocyte (IEL) immunophenotype on celiac patients on gluten-containing diet (GCD) compared to those on gluten-free diet (GFD). Another aim was to evaluate the clinical utility of IELs in the CD diagnosis, especially in selected patients on GFD where diagnostic uncertainty remains. IEL immunophenotype (TCRγδ and NK-like IELs) were studied by flow cytometry in 111 children with CD (81 children with CD on GCD and 30 celiac patients on GFD) and a control group (10 children). Duration of GFD was 5.4 ± 1.6 years. TCRγδ IELs in celiac patients receiving a GCD or GFD were significantly higher (p < 0.001) than in the control group. NK-like IELs in patients receiving a GCD or GFD were significantly lower than in the control group (p < 0.001). We observed a permanent decrease of NK-like IELs and an increment of TCRγδ IELs after following an adequate establishment and compliance of a long-term GFD in celiac patients. Recognition of IELs changes in the intestinal mucosa on celiac patients after long-term establishment of a GFD could constitute a useful tool for CD diagnosis in various situations: in which there is doubt about the initial diagnosis and repeat biopsy is necessary (avoiding the need of gluten challenges), and in those patients with symptoms/signs suggestive of CD who maintain a low gluten diet.     

HLA-B7-restricted EBV-specific CD8+ T cells are dysregulated in multiple sclerosis

It was hypothesized that the EBV-specific CD8(+) T cell response may be dysregulated in multiple sclerosis (MS) patients, possibly leading to a suboptimal control of this virus. To examine the CD8(+) T cell response in greater detail, we analyzed the HLA-A2-, HLA-B7-, and HLA-B8-restricted EBV- and CMV-specific CD8(+) T cell responses in a high number of MS patients and control subjects using tetramers. Content in cytolytic granules, as well as cytotoxic activity, of EBV- and CMV-specific CD8(+) T cells was assessed. We found that MS patients had a lower or a higher prevalence of HLA-A2 and HLA-B7, respectively. Using HLA class I tetramers in HLA-B7(+) MS patients, there was a higher prevalence of MS patients with HLA-B*0702/EBV(RPP)-specific CD8(+) T cells ex vivo. However, the magnitude of the HLA-B*0702/EBV(RPP)-specific and HLA-B*0702/CMV(TPR)-specific CD8(+) T cell response (i.e., the percentage of tetramer(+) CD8(+) T cells in a study subject harboring CD8(+) T cells specific for the given epitope) was lower in MS patients. No differences were found using other tetramers. After stimulation with the HLA-B*0702/EBV(RPP) peptide, the production of IL-2, perforin, and granzyme B and the cytotoxicity of HLA-B*0702/EBV(RPP)-specific CD8(+) T cells were decreased. Altogether, our findings suggest that the HLA-B*0702-restricted viral (in particular the EBV one)-specific CD8(+) T cell response is dysregulated in MS patients. This observation is particularly interesting knowing that the HLA-B7 allele is more frequently expressed in MS patients and considering that EBV is associated with MS.     

Gliadin regulates the NK-dendritic cell cross-talk by HLA-E surface stabilization

We analyzed the autologous NK cell interaction with gliadin-presenting dendritic cells. Gliadin is the known Ag priming the celiac disease (CD) pathogenesis. We demonstrate that gliadin prevents immature dendritic cells (iDCs) elimination by NK cells. Furthermore, cooperation between human NK cells-iDCs and T cells increases IFN-gamma production of anti-gliadin immune response. Gliadin fractions were analyzed for their capability to stabilize HLA-E molecules. The alpha and omega fractions conferred the protection from NK cell lysis to iDCs and increased their HLA-E expression. Gliadin pancreatic enzyme digest and a peptide derived from gliadin alpha increased HLA-E levels on murine RMA-S/HLA-E-transfected cells. Analysis of HLA-E expression in the small intestinal mucosa of gluten-containing diet celiac patients and organ culture experiments confirmed the in vitro data.     

Emergence and kinetics of simian immunodeficiency virus-specific CD8(+) T cells in the intestines of macaques during primary infection

In this report, three Mamu-A*01(+) rhesus macaques were examined to compare the emergence of simian immunodeficiency virus (SIV)-specific CD8(+) T cells in the intestines and blood in early SIV infection using a major histocompatibility complex class I tetramer complexed with the Gag(181-189) peptide. Fourteen days after intravenous inoculation with SIVmac251, large numbers of SIV Gag(181-189)-specific CD8(+) T cells were detected in the intestinal mucosa (3.1 to 11.5% of CD3(+) CD8(+) lymphocytes) as well as in the blood (3.1 to 13.4%) of all three macaques. By 21 days postinoculation, levels of tetramer-binding cells had dropped in both the intestines and blood. At day 63, however, levels of SIV Gag(181-189)-specific CD8(+) T cells in the intestines had rebounded in all three macaques to levels that were higher (8.6 to 18.7%) than those at day 21. In contrast, percentages of tetramer-binding cells in the peripheral blood remained comparatively stable (2.5 to 4.5%) at this time point. In summary, SIV Gag(181-189)-specific CD8(+) T cells appeared in both the intestinal mucosa and peripheral blood at a comparable rate and magnitude in primary SIV infection. Given that the intestine is a major site of early viral replication as well as the site where most of the total body lymphocyte pool resides, these data indicate that it is also an early and important site of development of antiviral immune responses.     

miRNA-regulated gene expression differs in celiac disease patients according to the age of presentation

Celiac disease is an intestinal disease which shows different symptoms and clinical manifestations among pediatric and adult patients. These variations could be imputable to age-related changes in gut architecture and intestinal immune system, which could be characterized by gene expression differences possibly regulated by miRNAs. We analyzed a panel of miRNAs and their target genes in duodenal biopsies of Marsh 3AB and 3C pediatric celiac patients, compared to controls. Moreover, to assess variation of expression in plasma samples, we evaluated circulating miRNA levels in controls and patients at diagnosis or on gluten-free diet. We detected a decreased miR-192-5p expression in celiac patients, but no variations in NOD2 and CXCL2, targets previously identified in adults. Conversely, we detected a significant increase in mRNA and protein levels of another target, MAD2L1, protein related to cell cycle control. miR-31-5p and miR-338-3p were down-regulated and their respective targets, FOXP3 and RUNX1, involved in Treg function, resulted up-regulated in celiac patients. Finally, we detected, in celiac patients, an increased expression of miR-21-5p, possibly caused by a regulatory loop with its putative target STAT3, which showed an increased activation in Marsh 3C patients. The analysis of plasma revealed a trend similar to that observed in biopsies, but in presence of gluten-free diet we could not detect circulating miRNAs values comparable to controls. miRNAs and their gene targets showed an altered expression in duodenal mucosa and plasma of celiac disease pediatric patients, and these alterations could be different from adult ones.

Electronic supplementary material

The online version of this article (doi:10.1007/s12263-015-0482-2) contains supplementary material, which is available to authorized users.     

Effect of celiac disease and gluten-free diet on growth hormone-binding protein, insulin-like growth factor-I, and insulin-like growth factor-binding proteins

Failure to thrive is common in children with celiac disease. As alterations in the growth hormone-insulin-like growth factor I (GH-IGF-I) growth axis have been reported in these patients, we studied the behavior of growth hormone-binding proteins (GH-BPs I and II), IGF-I and its binding proteins in 14 children with celiac disease, either before or after a 6-month gluten-free diet. GH-BP II levels were significantly lower in patients during the active phase of the disease than after the diet or in comparison with control subjects, appropriate for age and sex. There was no difference in the GH-BP-I levels of patients and controls, nor did they change after the diet. Blood levels of IGF-I and IGFBP-3 were reduced before the diet in all patients while ligand blotting showed that IGFBP-2 and 1 were increased. All of these parameters normalized after the gluten-free diet. IGFBP-4 was not greatly influenced by the disease. Furthermore, we found a significant, positive correlation between GH-BP II and IGF-I or IGFBP-3 levels. The height standard deviation scores and body mass indices of the patients improved significantly after the diet. The body mass index significantly and positively correlated with GH-BP II, IGF-I or IGFBP-3 levels. In conclusion, our data show that celiac children had multiple alterations in the growth axis during the active phase of the disease which disappeared during the gluten-free diet.     

Phenotypic and functional heterogeneity of EBV epitope-specific CD8+ T cells

High frequencies of EBV-specific CD8(+) T cells have been detected during acute EBV infection, yet persistent infection inevitably results. To address this issue, we characterized the phenotype and function of epitope-specific CD8(+) T cell populations from presentation with acute through latent infection. Considerable phenotypic and functional heterogeneity within, as well as between, two different epitope-specific populations was observed over time following acute infection. B7 EBV-encoded nuclear Ag (EBNA)-3A-specific CD8(+) T cells expressed only CD45RO from acute through latent EBV infection. A2 BMLF-1-specific CD8(+) T cells expressed CD45RO during acute infection and either CD45RA or CD45RO during latent EBV infection. This difference in CD45 isoform expression between the two epitope-specific populations did not translate into differences in perforin content, the ability to produce IFN-gamma, or the ability to proliferate in response to Ag in vitro. In individuals with latent EBV infection, the frequencies of A2 BMLF-1- or B7 EBNA-3A-specific CD8(+) T cells that expressed CD45RA, CD45RO, CD62 ligand, CCR7, and perforin were stable over time. However, the expression of CD62 ligand and CCR7 was significantly higher among EBNA-3A-specific CD8(+) T cells than among BMLF-1-specific CD8(+) T cells. Further work is necessary to understand how phenotypic and functional differences between EBV epitope-specific CD8(+) T cells are related to the biology of the virus and to the equilibrium between the virus and the host during persistent infection.     

Autoimmunity to both proinsulin and IGRP is required for diabetes in nonobese diabetic 8.3 TCR transgenic mice

T cells specific for proinsulin and islet-specific glucose-6-phosphatase catalytic subunit related protein (IGRP) induce diabetes in nonobese diabetic (NOD) mice. TCR transgenic mice with CD8(+) T cells specific for IGRP(206-214) (NOD8.3 mice) develop accelerated diabetes that requires CD4(+) T cell help. We previously showed that immune responses against proinsulin are necessary for IGRP(206-214)-specific CD8(+) T cells to expand. In this study, we show that diabetes development is dramatically reduced in NOD8.3 mice crossed to NOD mice tolerant to proinsulin (NOD-PI mice). This indicates that immunity to proinsulin is even required in the great majority of NOD8.3 mice that have a pre-existing repertoire of IGRP(206-214)-specific cells. However, protection from diabetes could be overcome by inducing islet inflammation either by a single dose of streptozotocin or anti-CD40 agonist Ab treatment. This suggests that islet inflammation can substitute for proinsulin-specific CD4(+) T cell help to activate IGRP(206-214)-specific T cells.