Rapid Anti-tTG-IgA Screening Test for Early Diagnosis of Celiac Disease in Pediatric Populations.

A large number of patients with celiac disease (CD) remain undiagnosed because they do not fulfill the criteria for entry into the conventional diagnostic workflow. This study evaluated the clinical utility of anti-tissue transglutaminase IgA antibody lateral flow immunoassays (anti-tTG-IgA LFIA) in the undiagnosed-CD-based pediatric population and the impact of a gluten-free diet (GFD) on screening-detected CD. A total of 576 volunteers were tested for anti-tTG-IgA. Gluten consumption habits, CD related symptoms, and risk factors for CD development were evaluated. Volunteers testing positive for anti-tTG-IgA were referred to the conventional CD diagnostic workflow, and the impact of the GFD on health-related quality of life (HR-QoL) was measured. Among them, 13 had a positive anti-tTG-IgA LFIA test result: 11 had confirmed CD (1.91%), one refused confirmatory tests, and another is undergoing diagnosis. Regarding the CD prevalence, no significant differences were observed among risk (1.89%) and symptomatic (2.65%) groups and the entire tested population (1.55%). Rapid anti-tTG-IgA LFIAs could be of clinical utility in primary care for the early identification of children with CD unidentified by the conventional diagnostic workflow. It could potentially reduce the costs of undiagnosed CD, avoiding unnecessary referrals to gastroenterologists, reducing diagnosis delays and long-term problems, and improving patients' HR-QoL.

Evaluation of the Usefulness of an Automatable Immunoassay for Monitoring Celiac Disease by Quantification of Immunogenic Gluten Peptides in Urine.

A gluten-free diet (GFD) is currently the only treatment available for patients with celiac disease (CD). However, adherence to a GFD can be challenging because gluten is present in many foods. A lifelong follow-up of patients with CD must be performed to promote adherence to a GFD and to identify the appearance of symptoms and the associated diseases. Therefore, the development of tools to analyze gluten exposure in these patients is important. This study proposes the development of the first automatable ELISA to monitor adherence to a GFD through the quantification of urine gluten immunogenic peptides (u-GIP). Seven healthy volunteers without suspicion of CD and 23 patients with CD were monitored as part of this study to optimize, validate, and apply this assay. Non-interference was found in the urine matrix, and the recovery percentage for spiked samples was 81-101%. The u-GIP was stable for up to 16 days when the samples were stored at different temperatures. Overall, 100% of the patients had detectable u-GIP at diagnosis (range of 0.39-2.14 ng GIP/mL), which reduced to 27% after 12 months on a GFD. Therefore, this highly sensitive immunoassay would allow the analysis of u-GIP from a large battery of samples in clinical laboratories of specialized healthcare centers.

Clinical utility of urinary gluten immunogenic peptides in the follow-up of patients with coeliac disease.

BACKGROUND: Gluten-free diet (GFD) is the only treatment for patients with coeliac disease (CD) and its compliance should be monitored to avoid cumulative damage. AIMS: To analyse gluten exposures of coeliac patients on GFD for at least 24 months using different monitoring tools and its impact on duodenal histology at 12-month follow-up and evaluate the interval of determination of urinary gluten immunogenic peptides (u-GIP) for the monitoring of GFD adherence. METHODS: Ninety-four patients with CD on a GFD for at least 24 months were prospectively included. Symptoms, serology, CDAT questionnaire, and u-GIP (three samples/visit) were analysed at inclusion, 3, 6, and 12 months. Duodenal biopsy was performed at inclusion and 12 months. RESULTS: At inclusion, 25.8% presented duodenal mucosal damage; at 12 months, this percentage reduced by half. This histological improvement was indicated by a reduction in u-GIP but did not correlate with the remaining tools. The determination of u-GIP detected a higher number of transgressions than serology, regardless of histological evolution type. The presence of >4 u-GIP-positive samples out of 12 collected during 12 months predicted histological lesion with a specificity of 93%. Most patients (94%) with negative u-GIP in ≥2 follow-up visits showed the absence of histological lesions (p < 0.05). CONCLUSION: This study suggests that the frequency of recurrent gluten exposures, according to serial determination of u-GIP, could be related to the persistence of villous atrophy and that a more regular follow-up every 6 months, instead of annually, provides more useful data about the adequate adherence to GFD and mucosal healing.

A Highly Sensitive Method for the Detection of Hydrolyzed Gluten in Beer Samples Using LFIA.

Most gluten analysis methods have been developed to detect intact gluten, but they have shown limitations in certain foods and beverages in which gluten proteins are hydrolyzed. Methods based on G12/A1 moAbs detect the sequences of gluten immunogenic peptides (GIP), which are the main contributors to the immune response of celiac disease (CD). Immunogenic sequences with tandem epitopes for G12/A1 have been found in beers with <20 mg/kg gluten, which could be consumed by CD patients according to the Codex Alimentarius. Therefore, an accurate method for the estimation of the immunogenicity of a beer is to use two moAbs that can recognize celiac T cell epitopes comprising most of the immunogenic response. Here, a specific and sensitive method based on G12/A1 LFIA was developed to detect GIP in beers labeled gluten-free or with low gluten content, with an LOD of 0.5 mg/kg. A total of 107 beers were analyzed, of those 6.5% showed levels higher than 20 mg/kg gluten and 29% showed levels above the LOD. In addition, G12/A1 LFIA detected gluten in 15 more beer samples than competitive ELISA with another antibody. Despite their labeling, these beers contained GIP which may cause symptoms and/or intestinal damage in CD patients.

Food Safety and Cross-Contamination of Gluten-Free Products: A Narrative Review.

A gluten-free diet (GFD) is currently the only effective treatment for celiac disease (CD); an individual's daily intake of gluten should not exceed 10 mg. However, it is difficult to maintain a strict oral diet for life and at least one-third of patients with CD are exposed to gluten, despite their best efforts at dietary modifications. It has been demonstrated that both natural and certified gluten-free foods can be heavily contaminated with gluten well above the commonly accepted threshold of 20 mg/kg. Moreover, meals from food services such as restaurants, workplaces, and schools remain a significant risk for inadvertent gluten exposure. Other possible sources of gluten are non-certified oat products, numerous composite foods, medications, and cosmetics that unexpectedly contain "hidden" vital gluten, a proteinaceous by-product of wheat starch production. A number of immunochemical assays are commercially available worldwide to detect gluten. Each method has specific features, such as format, sample extraction buffers, extraction time and temperature, characteristics of the antibodies, recognition epitope, and the reference material used for calibration. Due to these differences and a lack of official reference material, the results of gluten quantitation may deviate systematically. In conclusion, incorrect gluten quantitation, improper product labeling, and poor consumer awareness, which results in the inadvertent intake of relatively high amounts of gluten, can be factors that compromise the health of patients with CD.

New Insights into Non-Dietary Treatment in Celiac Disease: Emerging Therapeutic Options.

To date, the only treatment for celiac disease (CD) consists of a strict lifelong gluten-free diet (GFD), which has numerous limitations in patients with CD. For this reason, dietary transgressions are frequent, implying intestinal damage and possible long-term complications. There is an unquestionable need for non-dietary alternatives to avoid damage by involuntary contamination or voluntary dietary transgressions. In recent years, different therapies and treatments for CD have been developed and studied based on the degradation of gluten in the intestinal lumen, regulation of the immune response, modulation of intestinal permeability, and induction of immunological tolerance. In this review, therapeutic lines for CD are evaluated with special emphasis on phase III and II clinical trials, some of which have promising results.

Challenges of Monitoring the Gluten-Free Diet Adherence in the Management and Follow-Up of Patients with Celiac Disease.

Celiac disease (CD) is a chronic gluten-responsive immune mediated enteropathy and is treated with a gluten-free diet (GFD). However, a strict diet for life is not easy due to the ubiquitous nature of gluten. This review aims at examining available evidence on the degree of adherence to a GFD, the methods to assess it, and the barriers to its implementation. The methods for monitoring the adherence to a GFD are comprised of a dietary questionnaire, celiac serology, or clinical symptoms; however, none of these methods generate either a direct or an accurate measure of dietary adherence. A promising advancement is the development of tests that measure gluten immunogenic peptides in stools and urine. Causes of adherence/non-adherence to a GFD are numerous and multifactorial. Inadvertent dietary non-adherence is more frequent than intentional non-adherence. Cross-contamination of gluten-free products with gluten is a major cause of inadvertent non-adherence, while the limited availability, high costs, and poor quality of certified gluten-free products are responsible for intentionally breaking a GFD. Therefore, several studies in the last decade have indicated that many patients with CD who follow a GFD still have difficulty controlling their diet and, therefore, regularly consume enough gluten to trigger symptoms and damage the small intestine.

Rapid, Effective, and Versatile Extraction of Gluten in Food with Application on Different Immunological Methods.

One of the main concerns in gluten analysis is to achieve efficient extraction of gluten proteins. Conventional ethanol-based extraction solutions are inefficient and, because of this, it is necessary to use reducing agents or acids for proper solubilization. The extraction recommended by CODEX Standard 118-1979 (revised 2008) utilizes Cocktail solution (patent WO 02/092633 A1). However, it is harmful with a disgusting odor and is not compatible with some immunological techniques. Here, the versatility and extraction capacity of a new Universal Gluten Extraction Solution (UGES) (patent ES 2 392 412 A1) were evaluated using different methodological conditions, food matrices, and various immunological methods. UGES includes safer compounds for both the user and the environment, and it displayed similar extraction efficiency to that of the extraction method recommended for sandwich enzyme-linked immunosorbent assay (ELISA). The extraction time was significantly reduced from 100 to 40 min, depending on the type of the sample. Furthermore, unlike the currently used solution, UGES is compatible with competitive ELISA.

Negative predictive value of the repeated absence of gluten immunogenic peptides in the urine of treated celiac patients in predicting mucosal healing: new proposals for follow-up in celiac disease.

BACKGROUND: The treatment of celiac disease (CD) is a lifelong gluten-free diet (GFD). The current methods for monitoring GFD conformance, such as a dietary questionnaire or serology tests, may be inaccurate in detecting dietary transgressions, and duodenal biopsies are invasive, expensive, and not a routine monitoring technique. OBJECTIVES: Our aim was to determine the clinical usefulness of urine gluten immunogenic peptides (GIP) as a biomarker monitoring GFD adherence in celiac patients and to evaluate the concordance of the results with the degree of mucosal damage. METHODS: A prospective observational study was conducted involving 22 de novo CD patients, 77 celiac patients consuming a GFD, and 13 nonceliac subjects. On 3 d of the week, urine samples were collected and the GIP concentrations were tested. Simultaneously, anti-tissue transglutaminase antibodies, questionnaire results, clinical manifestations, and histological findings were analyzed. RESULTS: Approximately 24% (18 of 76) of the celiac patients consuming a GFD exhibited Marsh II-III mucosal damage. Among this population, 94% (17 of 18) had detectable urine GIP; however, between 60% and 80% were asymptomatic and exhibited negative serology and appropriate GFD adherence based on the questionnaire. In contrast, 97% (31 of 32) of the celiac patients without duodenal damage had no detectable GIP. These results demonstrated the high sensitivity (94%) and negative predictive value (97%) of GIP measurements in relation to duodenal biopsy findings. In the de novo CD-diagnosed cohort, 82% (18 of 22) of patients had measurable amounts of GIP in the urine. CONCLUSIONS: Determining GIP concentrations in several urine samples may be an especially convenient approach to assess recent gluten exposure in celiac patients and appears to accurately predict the absence of histological lesions. The introduction of GIP testing as an assessment technique for GFD adherence may help in ascertaining dietary compliance and to target the most suitable intervention during follow-up.

Celiac Immunogenic Potential of α-Gliadin Epitope Variants from Triticum and Aegilops Species.

The high global demand of wheat and its subsequent consumption arise from the physicochemical properties of bread dough and its contribution to the protein intake in the human diet. Gluten is the main structural complex of wheat proteins and subjects affected by celiac disease (CD) cannot tolerate gluten protein. Within gluten proteins, α-gliadins constitute the most immunogenic fraction since they contain the main T-cell stimulating epitopes (DQ2.5-glia-α1, DQ2.5-glia-α2, and DQ2.5-glia-α3). In this work, the celiac immunotoxic potential of α-gliadins was studied within Triticeae: diploid, tetraploid, and hexaploid species. The abundance and immunostimulatory capacity of CD canonical epitopes and variants (with one or two mismatches) in all α-gliadin sequences were determined. The results showed that the canonical epitopes DQ2.5-glia-α1 and DQ2.5-glia-α3 were more frequent than DQ2.5-glia-α2. A higher abundance of canonical DQ2.5-glia-α1 epitope was found to be associated with genomes of the BBAADD, AA, and DD types; however, the abundance of DQ2.5-glia-α3 epitope variants was very high in BBAADD and BBAA wheat despite their low abundance in the canonical epitope. The most abundant substitution was that of proline to serine, which was disposed mainly on the three canonical DQ2.5 domains on position 8. Interestingly, our results demonstrated that the natural introduction of Q to H at any position eliminates the toxicity of the three T-cell epitopes in the α-gliadins. The results provided a rational approach for the introduction of natural amino acid substitutions to eliminate the toxicity of three T-cell epitopes, while maintaining the technological properties of commercial wheats.