Mucosal Damage In Celiac Disease Research Articles

Missing Insight Into T and B Cell Responses in Dermatitis Herpetiformis.

Dermatitis herpetiformis is a cutaneous form of celiac disease manifesting as an itching rash typically on the elbows, knees and buttocks. It is driven by the ingestion of gluten-containing cereals and characterized by granular deposits of immunoglobulin A in the papillary dermis. These antibodies target transglutaminase (TG) 3 and in the majority of patients they are also found in circulation. The circulating antibodies disappear and skin symptoms resolve as a result of gluten-free diet but the cutaneous anti-TG3 IgA deposits may persist for several years. In dermatitis herpetiformis, plasma cells secreting antibodies against TG3 are located in the intestinal mucosa similarly to those producing TG2 antibodies characteristic for celiac disease. In fact, both TG2- and TG3-specific plasma cells and gluten responsive T cells are found in dermatitis herpetiformis patients but the interplay between these cell populations is unknown. The small bowel mucosal damage in celiac disease is believed to be mediated by co-operation of cytotoxic intraepithelial T cells and the inflammatory milieu contributed by gluten-reactive CD4+ T cells, whereas the skin lesions in dermatitis herpetiformis appear to be devoid of gluten reactive T cells. Thus, how celiac disease-type intestinal T and B cell responses develop into an autoimmune condition affecting the skin is still incompletely understood. Finally, the skin and small bowel lesions may reappear upon reintroduction of gluten in patients treated with gluten-free diet but virtually nothing is known about the long-lived B cell and memory T cell populations activating in response to dietary gluten in dermatitis herpetiformis.

PARK7 Diminishes Oxidative Stress-Induced Mucosal Damage in Celiac Disease.

Coeliac disease (CD) is a chronic, immune-mediated small intestinal enteropathy, accompanied with gluten-triggered oxidative damage of duodenal mucosa. Previously, our research group reported an increased mucosal level of the antioxidant protein Parkinson's disease 7 (PARK7) in children with CD. In the present study, we investigated the role of increased PARK7 level on the epithelial cell and mucosal integrity of the small intestine. The presence of PARK7 was investigated using immunofluorescent staining on duodenal mucosa of children with CD and on FHs74Int duodenal epithelial cells. To investigate the role of oxidative stress, FHs74Int cells were treated with H2O2 in the absence or presence of Comp23, a PARK7-binding compound. Intracellular accumulation of reactive oxygen species (ROS) was determined by DCFDA-based assay. Cell viability was measured by MTT, LDH, and Annexin V apoptosis assays. Disruption of cytoskeleton and cell adhesion was investigated by immunofluorescence staining and by real-time RT PCR. Effect of PARK7 on mucosal permeability was investigated ex vivo using intestinal sacs derived from control and Comp-23-pretreated mice. Comp23 treatment reduced the H2O2-induced intracellular accumulation of ROS, thus preserving the integrity of the cytoskeleton and also the viability of the FHs74Int cells. Accordingly, Comp23 treatment increased the expression of antioxidants (NRF2, TRX1, GCLC, HMOX1, NQO1), cell-cycle regulators (TP53, CDKN1A, PCNA, BCL2, BAX), and cell adhesion molecules (ZO1, CDH1, VCL, ITGB5) of H2O2-treated cells. Pretreatment with Comp23 considerably decreased the small intestinal permeability. In this study, we demonstrate that PARK7-binding Comp23 reduces the oxidative damage of duodenal epithelial cells, via increased expression of NRF2- and P53-regulated genes. Our results suggest that PARK7 plays a significant role in the maintenance of mucosal integrity in CD.

Phenotypic shift of small intestinal intra-epithelial type 1 innate lymphoid cells in celiac disease is associated with enhanced cytotoxic potential

The small intestinal (SI) epithelium harbors a heterogeneous population of lymphocytes that mediate mucosal damage and repair in celiac disease (CD). The composition and roles of human proximal SI intra-epithelial innate lymphoid cells (ILCs), and their alterations in CD, are not well understood. We report that duodenal intra-epithelial ILCs predominantly consist of natural killer (NK)p44+ CD127- cytotoxic ILC1s and NKp44- CD127+ helper ILC1s, while ILC3s only represent a minor population. In patients with newly diagnosed or active CD (ACD) and refractory CD type 1 (RCD I), the frequency of SI NKp44+ ILCs is decreased, with restoration of NKp44+ ILC frequency observed in patients adhering to a gluten-free diet who show evidence of mucosal healing. Moreover, the frequency of SI NKp44- ILCs is increased in ACD and RCD I patients and correlates with the severity of villous atrophy and epithelial damage, as assessed by serum levels of fatty acid binding protein 2 (FABP2). We show that the ILC alterations in CD represent a phenotypic shift of cytotoxic ILC1s rather than an increase in helper ILC1s or transdifferentiation of ILC1sto ILC3s, and activation-induced loss of NKp44 by cytotoxic ILC1s is associated with increased interferon (IFN)-γ expression and release of lytic granules. These findings suggest that intra-epithelial NKp44- CD127- cytotoxic ILC1s may contribute to mucosal damage in CD.

Contributions of HLA haplotypes, IL8 level and Toxoplasma gondii infection in defining celiac disease's phenotypes

BackgroundIt is not clear why some patients with coeliac disease (CD) present with severe symptoms and small intestinal mucosal damages while others present with milder symptoms and no frank enteropathy. There is no study to assess the associated factors with mild/severe symptoms and enteropathy. The terminologies like latent, silent and potential are difficult to use and are unrepresentative. In the present study we describe coeliac disease’s phenotypes based on HLA haplotypes, IL8 production and past infection with Toxoplasma gondii (T. gondii) infection.MethodsIn this case-control study, sera originating from 150 healthy subjects and 150 patients diagnosed with CD during the years 2013–14 were analyzed for the presence of antibodies specific T. gondii of the IgG and IgM subclasses. The level of IL8 were measured and HLA-DQ2 and HLA-DQ8 alleles were genotyped. The correlation between these parameters and the damages in intestinal mucosal were assessed using an accepted histopathological classification.ResultsHigh levels of IgG antibodies against T. gondii were found in the sera of control group compared to the CD group (52.6% vs. 39.4%, P = 0.02). Mean serum levels of IL8 was significantly higher in CD patients compared with control (P ≤ 0.05). By comparing the level of anti- T. gondii IgG and mucosal damage in celiac disease, we found a significant relationship between the severity of mucosal damages and anti- T. gondii IgG level (P = 0.02). No correlation was detected between Toxoplasma gondii infection and types of HLA (P > 0.05). However, patients with severely abnormal histology carried HLA-DQ2 risk alleles (92 patients (61%)) more often than the controls and those with mild histological abnormalities.ConclusionsCD patients with severe histological changes had more often Toxoplasma gondii infection than those affected with mild histological features. This suggests that CD’s phenotypes are correlated to additional factors like infections and to particular HLA DQ2 alleles that may need additional investigations and potentially will require additional treatment.

Challenging in the celiac disease diagnosis; Prague consensus

Celiac disease (CD) was initially a real enigma. There were many questions about what immunogenetics markers could influence the immune response (1). Immunogenetic studies provided important contributions to improve the understanding of critical pathogenetic factors at molecular levels (DQ2, DQ8) (2). An important discovery was the recognition of more precise serological markers for CD, specifically anti-endomysial and anti-tissue transglutaminase antibodies leading to an exploration of their increasingly important role in clinical diagnosis of CD (3). Our aim is to explore new and old challenges with unanswered questions in CD diagnosis. The last Prague consensus report challenges Re-evaluation of CD diagnostic with intestinal biopsy is required in this consesus (4). However, many patients with CD are still inaccurately diagnosed (or remain undiagnosed), even in referral populations whom were evaluated at tertiary care centers: the diagnosis of CD was confirmed in only 64 patients from 107 patients with previous diagnosis of CD (5). Recently, it was noted that even for some experts, the quality of duodenal bulb biopsies was unsatisfactory. This fact may create confusion and false-positive diagnoses which seem to be increasing in the pediatric population(6). However, histopathological classifications of CD have been debated for decades and some investigators have recently expressed concern regarding specific architectural changes, the degree of change and their relevance: Marsh, Marsh modified (Oberhuber), or Corazza classification. (7, 8). As immunologically-based assays evolve, the authors of this paper believe that serological markers are more useful for screening purposes in CD compared to small intestinal mucosal biopsy alone. At present, the gold standard for diagnosis is still small intestinal mucosal biopsy. Although data is still needed, it is conceivable that serological markers may eventually even become more useful for diagnostic purposes in CD compared to small intestinal mucosal biopsy alone (9). But an obvious serological algorithm is not specified. At present, the IgA anti-tissue transglutaminase antibody test appears to be the most sensitive and specific markers of CD screening (10) but a minority of CD patients are seronegative (11). The main task of gastroenterologists was to recognize untreated CD. In countries with weaker economies (i.e., the so-called developing nations), it is generally believed that the rate of diagnosis is low (12). Controversies remain in areas that include application of screening methods and evaluation of high risk groups for the CD, as well as diagnostic tests that lead to a final diagnosis of the CD (13). In addition, a number of immunologically-related challenges in gastroenterology remain for those involved in a CD care. IL-21 was lower in potential CD, than in active CD, so a key role for IL-21 in the progression of mucosal damage in CD needs to be further elucidated (14). Furthermore, HLA DQ8 in Southwest Asia, South America and the Middle East was positive in 49% (n=69) of CD patients and 13% (n=21) of control group, respectively. In conclusion, HLA DQ8 was found to be significantly higher in these geographical regions compared to European countries (15). However, HLA-DQ2 and/or -DQ8 expression: 38.1% in northern, 31.4% in northeastern, and 36.4% in southern India, did not appear to correlate with CD prevalence: 8.53/1,000 and 3.70/1,000 in northern, 4.66/1,000 and 3.92/1,000 in northeastern, and 0.11/1,000 and 1.22/1,000 in the southern India (16). A number of other associated immune-mediated or autoimmune disorders may co-exist with CD, possibly owing to a common genetic background (17). So, novel diagnostic antibodies for CD are neccesary (18).

Soluble Syndecan-1: A Novel Biomarker of Small Bowel Mucosal Damage in Children with Celiac Disease.

Syndecan-1 (SDC1) is essential for maintaining normal epithelial barrier. Shedding of SDC1 ectodomain, reflected by serum soluble syndecan-1 (SSDC1) levels, is regulated by inflammation. Increased intestinal permeability plays a central role in celiac disease (CD). The association between SSDC1 levels and mucosal damage in CD has not been evaluated. To evaluate serum SSDC1 levels in children with CD and to determine its relationship with histological grading classified by modified Marsh criteria. This is a cross-sectional, pilot study, in which serum SSDC1 was analyzed by ELISA in a cohort of 49 untreated children with CD and 15 children with nonspecific abdominal pain (AP). CD was diagnosed based on positive celiac serology and small intestinal biopsy. SSDC1 levels at the time of biopsy were correlated with Marsh grading. Controls were defined by AP, negative celiac serology, normal upper endoscopy, and small intestinal biopsies. SSDC1 levels were significantly higher in CD patients compared to AP controls (116.2±161 vs. 41.3±17.5ng/ml, respectively, p<0.01). SSDC1 levels were significantly higher in patients with Marsh 3c lesion compared to AP controls (170.6±201 vs. 41.3±17.5ng/ml, respectively, p<0.05). SSDC1 concentrations displayed a significant correlation with mucosal damage defined by Marsh (r=0.39, p<0.05). This is the first study demonstrating elevated levels of serum SSDC1 in children with CD. Our results suggest that SSDC1 is a potentially novel marker of intestinal mucosal damage in patients with CD. Its applicability as a surrogate biomarker in CD remains to be determined.

TNFA Haplotype Genetic Testing Improves HLA in Estimating the Risk of Celiac Disease in Children.

BackgroundTNF-α and IFN-γ play a role in the development of mucosal damage in celiac disease (CD). Polymorphisms of TNFA and IFNG genes, as well as of the TNFRSF1A gene, encoding the TNF-α receptor 1, might underlie different inter-individual disease susceptibility over a common HLA risk background. The aims of this study were to ascertain whether five SNPs in the TNFA promoter (-1031T>C,-857C>T,-376G>A,-308G>A,-238G>A), sequence variants of the TNFRSF1A gene and IFNG +874A>T polymorphism are associated with CD in a HLA independent manner.Methods511 children (244 CD, 267 controls) were genotyped for HLA, TNFA and INFG (Real Time PCR). TNFRSF1A variants were studied (DHPLC and sequence).ResultsOnly the rare TNFA-1031C (OR=0.65, 95% CI:0.44-0.95), -857T (OR=0.42, 95% CI:0.27-0.65), -376A (OR=2.25, 95% CI:1.12-4.51) and -308A (OR=4.76, 95% CI:3.12-7.26) alleles were significantly associated with CD. One TNFRSF1A variant was identified (c.625+10A>G, rs1800693), but not associated with CD. The CD-correlated TNFA SNPs resulted in six haplotypes. Two haplotypes were control-associated (CCGG and TTGG) and three were CD-associated (CCAG, TCGA and CCGA). The seventeen inferred haplotype combinations were grouped (A to E) based on their frequencies among CD. Binary logistic regression analysis documented a strong association between CD and HLA (OR for intermediate risk haplotypes=178; 95% CI:24-1317; OR for high risk haplotypes=2752; 95% CI:287-26387), but also an HLA-independent correlation between CD and TNFA haplotype combination groups. The CD risk for patients carrying an intermediate risk HLA haplotype could be sub-stratified by TNFA haplotype combinations.Conclusion TNFA promoter haplotypes associate with CD independently from HLA. We suggest that their evaluation might enhance the accuracy in estimating the CD genetic risk.

Pediatric Typical Vs. Atypical Celiac Disease: Correlation of Duodenal Histology With Tissue Transglutaminase Levels

Background: Both the clinical presentation and the degree of mucosal damage in coeliac disease vary greatly. In view of conflicting information as to whether the mode of presentation correlates with the degree of villous atrophy, we reviewed a 103 pediatric patient with coeliac disease. This study also aimed to determine the correlation of anti tTG level and the degree of mucosal damage in typical and atypical cases. Methods: The medical records of patients with newly diagnosed CD during the period of 2006 - 2010 were retrospectively reviewed. Results: A total of 103 patients fulfilled the established CD diagnostic criteria (mean age: 6.6 years; mediana: 5 years). A typical CD presentation was observed for 66/103 ( 64 .1% ) children vs atypical 37/103 (35.9%). The levels of tTG antibody were correlated significantly with Marsh types in the entire population by Spearman rank correlation (r = 0.661, P < 0.0001), and separately for typical(r = 0.633, P < 0.001) and atypical (r = 0.574, P < 0.0001) groups. Histopathological evaluation of intestinal biopsies revealed total or subtotal VA in 44 patients (66.6%) in the typical group, and 24 patients (64.8%) in the atypical group. Partial VA was observed in 5 patients (7.5%) in the typical group and 1 (2.7%) patients in atypical group. There was no correlation of mode of presentation (typical and atypical) with the degree of villous atrophy by a non-parametric Mann-Whitney U test Z = ­ -1.437, P = 0.151. Conclusion: There is a positive correlation between tTG antibody serum level and duodenal histopathology in typical as in atypical pediatric CD. The degree of villous atrophy did not correlate with the mode of presentation, indicating that factors other than the degree of villous atrophy must account for typical simptoms in coeliac disease. doi: http://dx.doi.org/10.4021/ijcp57w

Optical coherence tomography in children with coeliac disease

Coeliac disease (CD) is a gluten-sensitive enteropathy occurringingeneticallypredisposedindividualsandcharacterisedbychronicinflammationofthesmallintestinalmucosathatgraduallyleadstovillous atrophy. It typically results in malabsorption of most nutri-ents, but its clinical expression is highly variable [1].The gold standard for the diagnosis of CD is the histologicaldemonstration of characteristic pathological changes that are cat-egorised according to the Marsh classification and modificationsthereof [2]. As the small bowel mucosal damage in CD can be dis-tributed in a patchy fashion, with parts of the mucosa apparentlynormalintermingledwithothersseverelydiseased(‘patchyvillousatrophy’),itisimportantthatfourtosixbiopsies,includingaduode-nal bulb biopsy, are taken from the proximal small intestine duringupper endoscopy [3].The diagnosis of CD may, therefore, also depend on the endo-scopic appearance of the duodenal mucosa. Several endoscopicmarkers related to CD have been identified and include featuressuch as a nodular and/or a mosaic pattern of bulb and duodenalmucosa, reduction or loss of duodenal folds, scalloping of valvu-laeconniventes,visibilityofsubmucosalvessels[4].Thesemarkersare often used both to determine whether duodenal biopsies areindicated and to target the sites of the biopsy. Among several stud-ies, the overall sensitivity and specificity of endoscopic markersof CD vary from 6% to 94% and from 83% to 100%, respectively[4]. In clinical practice, recognising any suspicious mucosal fea-tures for CD should alert the endoscopist to the need to biopsy themucosa. However, the rather low sensitivity of these features sug-gests that their absence does not exclude an underlying CD andbiopsies should always be obtained when CD is suspected, eitherclinically or by serological assays. In individuals undergoing upperendoscopyforsymptomsotherthanmalabsorption(e.g.functionaldyspepsia, acid-related disease) duodenal biopsies might not betaken and diagnosis of CD missed. On the other hand searching forCDinsuchpatientsisnotrealistic,consideringthesignificantcostsof pathological processing.

Increased expression of mucosal addressin cell adhesion molecule 1 in the duodenum of patients with active celiac disease is associated with depletion of integrin α4β7-positive T cells in blood

Mucosal addressin cell adhesion molecule 1, expressed on gut endothelial cells, in conjunction with integrin alpha(4)beta(7), expressed on lymphocytes, is critical in lymphocyte homing to the gut. The mucosal addressin cell adhesion molecule 1/integrin alpha(4)beta(7) pathway is involved in the pathogenesis of chronic intestinal inflammation by recruiting lymphocytes into inflamed gut. We explored the duodenal expression of mucosal addressin cell adhesion molecule 1 and the peripheral T-cell expression of integrin alpha(4)beta(7) in patients with celiac disease. Duodenal biopsies and a peripheral blood sample were collected from 15 celiac patients, before and after 12 months of gluten-free diet, and from 12 control subjects. Treated celiac biopsies were cultured with peptic-tryptic digest of gliadin and/or an anti-interferon alpha neutralizing antibody. Mucosal addressin cell adhesion molecule 1 was determined by confocal immunofluorescence microscopy and immunoblotting. Integrin beta(7)-positive T cells were analyzed by flow cytometry. Mucosal addressin cell adhesion molecule 1 expression was significantly higher in active celiac disease than in normal mucosa. After gluten-free diet, a dramatic reduction of mucosal addressin cell adhesion molecule 1 was also observed. No difference was seen between patients with celiac disease after treatment and controls. Ex vivo peptic-tryptic digest of gliadin challenge induced a marked increase of mucosal addressin cell adhesion molecule 1 expression. Blocking interferon alpha inhibited the peptic-tryptic digest of gliadin-induced mucosal addressin cell adhesion molecule 1 overexpression. The percentage of circulating beta(7)-positive T cells was significantly lower in untreated celiac disease in comparison to controls but normalized after gluten-free diet. Mucosal addressin cell adhesion molecule 1 is strongly up-regulated in active celiac disease dependent on interferon alpha and is associated with peripheral depletion of integrin alpha(4)beta(7)-expressing T cells. We conclude that mucosal addressin cell adhesion molecule 1 may represent an important determinant for the generation of mucosal damage in celiac disease.

Serological Responses to Microbial Antigens in Celiac Disease Patients During a Gluten-Free Diet

Immunoglobulin A (IgA) autoantibodies to tissue transglutaminase (tTG) are commonly used for screening and diagnosing of celiac disease (CD). Seroreactivity for anti-Saccharomyces cerevisiae antibody (ASCA) and bacterial antigens have also been detected in CD patients. The aim of this study was to examine prospectively serologic responses to microbial targets in adult CD patients at the time of diagnosis and during a gluten-free diet (GFD). Further, we wanted to evaluate whether these serologic specificities could provide new tools for the follow-up of CD patients. Data on 55 adult biopsy-proven CD patients were available for follow-up study. Upper gastrointestinal endoscopy was performed on all patients. Sera from patients were tested for antibodies to tTG and ASCA and additionally analyzed with IgA enzyme-linked immunosorbent assays to Pseudomonas fluorescens-associated sequence, I2, and to a Bacteroides caccae TonB-linked outer membrane protein, OmpW. At the time of diagnosis, 91% of CD cases were positive for tTG and 49% for ASCA; positive seroreactivity to I2 was found in 86% and to OmpW in 60% of CD patients at the time of diagnosis. The frequency of seropositivity and serum levels of these antibodies decreased during GFD. Moreover, we found that the decline in the serum levels was significant in all of these markers (p < 0.005). Interestingly, we also found that serum levels of ASCA correlated with the grade of mucosal morphology (p = 0.021), as the ASCA serum levels declined in accordance with mucosal healing. Commensal enteric bacteria seem to play a role in the small intestinal mucosal damage in CD. This was proven by the serological responses to different microbial antigens shown in this study. Serum levels of ASCA, anti-I2, and anti-OmpW antibodies decreased significantly during GFD, indicating that these serologic markers are gluten dependent in CD patients. These specificities could provide new tools in the follow-up of CD patients.

Elevated Serum Anti-Saccharomyces cerevisiae, Anti-I2 and Anti-OmpW Antibody Levels in Patients with Suspicion of Celiac Disease

Expression of anti-Saccharomyces cerevisiae antibodies (ASCA) identifies patients and individuals at risk for Crohn's disease and has also been reported in 40-60% of celiac disease (CD) cases, suggesting a role of host response to enteric microbiota in the development of inflammatory lesions. In this prospective study in patients with suspicion of CD, we evaluate the frequency and association of ASCA to serological responses for other host microbial targets formally associated with Crohn's disease, including the Pseudomonas fluorescens associated sequence I2 and a Bacteroides caccae TonB-linked outer membrane protein, OmpW. Small bowel mucosal biopsies were taken from 242 patients with suspicion of CD, their sera were tested for antibodies to tissue transglutaminase (tTG), ASCA, I2, and OmpW. Eighty adult healthy blood donors were used as controls. The diagnosis of CD was confirmed on biopsy in 134 cases. The occurrence of ASCA and I2 positivity was significantly higher in adult CD patients than in patients with non-CD disease. Anti-I2 levels in the sera were significantly higher in adult CD patients than in non-CD disease or the controls and anti-OmpW levels in CD and non-CD patients when compared to controls. Positive seroreactivity to OmpW seemed to increase with age. Of the CD patients, 90% were seropositive for at least one microbial antigen tested. This study demonstrates a mosaic of disease-related serological responses to microbial antigens in patients with CD. Immune responses to commensal enteric bacteria may play a role in the small intestine mucosal damage in CD.

Histopathologic Diagnosis of Celiac Disease in Children Without Clinical Evidence of Malabsorption

Diverse clinical and pathologic experiences seem to have led to the idea that celiac disease is a spectrum in both categories. Conflicting results emerging from different reports have produced a large amount of confusion on the subject. This article discussed histopathology findings in 10 children with positive autoantibodies for celiac disease but without clinical evidence of malabsorption. The patients were evaluated following a detailed video-endoscopic study sampling the proximal (first and second) and distal (third and fourth) duodenal parts separately and processed apart. The procedure consistently revealed advanced villous atrophy in the proximal duodenal mucosa associated with mild to absent involvement of the distal segments. The data here presented favor the interpretation that (1) the presence of autoantibodies for celiac disease is always associated with mucosal damage, (2) mucosal damage in the absence of malabsorption is always evident in the proximal duodenum, and (3) the mucosal biopsy in search for the telltale damage needs to be done in separate samples of proximal and distal duodenal mucosa. This procedure may result in a better understanding of the dynamics of mucosal damage in celiac disease.

Lack of correlation of degree of villous atrophy with severity of clinical presentation of coeliac disease

Background and aim Both the clinical presentation and the degree of mucosal damage in coeliac disease vary greatly. In view of conflicting information as to whether the mode of presentation correlates with the degree of villous atrophy, we reviewed a large cohort of patients with coeliac disease. Patients and methods We correlated mode of presentation (classical, diarrhoea predominant or atypical/silent) with histology of duodenal biopsies and examined their trends over time. Results The cohort consisted of 499 adults, mean age 44.1 years, 68% females. The majority had silent coeliac disease (56%) and total villous atrophy (65%). There was no correlation of mode of presentation with the degree of villous atrophy ( p = 0.25). Sixty-eight percent of females and 58% of males had a severe villous atrophy ( p = 0.052). There was a significant trend over time for a greater proportion of patients presenting as atypical/silent coeliac disease and having partial villous atrophy, though the majority still had total villous atrophy. Conclusions Among our patients the degree of villous atrophy in duodenal biopsies did not correlate with the mode of presentation, indicating that factors other than the degree of villous atrophy must account for diarrhoea in coeliac disease.

The liver in celiac disease.

The liver in celiac disease.

Electrogenic Ion Transport in Duodenal Biopsies from Children with Coeliac Disease

Background: The aim of this work was to study whether different degrees of duodenal mucosal damage in coeliac disease (CD) influenced secretory responses to well-known secretagogues. Methods: Intestinal biopsies from 53 patients in different clinical phases of CD and 34 patients without CD and with normal histology were studied in a modified Ussing chamber. The electrogenic responses--with and without pretreatment with indomethacin--to prostaglandin E 2 , aminophylline, dibutyryl-cAMP and acetylcholine were followed by continuous measurements of potential difference (Pd). Tissue resistance and epithelial current (I m ) were calculated. Results: All secretagogues induced a similar pattern, with a greater increase in Pd and I m in biopsies with villous atrophy compared to controls. The electrophysiological response was correlated to the serum levels of IgA gliadin antibodies. The most prominent electrophysiological increase was found in the biopsies with partial atrophy. Indomethacin had a greater impact on the response to secretagogues in the more severely damaged mucosa. Conclusion: The electrogenic secretory response in the proximal small intestine was enhanced and related to serum levels of IgA gliadin antibodies and to the degree of mucosal damage in biopsies from children with active CD compared to controls.

Sensitivity of a hyperosmolar or “low”-osmolar test solution for sugar absorption in recognizing small intestinal mucosal damage in coeliac disease

Reliability of differential sugar absorption tests is hampered by a lack of standardization of the content and osmolarity of the test solutions. We evaluated the effect of osmolarity of the test solution of the sugar absorption test on the 5 hour urine excretion of orally administered lactulose and mannitol. A group of 28 controls and 14 coeliacs, with villous atrophy grade 11 to IV, ingested a hyperosmolar sugar absorption test solution and a “low”-osmolar solution, respectively. After an overnight fast, each subject ingested hyperosmolar sugar absorption test solution (2 g mannitol, 5 g lactulose and 40 g sucrose/100 ml (around 1560 mmol/1]]. After two days, this procedure was repeated with low-osmolar solution (2 g mannitol and 5 g lactulose/100 ml (around 375 mmol/1). The influence of the sequence of the tests on the results had previously been excluded. All urine from the 5 h-period following ingestion of the test solution was collected. To calculate the low-osmolar solution ratio, samples were analysed for lactulose and mannitol concentrations by gas chromatography. The sensitivity of hyperosmolar SAT solution and low-osmolar solution for the detection of mucosal abnormalities in coeliacs was 64% and 43%, respectively. In conclusion, a hyperosmolar solution discriminates better between normal and damaged mucosa of the small bowel such as villous atrophy due to a relative increase in permeability for lactulose.

Selective identification by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry of different types of gluten in foods made with cereal mixtures

The gluten toxic fractions responsible for the mucosal damage in coeliac disease (CD), so-called gliadins, hordeins, secalins and avenins from a large number (30–40) of wheat, barley, rye and oats cultivars respectively, have been mass analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF–MS). Gliadin, secalin and avenin characteristic mass profiles are nearly identical amongst distinct cultivars from the corresponding cereal, while hordeins profiles show more variability depending on the particular barley cultivar. On the basis of these four distinguishable characteristic mass patterns spreading within the 20 000–40 000 Da range, MALDI-TOF-MS has permitted the direct and simultaneous visualization of gliadins, hordeins, secalins and avenins in foods elaborated with cereal mixtures of wheat, barley, rye and oats. This capacity has been demonstrated by mass analyzing foods made with these four cereals in varying ratios. Thus MALDI-TOF-MS can be preliminarily established as a unique system with the ability to discriminate the specific type of gluten toxic fractions present in food samples.

THE IN VITRO-INFLUENCE OF GLIADIN PEPTIDES ON THE HYDROLASES OF THE DUODENAL BRUSH BORDER MEMBRANE IN COELIAC DISEASE IN REMISSION

Aims: The molecular mechanism inducing the mucosal damage in coeliac disease (CD) is still unknown. Gliadin causes villus atrophy in CD, and patients on a gluten-free diet recover completely. On the basis of the organ culture of intestinal biopsies we developed an in vitro-system for CD in remission. We used this model to verify our hypothesis postulating a correlation between the mucosal damage caused by gliadin and the biosynthesis of the intestinal hydrolases. Besides we measured the enzyme activity to prove whether this level is afflicted additionally or alternatively. Methods: Intestinal biopsies from 14 coeliac patients in remission (following a gluten-free diet for at least 2 years) and 23 healthy children (control group) were organ cultured for 24 hours in presence or absence of gliadin peptides. A peptictryptic digest of maize prolamine operated as the non-toxic control. The effect of these additives on the enzyme activity of the disaccharidases was determined by a modificated method of Dahlqvist. The influence on the biosynthesis of several hydrolases (LPH, SI, MGA, ACE, APN, DPP IV) was studied by biosynthetical protein-labelling during organ culture, followed by immunoprecipitation, SDS-PAGE, autoradiography and quantified by densitometric scanning. Results: In our model we demonstrated a rapid and specific gliadin-induced reduction of the biosynthesis of all studied hydrolases in biopsies from coeliac patients on gluten-free diet. This phenomenon was neither seen after incubation with maize prolamine nor in healthy children. We could not show an effect of gliadin on the disaccharidase activities after 24 hours of incubation. Because of beginning necrosis in the biopsies a longer incubation was not possible, so we were not able to examine later effects. Conclusions: Our model is suitable for studying the in vitro-situation of CD in remission. Moreover, our results indicate a fast effect of gliadin peptides on inhibiting the biosynthesis of hydrolases of duodenal brush border membranes. It remains unclear whether the gliadin-induced reduction primarily influences the protein biosynthesis or has to be interpreted as a secondary effect of an impaired mRNA synthesis. We suppose that the effect occurs at the translational or transcriptional level. The early modifications within 24 hours of incubation are hardly to explain by activities of the immune system. More likely the results suggest a toxic effect of gliadin peptides on the intestinal mucosa preceeding immunological changes mediated by cytokines.

Reappraisal of the 'lectin hypothesis' in the aetiopathogenesis of coeliac disease.

The agglutinating properties of a crude gluten digest, purified gliadin fractions and established plant lectins were investigated using mammalian erythrocytes, rat enterocytes and normal and coeliac human enterocytes as the target systems. Gliadin preparations failed to cause agglutination of any of the cells tested, whereas established pure plant lectins were active cell agglutinins. These studies indicate that gliadin peptides do not interact with intestinal cells in a polyvalent, lectin-like manner and as such cannot be regarded as true lectins. Mucosal damage in coeliac disease is unlikely therefore to be related to lectin-like activity of gliadin.