Coeliac disease (CD) is a disorder resulting from interactions between diet, genome and immunity. This research seeks to further our understanding of the pathology of CD in regard to its secondary effects on the diversity of enteral microflora via changes in immune tolerance. It proposes that enteral mucosal pro-inflammatory change in CD is associated with a decrease in microbial diversity whilst remission from inflammation may result in an increase in enteral microbial diversity that could contribute to the restoration of tolerance. The first study analyses whether remission from active CD is associated with change in generic enteral microbial diversity by assessing people at diagnosis and following their response to gluten exclusion. A comparison is made to people without CD consuming a ’normal diet’. DGGE profiling of faecal microflora in subjects with CD at diagnosis (confirmed by serology and by duodenal biopsy) and over three consecutive months on a gluten-free diet (GFD) was performed and profiles were compared with those of age and gender matched control subjects taken at monthly intervals. Diversity of faecal microflora (measured as Simpsons Index ) was significantly lower in people with CD than in control subjects. It was possible to distinguish the profiles of coeliac subjects at diagnosis from those obtained after three months on a GFD but it was not possible to distinguish between the samples from control subjects taken at monthly intervals. The profiles of CD subjects after three months on a GFD were more dissimilar to those of the control subjects than those obtained prior to dietary treatment, chiefly on the basis of three bands that were not found in the faeces of any control subjects. The second study analyses dietary intake to determine if a lack of nutrients at diagnosis (before institution of a GFD) and at monthly intervals for three consecutive months post diagnosis (on a GFD) exists, as it is known that CD is associated with nutrient deficiencies resulting from malabsorption due to intestinal inflammation and damage. Subjects completed a customised food questionnaire at each sampling period. Dietary intake was analysed using Foodworks Professional 2007. Significant differences were identified in gluten, starch and carbohydrate intake but not in other macronutrients. Contrary to established literature, these analyses identified few significant differences in micronutrient intake within coeliac subjects over time, however, significant differences were found in iron and sodium.

Magister Scientiae - MSc / Coeliac disease (CD) is a gluten intolerance condition that results in the flattening of the villi, which line the bowel. It is the most common cause of malabsorption of food nutrients. This inability to absorb sufficient levels of nutrients causes many of the common symptoms experienced by CD patients. Some of the symptoms, which lead to an increase in mortality rate, include chronic diarrhea, fatigue, iron-deficient anemia and osteoporosis. People with CD have higher than normal levels of certain antibodies in their blood. Thus, the concentration of anti-transglutaminase antibody (anti-tTG) in human sera is an important analytical marker for the diagnosis of CD. An immunosensor is a type of biosensor that has an antigen or antibody fragment as its biological recognition component. The specificity of the molecular recognition of antigens by antibodies to form a stable complex is the basis of immunosensor technology. In this work, overoxidized polypyrrole (OvoxPpy) was electrosynthesized as a noval sensor platform on a glassy carbon electrode (GCE). The OvoxPpy was then doped with gold-nanoparticles (GNP) by electrodeposition using cyclic voltammetry to form GNP|OvoxPpy||GCE electrode system. Morphology and size of the GNP|OvoxPpy||GCE nanocomposite were determined using scanning electron microscopy. The electrochemical immunosensor for anti-tTG antibodies was prepared by immobilizing transglutaminase antigen (tTG-antigen) onto the GNP|OvoxPpy||GCE by drop coating and allowed to incubate for 2 hrs. The electrochemical characterization of the nanocomposite platform and immunosensor were studied by voltammetry and electrochemical impedance spectroscopy (EIS). Charge transfer resistance, Rct (obtained from EIS data fitting) of [Fe (CN)6]3-/4- redox probe was used as the analytical parameter for measuring the interfacial kinetics which occurred as a result of the bio-recognition event (affinitive binding) between the tTG-antigen and anti-tTG antibody. Rct was found to increase with increased concentration of the antibody as a result of the reluctance to the flow of redox probe charge across the interface. Antibody concentration as low as in 1:4000 dilutions was detected. / South Africa

This diploma thesis deals with theme "Quality evaluation of gluten-free food, concretely gluten-free pastries". Theoretical part is concerned about coeliac disease in general, their symptoms, forms and diagnostic. There is described basic principal of coeliac disease and approachable gluten-free food. Then the basics of sensory analyse are described. Practical part is focused on finding suitable gluten-free flour and baking samples of bread based on recipe with care about improving nutritional value. Baked bread samples, made from commixture of gluten-free flour with enrichment of different amount of flaxseed, were evaluated with suitable methods of sensory analysis. This analyse shows which sample is the most preferred and based on those information it is proposed to enlarge offer in gluten-free pastry.

Yes / Objectives. Coeliac disease (CD) is an autoimmune condition managed by a lifelong therapeutic gluten-free diet. Previous research suggests that the chronicity of CD, the limitations imposed by the gluten-free diet, and the risk of other associated diseases can have a negative impact on health-related quality of life (HRQoL) and psychological well-being. The aim of this study was to explore the illness perceptions and self-efficacy beliefs of adults with CD in the United Kingdom and to report their subjective levels of HRQoL and psychological well-being. Design. The study employed a cross-sectional postal questionnaire design. Method. Participants (n= 288) were adults with CD recruited via Coeliac UK. Measures of well-being, HRQoL, self-efficacy, illness perceptions, and dietary self-management were analysed. Preliminary descriptive and univariate procedures were employed before bivariate tests of association or difference were carried out. Backward stepwise multiple regression analysis was used to investigate the predictive strength of variables on well-being, quality of life, and self-efficacy. Logistic regression was used to look at the influence of variables on adherence. Results. Results indicate that HRQoL and psychological well-being were comparable to those found in previous related studies. Participants with weak beliefs in the serious consequences of CD and poorer emotional reactions to the condition had a greater likelihood of having enhanced HRQoL, improved psychological well-being, and higher self-efficacy. Strong beliefs in personal control and a greater perceived understanding of CD were associated with greater self-efficacy. Conclusions. Perceived self-efficacy and illness perceptions could play a role in informing psychological interventions for individuals with CD.

<p>This study, based on daily recordings of infant feeding, comprised 506 infants from Uppsala, Sweden. All mothers had had previous breastfeeding experience of at least 4 months, and were planning to breastfeed the index child for ≥6 months.</p><p>Among exclusively breastfed infants there were wide variations in breastfeeding frequency and suckling duration per 24 hours both between infants and in the individual infant over time in the first 6 months. Most infants had an average of 1.0-2.9 feeds per night. Infants using a pacifier had fewer feeds and a shorter total suckling duration per 24 hours, and stopped breastfeeding earlier than infants not using a pacifier. These associations were not found for thumb sucking.</p><p>Accustoming the infants to solids was a lengthy process, the longer the younger the infant at introduction, and was associated with small changes in pattern and duration of breastfeeding. In contrast, formula was usually given in large amounts from the beginning, and when formula was given regularly the daily breastfeeding frequency and suckling duration declined swiftly. The younger an infant at the start of regular formula feeds, the shorter the breastfeeding duration. Occasional formula feeds did not affect the breastfeeding duration.</p><p>It is important for health personnel and parents to keep in mind that exclusively breastfed infants are not a homogeneous group, but rather members of distinct 'breastfeeding entities'. Moreover, if the aim is to introduce other foods 'under the protection of breast milk' it is important to realise that formula is also 'another food' and needs to be treated as such.</p>

<p>A diagnosis of coeliac disease (CD) in adults relies on the presence of a structurally abnormal intestinal mucosa, followed by a clear clinical remission on a gluten-free diet. There is a clear need for a rapid, simple, safe and sensitive method to determine the type and intensity of inflammation in the gut mucosa in clinical practice. The overall aims of our studies were to develop and evaluate a new technique, “the mucosal patch technique”, to characterize rectal local inflammatory process after rectal food challenge in patients with CD<b>. In study 1</b> we evaluated the potential of the new technique. The technique was well tolerated and easily applied. Pronounced neutrophil and eosinophil involvement in ulcerative colitis (UC) was demonstrated. With the high sensitivity of the technique, low-degree mucosal neutrophil activation could also be quantified in patients with collagen colitis,UC in clinical remission and in patients with irritable bowel syndrome. <b>In study 2 and 3</b> the aim was to elucidate the dynamics of the rectal inflammatory response and nitric oxide (NO) production after rectal gluten challenge. We found a pronounced neutrophil activation in coeliac patients after rectal gluten challenge. This activation was apparent 4 hours after challenge and remains for at least 48 hours. A more modest eosinophil activation started 1-2 hours later and remained at least for 48 hours. The biphasic pattern of neutrophil and eosinonphil activation after challenge suggests a biphasic inflammatory reaction. The activation of neutrophils and eosinophils precedes a pronounced enhancement of mucosal NO production. Some of our coeliac patients displayed signs of an inflammatory reaction after rectal corn gluten challenge. <b>In study 4</b> the aim was to investigate the local inflammatory reaction to gluten and cow’s milk protein in CD patients in remission. The findings indicate that not only gluten sensitivity but also cow’s milk (CM) protein sensitivity is common in CD. The data support the hypothesis that CM sensitivity may contribute to persistent symptoms in coeliac patients on gluten-free diet.</p>

This study, based on daily recordings of infant feeding, comprised 506 infants from Uppsala, Sweden. All mothers had had previous breastfeeding experience of at least 4 months, and were planning to breastfeed the index child for ≥6 months. Among exclusively breastfed infants there were wide variations in breastfeeding frequency and suckling duration per 24 hours both between infants and in the individual infant over time in the first 6 months. Most infants had an average of 1.0-2.9 feeds per night. Infants using a pacifier had fewer feeds and a shorter total suckling duration per 24 hours, and stopped breastfeeding earlier than infants not using a pacifier. These associations were not found for thumb sucking. Accustoming the infants to solids was a lengthy process, the longer the younger the infant at introduction, and was associated with small changes in pattern and duration of breastfeeding. In contrast, formula was usually given in large amounts from the beginning, and when formula was given regularly the daily breastfeeding frequency and suckling duration declined swiftly. The younger an infant at the start of regular formula feeds, the shorter the breastfeeding duration. Occasional formula feeds did not affect the breastfeeding duration. It is important for health personnel and parents to keep in mind that exclusively breastfed infants are not a homogeneous group, but rather members of distinct 'breastfeeding entities'. Moreover, if the aim is to introduce other foods 'under the protection of breast milk' it is important to realise that formula is also 'another food' and needs to be treated as such.

A diagnosis of coeliac disease (CD) in adults relies on the presence of a structurally abnormal intestinal mucosa, followed by a clear clinical remission on a gluten-free diet. There is a clear need for a rapid, simple, safe and sensitive method to determine the type and intensity of inflammation in the gut mucosa in clinical practice. The overall aims of our studies were to develop and evaluate a new technique, “the mucosal patch technique”, to characterize rectal local inflammatory process after rectal food challenge in patients with CD<b>. In study 1</b> we evaluated the potential of the new technique. The technique was well tolerated and easily applied. Pronounced neutrophil and eosinophil involvement in ulcerative colitis (UC) was demonstrated. With the high sensitivity of the technique, low-degree mucosal neutrophil activation could also be quantified in patients with collagen colitis,UC in clinical remission and in patients with irritable bowel syndrome. <b>In study 2 and 3</b> the aim was to elucidate the dynamics of the rectal inflammatory response and nitric oxide (NO) production after rectal gluten challenge. We found a pronounced neutrophil activation in coeliac patients after rectal gluten challenge. This activation was apparent 4 hours after challenge and remains for at least 48 hours. A more modest eosinophil activation started 1-2 hours later and remained at least for 48 hours. The biphasic pattern of neutrophil and eosinonphil activation after challenge suggests a biphasic inflammatory reaction. The activation of neutrophils and eosinophils precedes a pronounced enhancement of mucosal NO production. Some of our coeliac patients displayed signs of an inflammatory reaction after rectal corn gluten challenge. <b>In study 4</b> the aim was to investigate the local inflammatory reaction to gluten and cow’s milk protein in CD patients in remission. The findings indicate that not only gluten sensitivity but also cow’s milk (CM) protein sensitivity is common in CD. The data support the hypothesis that CM sensitivity may contribute to persistent symptoms in coeliac patients on gluten-free diet.

La maladie cœliaque est une entéropathie inflammatoire chronique se développant chez des individus génétiquement prédisposés par l’expression des haplotypes HLA-DQ2 ou HLA-DQ8 et survenant suite à la consommation de gluten. Elle se caractérise par le développement d’une atrophie des villosités de la muqueuse intestinale débouchant sur un syndrome de malabsorption alimentaire. La seule thérapie actuelle est le suivi d’une diète sans gluten mais cette éviction totale du gluten n’est pas toujours efficace et est lourde en concessions. Il est par conséquent urgent de développer des thérapies alternatives mais ce domaine constitue un pipeline évoluant lentement, notamment suite à l’absence d’un modèle animal pertinent et complet sur le plan physiologique. L’objectif de cette thèse doctorale est de répondre à ce besoin crucial en développant un modèle murin capable de récapituler les caractéristiques de la maladie. Le chapitre 1 dresse le portrait de la maladie en quatre parties amenant progressivement le lecteur dans les détails de sa pathogenèse. Cette introduction débute par un rappel sur la physiologie et l’immunité intestinale puis elle définit la face clinique de la maladie. Ensuite, le lecteur évolue dans une partie plus détaillée de la pathogenèse aidant au discernement de ses acteurs cellulaires et moléculaires. Finalement, elle se termine par une revue de la littérature sur les actuels modèles animaux. Le chapitre 2 brossent les objectifs de la thèse sur base de données clés de la littérature, notamment, les patients présentent au minimum une copie de l’halplotype HLA-DQ2 ou HLA-DQ8 et plus des deux-tiers sur-expriment la cytokine pro-inflammatoire interleukine-15 au niveau de leur muqueuse intestinale. Il est donc raisonnable de penser qu’ensemble, le gluten, l’haplotype HLA et l’interleukine-15 contribuent activement à la pathogenèse. Bien que soupçonnés, leurs rôles et interactions nécessitent l’apport de preuves tangibles in vivo. Le chapitre 3 détaille ainsi ces interactions démontrées à l’aide du développement de notre nouveau modèle murin. Ce dernier est caractérisé par la surexpression de l’interleukine-15 dans l’épithélium et dans la lamina propria intestinale et par l’expression de l’haplotype HLA-DQ8. Ce travail démontre que l’exposition de cette souris au gluten s’accompagne d’une atrophie villositaire et de la signature complète de la maladie, tant sur le plan sérologique, cellulaire que transcriptionnel. Nous démontrons que la surexpression simultanée de l’interleukine-15 dans les deux compartiments de la muqueuse intestinale que sont la lamina propria et l’épithélium est une condition sine qua non au développement de l’atrophie. Aussi, cette étude permet de mettre en lumière la nécessité des cellules T CD4+ et de l’interféron-gamma dans l’activation des lymphocytes intraépithéliaux et le développement de l’atrophie. Finalement, cette recherche établit le rôle central joué par l’haplotype HLA-DQ8 et par l’enzyme transglutaminase II tissulaire dans la survenue de ces lésions. De manière générale, les résultats issus de ce modèle et présentés au chapitre 3 reflètent toute la complexité des interactions entre le gluten, la génétique et l’IL-15 dans le développement de la maladie cœliaque. Enfin, le chapitre 4 apporte une conclusion à ce travail et le chapitre 5 discute des futures directions envisagées pour ce modèle préclinique. Ce dernier va sans doute contribuer à une meilleure compréhension de la maladie cœliaque et permettre l’identification de potentielles cibles thérapeutiques. / Coeliac disease is a chronic inflammatory enteropathy characterized by autoimmune features. This disease occurs in genetically predisposed individuals expressing HLA-DQ2 or HLA-DQ8 haplotypes and is triggered following gluten consumption. The disease is characterized by the development of intestinal villous atrophy leading to malabsorption. The only current therapy is the adherence to a gluten-free diet, but the diet is not always effective and is heavy in concessions. Therefore, the development of alternative therapies is urgent but is a slowly evolving pipeline, mainly due to the absence of a physiologically relevant animal model. The aim of this thesis is to answer this unmet need by developing an animal model capable of recapitulating the main characteristics of the disease. Chapter 1 depicts a portrait of the disease in four points, gradually leading the reader into the details of its pathogenesis. This introduction begins with a review of the physiology and intestinal immunity and then draws a clinical portrait of the disease. Third, the reader evolves in a more detailed part of the pathogenesis helping him to discern its cellular and molecular actors. Finally, the introduction ends with a review of the literature on current animal models. Chapter 2 outlines the thesis objectives based on key data from the literature, in particular, patients present at least one copy of the HLA-DQ2 or HLA-DQ8 haplotype and more than two-thirds over-express the proinflammatory cytokine interleukin-15 at the level of their intestinal mucosa. It is therefore reasonable to hypothesize that gluten, HLA haplotype and interleukin-15 together contribute to the pathogenesis. Although suspected, their roles and interactions still require the provision of tangible evidence in vivo. Chapter 3 details these interactions based on the proposed new mouse model. This model is characterized by the overexpression of interleukin-15 in the intestinal epithelium and lamina propria and by the expression of the HLA-DQ8 haplotype. This work demonstrates that the exposure of this mouse to gluten is accompanied by villous atrophy and the complete serological, cellular and transcriptional signature of the disease. We also demonstrate that simultaneous overexpression of interleukin-15 in both mucosal compartments is a prerequisite for the development of atrophy. This study also highlights the need for CD4+ T cells and interferon-gamma in the activation of intraepithelial lymphocytes and the development of villous atrophy. Finally, this research establishes the central role played by the HLA-DQ8 haplotype and the enzyme tissue transglutaminase II in the occurrence of these lesions. In general, the results from this model presented in Chapter 3 reflects the complexity of the interactions between gluten, genetics and IL-15 in the development of coeliac disease. Finally, chapter 4 concludes this work and chapter 5 discusses future directions for this powerful preclinical model that will undoubtedly contribute to a better understanding of coeliac disease and will allow the identification of new potential therapeutic targets.