Vincenza Lombardo

Evidence for the presence of non-celiac gluten sensitivity in patients with functional gastrointestinal symptoms: Results from a multicenter randomized double-blind placebo-controlled gluten challenge

Non-celiac gluten sensitivity (NCGS) is characterized by the onset of symptoms after eating gluten-containing food. We aimed to single out NCGS subjects among subjects with functional gastrointestinal symptoms. Patients were enrolled in a multicenter double-blind placebo-controlled trial with crossover. Symptoms and quality of life were evaluated by means of 10-cm VAS and SF36. Iron parameters, transaminases and C reactive protein (CRP) were evaluated. After a three-week-long gluten-free diet (GFD), responsive patients were randomly assigned to gluten intake (5.6 g/day) or placebo for seven days, followed by crossover. The primary endpoint was the worsening of symptoms (VAS increase ≥3 cm) during gluten ingestion compared to placebo. One hundred and forty patients were enrolled and 134 (17 males, mean age 39.1 ± 11.7 years, BMI 22.4 ± 3.8) completed the first period. A total of 101 subjects (10 males, mean age 39.3 ± 11.0 years, BMI 22.3 ± 4.0) reported a symptomatic improvement (VAS score 2.3 ± 1.2 vs. 6.5 ± 2.2 before and after GFD, p = 0.001). 98 patients underwent the gluten challenge and 28 (all females, mean age 38.9 ± 12.7 years, BMI 22.0 ± 2.9) reported a symptomatic relapse and deterioration of quality of life. No parameters were found to be statistically associated with positivity to the challenge. However, 14 patients responded to the placebo ingestion. Taking into account this finding, about 14% of patients responding to gluten withdrawal showed a symptomatic relapse during the gluten challenge. This group is suspected to have NCGS.

Evaluation of a Modified Italian European Prospective Investigation into Cancer and Nutrition Food Frequency Questionnaire for Individuals with Celiac Disease

BACKGROUNDnTo date, it is unclear whether individuals with celiac disease following a gluten-free (GF) diet for several years have adequate intake of all recommended nutrients. Lack of a food frequency questionnaire (FFQ) for individuals with celiac disease could be partly responsible for this still-debated issue.nnnOBJECTIVEnThe aim of the study is to evaluate the performance of a modified European Prospective Investigation into Cancer and Nutrition (EPIC) FFQ in estimating nutrient and food intake in a celiac population.nnnDESIGNnIn a cross-sectional study, the dietary habits of individuals with celiac disease were reported using a modified Italian EPIC FFQ and were compared to a 7-day weighed food record as a reference method.nnnPARTICIPANTS/SETTINGnA total of 200 individuals with histologically confirmed celiac disease were enrolled in the study between October 2012 and August 2014 at the Center for Prevention and Diagnosis of Celiac Disease (Milan, Italy).nnnMAIN OUTCOME MEASURESnNutrient and food category intake were calculated by 7-day weighed food record using an Italian food database integrated with the nutrient composition of 60 GF foods and the modified EPIC FFQ, in which 24 foods were substituted with GF foods comparable for energy and carbohydrate content.nnnSTATISTICAL ANALYSES PERFORMEDnAn evaluation of the modified FFQ compared to 7-day weighed food record in assessing the reported intake of nutrient and food groups was conducted using Spearmans correlation coefficients and weighted κ.nnnRESULTSnOne hundred individuals completed the study. The Spearmans correlation coefficients of FFQ and 7-day weighed food record ranged from .13 to .73 for nutrients and from .23 to .75 for food groups. A moderate agreement, which was defined as a weighted κ value of .40 to .60, was obtained for 30% of the analyzed nutrients, and 40% of the nutrients showed values between .30 and .40. The weighted κ exceeded .40 for 60% of the 15 analyzed food groups.nnnCONCLUSIONSnThe modified EPIC FFQ demonstrated moderate congruence with a weighed food record in ranking individuals by dietary intakes, particularly food groups.

Are Treated Celiac Patients at Risk for Mycotoxins? An Italian Case-Study

Urinary biomarkers of mycotoxin exposure were evaluated in a group of celiac patients (n = 55) and in a control group of healthy subjects (n = 50) following their habitual diet. Deoxynivalenol (DON), zearalenone (ZEN), and fumonisin B1 (FB1) were monitored in 105 urinary samples collected from the two groups. Dietary habits were also recorded through compilation of a seven-day weighed dietary diary. Biomarkers of mycotoxin exposure were detected in 21 celiac patients and in 15 control subjects, corresponding to about 34% of total participants. In particular, ZEN was the most detected mycotoxin among all the studied subjects with a total of 19 positive cases. Results did not show a statistically significant difference in mycotoxin exposure between the two groups, and the presence of specific mycotoxins was not related to the intake of any particular food category. Our findings suggest little urgency of specific regulation for gluten free products, although the prevalence of exposure observed in free-living diets of both celiac and healthy subjects underlines the need of a constant surveillance on mycotoxins occurrence at large.

Gliadin effect on the oxidative balance and DNA damage: An in-vitro, ex-vivo study

BACKGROUNDnGliadins are involved in gluten-related disorders and are responsible for the alteration of the cellular redox balance. It is not clear if the gliadin-related oxidative stress can induce DNA damage in enterocytes.nnnAIMnTo investigate any possible genotoxicity caused by gliadin and to assess its relationship with oxidative stress in vitro and ex vivo.nnnMETHODSnCaco-2 cells were exposed for 6-12-24u202fh to increasing concentrations (250u202fμg/mL-1000u202fμg/mL) of digested gliadin. We investigated: cytotoxicity, oxidative balance (reactive oxygen species, ROS), DNA damage (comet assay and γ-H2AX detection), transglutaminase type 2 (TG2) activity and annexin V expression. H2AX and 8-OHG immunohistochemistry has been evaluated on duodenal biopsies of celiac subjects and controls.nnnRESULTSnGliadin induced a significant increase (+50%) of ROS after 12u202fh of exposition starting with a 500u202fμg/mL dose of gliadin. Comet assay and γ-H2AX demonstrated DNA damage, evident at the gliadin concentration of 500u202fμg/mL after 24u202fh. TG2 activity increased in chromatin and cytoskeleton cellular compartments at different gliadin doses (250/500/1000u202fμg/mL). The γ-H2AX and 8-OHG immunohistochemistry was altered in the duodenal biopsies of celiac patients.nnnCONCLUSIONSnGliadin induces cellular oxidative stress, DNA damage and pro-apoptotic stimulation in Caco-2 cells and in the duodenal mucosa of celiac patients.

Vitamin D3 Versus Gliadin: A Battle to the Last Tight Junction

Celiac disease (CD), the most common chronic autoimmune enteropathy present in Western populations, is triggered by gluten ingestion in genetically susceptible individuals carrying the HLA DQ2 and/or DQ8 loci [1]. Gluten, a high molecular weight protein present in the endosperm of grass-related grains, including wheat, barley, and rye, is stored within seeds in order to ensure a stable nutrient supply supporting the germination and development of young plants. Gluten-containing cereals, the most important crop in the world, are used to make food products such as pasta, bread, other baked and pastry products. The viscoelastic and stabilizing properties of gluten have fostered its use as an additive in the baking industry; furthermore, it gives food greater palatability due to the creation of disulfide bonds that in combination with atmospheric oxygen and nitrogen alter the properties of the dough as a function of the sulfhydryl and disulfide content [1]. Gluten is a composite of two classes of protein, glutenins and prolamins (gliadin, secalin, and hordein), which can be further fractionated to produce peptides. Pepsin– trypsin-resistant gliadin (PT-G) is an undigested gliadin fragment that substantially contributes to the pathogenesis of CD by altering intercellular tight junctions (TJs) [2]. In CD, a genetic predisposition and exposure to environmental triggers cooperate in the loss of the intestinal barrier function. Gluten peptides and gliadins permeabilize the gut, which provokes an immunomodulatory cytotoxic effect and opens TJs. Subsequently, the deregulated traffic of macromolecules, due to the ‘‘leaky gut,’’ severely damages the intestine, thus fueling the chronic inflammatory process. The TJ, a dynamic protein complex, is the primary determinant of the paracellular flux of fluid and solutes in a healthy epithelium. The integrity of the barrier is important for the separation of the two compartments: the luminal and the submucosal sides [3]. TJs govern the permeability of the intestinal mucosal barrier; even minimal alterations in TJ function allow the passage of potentially toxic macromolecules through the intercellular spaces in the subcellular matrix, a condition called ‘‘leaky gut syndrome’’ that is common in many inflammatory diseases of the small bowel [4–6]. The three-dimensional structure of many cells is governed by a network of polymeric proteins including actin filaments termed the cytoskeleton, the organization of which is implicated in the pathogenesis of CD. Gliadin causes the disarrangement and disappearance of epithelial cellular organization in CD patients [7]. TJs are demonstrably associated with the actin filaments in epithelial cells assembling in a fusion structure called ‘‘kissing points.’’ The inflammation and autoimmunity that occurs secondary to the ingestion of gluten in predisposed subjects disconnects innate and adaptive immunity. Zonulin is the only known modulator of intercellular TJs, regulating intestinal permeability [8]. During the acute phase of CD, the zonulin pathway is upregulated. For all the aforementioned reasons, investigation is extremely active into drugs & Luca Elli luca.elli@policlinico.mi.it