Lectin binding sites in duodeno-jejunal mucosae from coeliac children

The distribution pattern of some labelled lectins (WGA, SBA, PNA, DBA, ConA, LCA) has been investigated in small intestinal mucosa of coeliac children (119 cases) and normal (short stature) and pathological (postenteritis syndrome) controls. The oligosaccharide side chains of glycoproteins present in the cytoplasm and in the brush border of enterocytes, goblet cells and luminal crypt surface have been revealed. The most important differences in lectin binding reactivity between coeliac and controls mucosae are discussed.     

PROgnosticating COeliac patieNts SUrvivaL: The PROCONSUL Score

Introduction

It has been shown that mortality rates of coeliac patients correlate with age at diagnosis of coeliac disease, diagnostic delay for coeliac disease, pattern of clinical presentation and HLA typing. Our aim was to create a tool that identifies coeliac patients at higher risk of developing complications.

Methods

To identify predictors of complications in patients with coeliac disease, we organised an observational multicenter case-control study based on a retrospective collection of clinical data. Clinical data from 116 cases (patients with complicated coeliac disease) and 181 controls (coeliac patients without any complications) were collected from seven European centres. For each case, one or two controls, matched to cases according to the year of assessment, gender and age, were selected. Diagnostic delay, pattern of clinical presentation, HLA typing and age at diagnosis were used as predictors.

Results

Differences between cases and controls were detected for diagnostic delay and classical presentation. Conditional logistic models based on these statistically different predictors allowed the development of a score system. Tertiles analysis showed a relationship between score and risk of developing complications.

Discussion

A score that shows the risk of a newly diagnosed coeliac patient developing complications was devised for the first time. This will make it possible to set up the follow-up of coeliac patients with great benefits not only for their health but also for management of economic resources.

Conclusions

We think that our results are very encouraging and represent the first attempt to build a prognostic score for coeliac patients.     

Differences between the fecal microbiota of coeliac infants and healthy controls

Coeliac disease (CD) is an immune-mediated enteropathy with a multifactorial aetiology, characterized by chronic inflammation of the small intestinal mucosa. Although evidence suggests that the gut microbiota contributes to other chronic inflammatory disorders, its possible role in CD has not been determined. In this study, the composition of the fecal microbiota of coeliac children and age-matched controls was investigated by culture-dependent and -independent methodologies, using fluorescent in situ hybridization (FISH). The levels of Bacteroides, Clostridium and Staphylococcus were significantly higher (p < 0.05) in fecal samples from coeliac patients than in healthy subjects when analysed by culture methods. The numbers of Bacteroides-Prevotella, Clostridium histolyticum, Eubacterium rectale-C. coccoides, Atopobium, and sulfate reducing bacterial groups were also significantly higher (p < 0.05) in fecal samples from coeliac infants when analysed by FISH. The counts of Bifidobacterium tended to be higher in healthy controls by the two type of analysis but the differences were not significant. This is the first report on the identification of the specific bacterial groups responsible for alterations in the intestinal microecology of children with active CD. The bacterial pattern detected in coeliac patients, correlates with the epidemiological data and metabolic deviations associated with CD, and involve bacterial groups link to other chronic inflammatory disorders.     

Differences in faecal bacterial communities in coeliac and healthy children as detected by PCR and denaturing gradient gel electrophoresis

Coeliac disease (CD) is a chronic inflammatory disorder of the small intestinal mucosa. Scientific evidence supports a role of the gut microbiota in chronic inflammatory disorders; yet information is not specifically available for CD. In this study, a comparative denaturing gradient gel electrophoresis analysis of faecal samples from coeliac children and age-matched controls was carried out. The diversity of the faecal microbiota was significantly higher in coeliac children than in healthy controls. The presence of the species Lactobacillus curvatus, Leuconostoc mesenteroides and Leuconostoc carnosum was characteristic of coeliac patients, while that of the Lactobacillus casei group was characteristic of healthy controls. The Bifidobacterium population showed a significantly higher species diversity in healthy children than in coeliacs. In healthy children, this population was characterized by the presence of Bifidobacterium adolescentis. Overall, the results highlighted the need for further characterization of the microbiota in coeliac patients, and suggested a potential role of probiotics and/or prebiotics in restoring their gut microbial balance.     

Body content of selenium in coeliac disease.

Concentrations of selenium in whole blood, plasma, and leucocytes were determined in 16 patients with coeliac disease confirmed by biopsy and 32 controls. All the patients were clinically well and receiving gluten free diets. The concentrations of selenium were significantly lower in the leucocytes, blood, and plasma of patients compared with controls, probably indicating a decrease in the body content of selenium. A high incidence of malignancy in coeliac disease has been reported. As a protective role for selenium against cancer has been postulated, the importance of this unexpected observation of lowered tissue concentrations of selenium requires further investigation.     

HLA-DR typing in coeliac disease: evidence for genetic heterogeneity.

Sixty nine propositi from a family study of coeliac disease were typed for HLA-DR antigens. Sixty three (91%) were found to carry the antigen DR3, which was a significantly greater proportion (p = 9.6 X 10(-24] than among the 168 controls (26%). Concurrently 42 children with the disease were DR typed. Not only was the frequency of DR3 significantly increased in these patients (86% versus 26% in controls; p = 3.1 X 10(-12] but so also was the frequency of DR7 (patients 60%, controls 29%; p = 5.8 X 10(-4]. When those propositi whose coeliac disease presented before the age of 20 were combined with the childhood coeliac group and a comparison made between these patients and the remainder of the propositi, all of whom presented when they were older than 20, the childhood onset group had a significant excess of DR7 (p = 2.2 X 10(-3] and a significant deficiency of DR2 (p = 3.5 X 10(-3]. These findings indicate that childhood coeliac disease and adult coeliac disease are genetically heterogeneous.     

Alpha gliadin antibody levels: a serological test for coeliac disease.

The diagnostic value in coeliac disease of circulating antibodies to casein, crude gliadin, and alpha gliadin was assessed using an adaption of the enzyme linked immunosorbent assay system. alpha Gliadin was the only antigen which consistently separated 26 patients with untreated coeliac disease from 26 normal controls and 13 patients with chronic inflammatory bowel disease. The mean assay index for the 26 patients was 3.1 (SD 1.2) compared with 1.05 (0.5) for the normal controls and 1.1 (0.6) for patients with chronic inflammatory bowel disease. The alpha gliadin antibody levels of six patients with coeliac disease who had maintained a gluten free diet for at least two years were not significantly higher than normal (1.0 (0.4)). The validity of the test was determined in 90 consecutive patients who were being investigated for the presence of coeliac disease. Levels of alpha gliadin antibody were raised in 36 out of 44 patients found to have histologically proved coeliac disease and in six out of 46 subjects whose jejunal mucosa was normal. Serial alpha gliadin concentrations were measured in 12 patients with coeliac disease who had repeat jejunal biopsies performed six months after starting a gluten free diet. The levels of antibody fell in seven of the eight patients whose jejunal mucosa improved on maintaining the diet. They remained raised in four patients who did not adhere to the diet and whose mucosa did not improve. Although a test measuring alpha gliadin antibodies is unlikely to replace jejunal biopsy in the diagnosis of coeliac disease it may be useful in screening for the disease among outpatients.     

Antibodies to bovine beta-casein in diabetes and other autoimmune diseases.

Cow's milk is thought to be an environmental trigger for autoimmune response in Type 1 diabetes. In the present study, our aim was to investigate the antibody response to bovine beta-casein in different immune- and non-immune-mediated diseases and to establish whether such an antibody response is specific to Type 1 diabetes. We measured antibodies to bovine beta-casein using an enzyme-linked immunosorbent assay in a total of 519 sera from subjects as follows: 71 patients with Type 1 diabetes, 33 patients with coeliac disease, 100 patients with latent autoimmune diabetes in adults (LADA), 50 patients with autoimmune thyroid disease (ATD), 50 patients with Type 2 diabetes, 24 patients with multiple sclerosis (MS), and 3 different groups of controls (n = 191). Significantly increased levels of antibodies to beta-casein were found in patients with Type 1 diabetes, coeliac disease and in LADA compared to age-matched controls (p = 0.01, p = 0.02 and p = 0.01, respectively). No differences were observed in beta-casein antibody titres between patients with other disease conditions (MS, and ATD) and age-matched controls. The highest antibody response to beta-casein in Type 1 diabetic patients and in patients with coeliac disease could reflect the gut mucosal immune disorders common to Type 1 diabetes and coeliac disease. Furthermore, the elevated beta-casein antibody levels found in LADA patients suggest that the antibody response to this protein may be relevant in autoimmune diabetes.     

Simultaneous demonstration of mucins and lysozyme in duodeno-jejunal biopsies of coeliac infants

Using a sequential method for the simultaneous demonstration of lysozyme and mucins, the possible existence of a transitional cell intermediate in form between Paneth and goblet cells has been investigated in childhood coeliac disease. Mixed staining for both mucins and lysozyme has not been encountered within intestinal cells of affected mucosae and controls.     

DNA synthesis by jejunal mucosa in responsive and non-responsive coeliac disease.

DNA synthesis by jejunal biopsy specimen from patients with coeliac disease and from controls was measured by an organ culture technique. The rate of synthesis in the mucosa of patients with untreated coeliac disease was almost eight times that in normal mucosa. Patients whose jejunal mucosa remained flat despite prolonged gluten withdrawal showed a rate of DNA synthesis significantly lower than that of the untreated patients, while those whose jejunal mucosa had responded to gluten withdrawal showed a rate similar to that of normal subjects. Impaired enterocyte production in nonresponsive coeliac disease may be responsible for the failure to regenerate villi after gluten withdrawal.     

CD69 expression on alpha-gliadin-specific T cells in coeliac disease

Coeliac disease (CD) is a T-cell mediated immunological disease of the small intestine which is triggered in susceptible individuals by ingestion of gluten. The pathogenic mechanism of coeliac disease, and the role that alpha-gliadin specific T cells play in mucosal lesions and their involvement in peripheral blood is not yet explained at all. Previous studies have reported proliferative response to alpha-gliadin measured with the classic assay of 3HTdR incorporation. We analysed the activation antigen CD69 on T cells from CD patients and normal individuals following stimulation with alpha-gliadin and different antigens (tetanus toxoid, peptides unrelated to gliadin and PHA). CD69 coexpression with T cell CD3+ and proliferation marker Ki67 was evaluated with time. CD69 coexpression with T cell CD3+, CD4+ and CD8+ was also evaluated. It was found that peripheral blood mononuclear cells (PBMC) of coeliac patients increased their percentage of CD69 positive T cells when stimulated with alpha-gliadin, in comparison with cells from controls. Significant T cell activation was found only in subjects not treated with the gluten free diet; a positive response was found also in two coeliac patients with selective IgA deficiency, anti-endomisium negative, without circulating IgA anti alpha-gliadin or anti-tissue transglutaminase antibodies. The CD69 expression after stimulation was compared with the standard method of 3HTdR incorporation. Our data show that CD69 expression is useful to asses a specific T cell response to alpha-gliadin in coeliac disease. in a very short time. Moreover, the method allows to investigate T cell response at the lymphocyte subsets level, which represents a useful tool in the diagnosis of coeliac disease.     

Fingerprint Changes in Coeliac Disease

Study of the fingerprints of 73 patients with coeliac disease, taken carefully, showed changes varying between moderate epidermal ridge atrophy and actual loss of fingerprint patterns. Of the patients 63 had these abnormalities, compared with 3 out of 485 controls. A high degree of correlation existed between ridge atrophy and changes in the clinical state of patients with coeliac disease.     

Persistent Changes in Circulating and Intestinal γδ T Cell Subsets,Invariant Natural Killer T Cells and Mucosal-Associated Invariant T Cells in Children and Adults with Coeliac Disease

Coeliac disease is a chronic small intestinal immune-mediated enteropathy precipitated by exposure to dietary gluten in genetically predisposed individuals. The only current therapy is a lifelong gluten free diet. While much work has focused on the gliadin-specific adaptive immune response in coeliac disease, little is understood about the involvement of the innate immune system. Here we used multi-colour flow cytometry to determine the number and frequency of γδ T cells (Vδ1, Vδ2 and Vδ3 subsets), natural killer cells, CD56⁺ T cells, invariant NKT cells, and mucosal associated invariant T cells, in blood and duodenum from adults and children with coeliac disease and healthy matched controls. All circulating innate lymphocyte populations were significantly decreased in adult, but not paediatric coeliac donors, when compared with healthy controls. Within the normal small intestine, we noted that Vδ3 cells were the most abundant γδ T cell type in the adult epithelium and lamina propria, and in the paediatric lamina propria. In contrast, patients with coeliac disease showed skewing toward a predominant Vδ1 profile, observed for both adult and paediatric coeliac disease cohorts, particularly within the gut epithelium. This was concurrent with decreases in all other gut lymphocyte subsets, suggesting a specific involvement of Vδ1 cells in coeliac disease pathogenesis. Further analysis showed that γδ T cells isolated from the coeliac gut display an activated, effector memory phenotype, and retain the ability to rapidly respond to in vitro stimulation. A profound loss of CD56 expression in all lymphocyte populations was noted in the coeliac gut. These findings demonstrate a sustained aberrant innate lymphocyte profile in coeliac disease patients of all ages, persisting even after elimination of gluten from the diet. This may lead to impaired immunity, and could potentially account for the increased incidence of autoimmune co-morbidity.     

Correctness of diagnosis of celiac disease based on personal data]

A way of coeliac disease diagnosis was assessed in 348 children, Only 14% of patients has been examined according to ESPGAN recommendations and in the proper time. Diagnosis has been completed in 123 patients, and coeliac disease has been diagnosed only in 38 of them. The I biopsy has not been performed in 96 children, the II biopsy--in 136 children, gluten-free diet has not been observed in 105 children after the I biopsy. Other causes of diagnostic failures included: prolongation of the consecutive stages and improper histological evaluation, change of biopsies order, erroneous clinical evaluation. Diagnosis according to ESPGAN recommendations is practically very difficult and time consuming. Immunological markers of coeliac disease (IgA EmA, ARA) should lead to simplification and adjustment of those recommendations in Poland.     

The use of Cellomics to study enterocyte cytoskeletal proteins in coeliac disease patients

Coeliac disease is characterised by inflammation of small intestinal mucosa accompanied by abnormal villous architecture. It is now accepted that some patients with positive coeliac serology tests may have minor mucosal lesions that may not be apparent on routine histopathological analysis. The aim of the study was to perform detailed examination of enterocyte morphology and cytoskeletal structures using a high content analysis technology. Duodenal biopsies from 14 untreated and 10 treated coeliac patients and from 20 non-coeliac controls were examined. Tissue sections from six patients (study group subjects) before and after the development of gluten-sensitive enteropathy were also investigated. Immunohistochemical studies were performed on paraffin-embedded sections using an anti-α-tubulin antibody. Significant differences in enterocyte morphology and intracellular cytoskeletal structures were demonstrated in patients with proven coeliac disease and in the study group subjects. These changes were present in study group biopsies before evidence of enteropathy, as assessed by routine microscopy. This is the first study to demonstrate detailed characteristics of enterocyte morphology in coeliac patients using a high content analysis approach. The use of this technology allows a quantitative analysis of enterocyte intracellular structures from routine biopsy material and permits detection of subtle changes that precede the characteristic histological lesion.     

The Incidence of Other Gastroenterological Disease following Diagnosis of Irritable Bowel Syndrome in the UK: A Cohort Study

Background

Guidelines recommend Irritable Bowel Syndrome (IBS) diagnosis and management in primary care with minimal investigations; however little evidence exists regarding risk of organic gastrointestinal conditions following diagnosis of IBS and how such risks vary over the long term. This study assesses excess incidence of coeliac disease, inflammatory bowel disease (IBD) and colorectal cancer (CRC) and variation with age and time after IBS diagnosis.

Methods

IBS patients and controls were identified within the UK Clinical Practice Research Dataset. Incidence rates were calculated and stratified by age and time since IBS diagnosis with incident rate ratios generated.

Results

Fifteen years after IBS diagnosis there is a significant cumulative excess incidence of coeliac disease, IBD and CRC in IBS of 3.7% compared to 1.7% in controls. For every 10000 patient years, IBS patients experienced an additional 4 diagnoses of coeliac disease, 13 of IBD and 4 CRCs. In each condition peak excess incidence was in the 6 months following diagnosis. After one year, increased incidence of coeliac disease remained consistent without variation by age. IBD incidence fell slowly, with higher rates in those under 30. CRC incidence was increased only in patients aged 30 to 74 during the first 5 years.

Conclusion

Some IBS patients later receive organic gastrointestinal diagnoses, with the early excess incidence likely detected during diagnostic investigation at the time of IBS diagnosis. More than 5 years after IBS diagnosis there is no increased risk of CRC compared to the general population, but a small excess risk of coeliac disease and IBD persists. Overall, though our findings provide reassurance that non-specialists, especially those in primary care, are unlikely to be missing an organic condition in the majority of their patients. This suggests that current guidelines suggesting avoidance of universal referral for these patients are appropriate.     

The pathogenesis of coeliac disease

Coeliac disease is a chronic enteropathy caused by intolerance to gluten proteins. The true prevalence of this condition is greater than previously thought, with increasing numbers of 'silent' cases being diagnosed. Untreated coeliac disease is associated with significant morbidity and increased mortality. There have been a number of advances in our understanding of the pathogenesis of coeliac disease, in particular the mechanisms whereby gluten epitopes are processed, become modified by tissue transglutaminase (tTG) and then interact with HLA restricted T cells. An improved understanding of the immune response to gluten is likely to lead to the development of novel strategies for the treatment of coeliac disease.     

Screening of coeliac disease in undetected adults and patients diagnosed with irritable bowel syndrome in Riyadh,Saudi Arabia

The present study is to determine the prevalence and implication of coeliac disease (CD) among adult Saudis and compared to those with diagnosed irritable bowel syndrome. This prospective study was conducted among 980 adults. Out of that, 482 subjects (staff and students of Riyadh Health Science College) were designated as control cohorts for undetected coeliac disease. Furthermore, another contingent of 498 subjects diagnosed with irritable bowel syndrome (IBS) at Prince Salman Hospital and Al-Iman General Hospital also constituted a segment of the overall initial 1020 subjects. Both cases and control were tested for serological markers of coeliac disease (tissues transglutaminase (tTGAs) and endomysial autoantibody (EMAs) and were confirmed by histopathology test. All the positive for cases of coeliac disease were screened for iron deficiency anaemia, Vitamin D deficiency, and osteoporosis and weight assessment. The percentage of coeliac disease in control subjects and patients diagnosed with irritable bowel syndrome (IBS) were found to be 1.9% and 9.6% respectively, about 38% of the total coeliac disease patients are among females of middle age (20–39-years) and 16% of the males in the same age range. Whereas, 20% and 25% of all coeliac disease cases with ages of 40–59 were remarked as females and males respectively. The identical nature and overlap of symptoms of the two conditions could possibly result in misdiagnosis of coeliac diseases or over-diagnosis of irritable bowel syndrome. The findings of the study might also give considerable implications of the disease in the nutritional level which is noticeable.     

Predictive value for coeliac disease of antibodies to gliadin,endomysium, and jejunum in patients attending for jejunal biopsy.

OBJECTIVE--To investigate the extent to which the detection of antibodies to gliadin, endomysium, and jejunum predicts the eventual diagnosis of coeliac disease according to the revised ESPGAN diagnostic criteria in a group of patients in whom there is a high suspicion of coeliac disease. DESIGN--Clinical assessment and laboratory analysis of patients with suspected coeliac disease. SETTING--Gastroenterology department of teaching hospital. PATIENTS--96 adults with suspected coeliac disease attending for jejunal biopsy. MAIN OUTCOME MEASURES--Diagnosis of coeliac disease with the revised criteria of the European Society of Paediatric Gastroenterology and Nutrition in patients with and without antibodies associated with coeliac disease. RESULTS--28 patients had a clinical diagnosis of coeliac disease, seven of other gastrointestinal diseases, and 12 of miscellaneous diseases; 49 had no diagnosis. Gliadin IgA detected by ELISA was found in all patients with coeliac disease and none of those without, giving a sensitivity, specificity, positive and negative predictive values, and predictive efficiency of 100% for diagnosing coeliac disease within the group. Endomysial IgA was found in 25 (89%) patients with coeliac disease and jejunal IgA in 21 (75%); neither IgA was found in patients without coeliac disease. CONCLUSION--Detection of gliadin IgA by ELISA and to a lesser extent the endomysial IgA should allow better selection of patients for jejunal biopsy and thus make diagnosing coeliac disease simpler and more efficient.     

Catecholamines and atrial natriuretic factor in Dahl and spontaneously hypertensive rats

We have previously demonstrated two different catecholaminergic patterns in genetic and experimental hypertension: a hyperdopaminergic state in spontaneously hypertensive (Okamoto) rats (SHR) and a hypernoradrenergic state in salt-sensitive Dahl rats. Plasma immunoreactive atrial natriuretic factor (IR ANF) concentrations increase in both models as a response to hypertension. To distinguish between the genetic and acquired components of these abnormalities, we measured adrenal dopamine-beta-hydroxylase (D beta H) activity and coeliac ganglionic atrial natriuretic factor (ANF) like immunoreactivity in the two animal strains. While adrenal D beta H activity was increased in Dahl S rats, it was diminished in SHR in the prehypertensive as well as in the hypertensive stages. In the hypertensive stage, the ANF-like immunoreactivity in the coeliac ganglia was lower in the Dahl S group but higher in SHR than in their respective normotensive controls; there were no changes in these animals when they were prehypertensive. Differences in D beta H activity, which determines the fine tuning of sympathoadrenomedullary catecholamine synthesis may account for the inheritance of mechanisms resulting in salt-sensitive hypertension (as in SHR) or salt-dependent hypertension (as in Dahl salt-sensitive rats). In contrast, plasma IR ANF concentrations may reflect a defense mechanism against hypertension. However ANF-like immunoreactivity in coeliac ganglia does not follow its plasma concentrations and changes in different directions in the two hypertensive strains; it may reflect a neuromodulatory function of ANF in the ganglionic neurotransmission and different implications of this role of ANF in the two hypertensive models.