Guidelines for best practices in monitoring established coeliac disease in adult patients.

Coeliac disease (CeD) is an immunological disease triggered by the consumption of gluten contained in food in individuals with a genetic predisposition. Diagnosis is based on the presence of small bowel mucosal atrophy and circulating autoantibodies (anti-type 2 transglutaminase antibodies). After diagnosis, patients follow a strict, life-long gluten-free diet. Although the criteria for diagnosis of this disease are well defined, the monitoring phase has been studied less and there is a lack of specific guidelines for this phase. To develop a set of clinical guidelines for CeD monitoring, we followed the Grading of Recommendations Assessment, Development and Evaluation methodology. Statements and recommendations with the level of evidence were developed and approved by the working group, which comprised gastroenterologists, pathologists, dieticians and biostatisticians. The proposed guidelines, endorsed by the North American and European coeliac disease scientific societies, make recommendations for best practices in monitoring patients with CeD based on the available evidence. The evidence level is low for many topics, suggesting that further research in specific aspects of CeD would be valuable. In conclusion, the present guidelines support clinicians in improving CeD treatment and follow-up and highlight novel issues that should be considered in future studies.

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 a Single Determination of Gluten Immunogenic Peptides in Urine from Unaware Celiac Patients to Monitor Gluten-Free Diet Adherence.

INTRODUCTION AND AIM: Usually, adherence to the gluten-free diet (GFD) in celiac patients is indirectly assessed through serological analysis, questionnaires, or invasive methods such as intestinal biopsy. The detection of gluten immunogenic peptides in urine (urinary gluten immunogenic peptides-uGIP) is a novel technique that directly evaluates the ingestion of gluten. The aim of this study was to evaluate the clinical efficacy of uGIP in the follow-up of celiac disease (CD). METHODS: From April 2019 to February 2020, CD patients reporting complete adherence to the GFD were prospectively enrolled but were unaware of the reason for the tests. Urinary GIP, the celiac dietary adherence test (CDAT), symptomatic visual analog scales (VAS), and tissue transglutaminase antibodies (tTGA) titres were evaluated. Duodenal histology and capsule endoscopy (CE) were performed when indicated. RESULTS: A total of 280 patients were enrolled. Thirty-two (11.4%) had a positive uGIP test (uGIP+). uGIP+ patients did not show significant differences in demographic parameters, CDAT, or VAS scores. The tTGA+ titre was not related to the positivity of uGIP (14.4% vs. 10.9% in patients with tTGA+ and tTGA-). Regarding histology, 66.7% of the GIP+ patients had atrophy compared to 32.7% of the GIP patients (p-value 0.01). However, the presence of atrophy did not correlate with tTGA. Mucosal atrophy was detected in 29 (47.5%) out of 61 patients by CE. With this method, no noticeable dependence on uGIP results (24 GIP- vs. 5 GIP+) was observed. CONCLUSIONS: The single uGIP test was positive in 11% of CD cases referring a correct GFD adherence. Furthermore, uGIP results significantly correlated with the duodenal biopsy, formerly considered the gold standard for assessing CD activity.

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.

Individual variability in patterns and dynamics of fecal gluten immunogenic peptides excretion after low gluten intake.

PURPOSE: Determination of Gluten Immunogenic Peptides (GIP) in feces is a direct tool for gluten exposure detection. The sensitivity of GIP detection methods for cases of unintentional low gluten intakes is unknown. We studied the interindividual variability in the kinetic of excretion under homogeneously controlled dietary conditions, and the sensitivity of fecal GIP tests after low amounts of punctual gluten ingestions. METHODS: Participants (n = 20) followed the same gluten-free menu for 12 days in which two separated doses of gluten (50 mg and 2 g) were ingested and all the depositions were collected. GIP from stool samples were analyzed by ELISA and lateral flow immunoassay (LFIA) tests. RESULTS: Most participants had detectable GIP after 50 mg and 2 g gluten ingestions using ELISA test (72.2% and 95%, respectively), whereas the LFIA test showed less sensitivity (22.2% and 80%, respectively). GIP were detected at higher either frequency or concentration in the range of 12-36 h after 50 mg intake, and 12-84 h after 2 g consumption. Considering this period, diagnostic sensitivity of GIP detection after a single 50 mg ingestion may be significatively increased analyzing three stool samples per individual. High variability among participants was found in the time and amount of GIP excretion; however, some individuals showed common patterns for both gluten intakes. CONCLUSION: Sporadic gluten exposure detection may require several fecal samples to achieve level of sensitivity above 90%. Interindividual variability in the dynamic of GIP excretion may suggest patterns of gluten metabolism.

Determination of gluten immunogenic peptides for the management of the treatment adherence of celiac disease: A systematic review.

BACKGROUND: Gluten is a complex mixture of proteins with immunogenic peptide sequences triggering the autoimmune activity in patients with celiac disease (CeD). Gluten immunogenic peptides (GIP) are resistant to gastrointestinal digestion and are then excreted via the stool and urine. Most common detection methods applied in the follow-up visits for CeD patients such as serology tests, dietetic interviews, questionnaires, and duodenal biopsy have been proved to be inefficient, invasive, or inaccurate for evaluating gluten-free diet (GFD) compliance. Determination of excreted GIP in stool and urine has been developed as a non-invasive, direct, and specific test for GFD monitoring. AIM: To summarize published literature about the clinical utility of GIP determination in comparison to the tools employed for GFD monitoring. METHODS: PubMed and Web of Science searches were performed using the keywords "gluten immunogenic peptides" or "gluten immunogenic peptide" and a combination of the previous terms with "feces", "stools", "urine", "celiac disease", "gluten-free diet", and "adherence" to identify relevant clinical studies published in English and Spanish between 2012 to January 2021. Reference lists from the articles were reviewed to identify additional pertinent articles. Published articles and abstracts reporting the clinical use of GIP determination in stool and/or urine for the follow-up of patients with CeD in comparison with other tools in use were included. Case reports, commentaries, reviews, conference papers, letters, and publications that did not focus on the aims of this review were excluded. RESULTS: Total of 15 publications were found that involved the use of GIP determination in stool and/or urine to monitor the adherence to the GFD in comparison to other tools. Studies included both children and adults diagnosed with CeD and healthy volunteers. Overall, these preliminary studies indicated that this novel technique was highly sensitive for the detection of GFD transgressions and therefore could facilitate the follow-up of patients with CeD. Tools identified in this work included the CeD-specific serology, dietetic questionnaires, symptomatology, and the duodenal biopsy. Review of the literature revealed that the rates of GFD adherence may vary between 30%-93% using either stool or urine GIP determination, 49%-96% by the serology, 59%-94% using the dietetic questionnaires, 56%-95% by the reported symptoms and 44%-76% with the duodenal biopsy. In addition, the association between the different methods and histological abnormalities (Marsh II-III) was found to be 33%-100% for GIP determination (stool and urine), 25%-39% for CeD-specific serology, 3%-50% for dietetic questionnaires, and 22%-28% for the symptomatology. CONCLUSION: Excreted GIP detection is the precise approach for determining voluntary or involuntary gluten consumption in CeD patients preventing future complications arising from gluten exposure.

Dynamics and Considerations in the Determination of the Excretion of Gluten Immunogenic Peptides in Urine: Individual Variability at Low Gluten Intake.

BACKGROUND: A lifelong strict gluten-free diet is the only available treatment for celiac disease, but total exclusion of gluten is difficult to achieve. The aim of this study was to determine the range of time and the amount of gluten immunogenic peptides (GIP) excreted in urine after specific gluten ingestions. METHODS: 20 healthy participants followed the same diet for 12 days in which 50 mg and 2 g of gluten were ingested and all the urinations were collected. GIP were analyzed by lateral flow immunoassay (LFIA) tests and quantified using an LFIA reader. RESULTS: GIP were detected in 15% and 95% of participants after 50 mg and 2 g gluten intakes, respectively. The higher frequency and concentration of GIP was found between 6 and 9 h after both gluten ingestions. The ranges of detection were 3-12 h (50 mg) and 0-15 h (2 g). CONCLUSIONS: An increase in the frequency of urine tests may be a suitable approach to avoid false negative results. The use of the LFIA test in three urine samples collected at different times may show a sensitivity of 19.6% for a gluten ingestion like 50 mg, increasing to 93% after 2 g consumption.

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.

Exposure sources, amounts and time course of gluten ingestion and excretion in patients with coeliac disease on a gluten-free diet.

BACKGROUND: A major deficit in understanding and improving treatment in coeliac disease (CD) is the lack of empiric data on real world gluten exposure. AIMS: To estimate gluten exposure on a gluten-free diet (GFD) using immunoassays for gluten immunogenic peptides (GIP) and to examine relationships among GIP detection, symptoms and suspected gluten exposures METHODS: Adults with biopsy-confirmed CD on a GFD for 24 months were recruited from a population-based inception cohort. Participants kept a diary and collected urine samples for 10 days and stools on days 4-10. 'Doggie bags' containing » portions of foods consumed were saved during the first 7 days. Gluten in food, stool and urine was quantified using A1/G12 ELISA. RESULTS: Eighteen participants with CD (12 female; age 21-70 years) and three participants on a gluten-containing diet enrolled and completed the study. Twelve out of 18 CD participants had a median 2.1 mg gluten per exposure (range 0.2 to >80 mg). Most exposures were asymptomatic and unsuspected. There was high intra-individual variability in the interval between gluten ingestion and excretion. Participants were generally unable to identify the food. CONCLUSIONS: Gluten exposure on a GFD is common, intermittent, and usually silent. Excretion kinetics are highly variable among individuals. The amount of gluten varied widely, but was typically in the milligram range, which was 10-100 times less than consumed by those on an unrestricted diet. These findings suggest that a strict GFD is difficult to attain, and specific exposures are difficult to detect due to variable time course of excretion.

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.

A compact SPR biosensor device for the rapid and efficient monitoring of gluten-free diet directly in human urine.

Celiac disease (CD) is a chronic autoimmune disorder induced in genetically susceptible individuals by the ingestion of gluten from wheat, rye, barley, or certain varieties of oats. A careful diet follow-up is necessary to avoid health complications associated with long-term gluten intake by the celiac patients. Small peptides (GIP, gluten immunogenic peptides) derived from gluten digestion, which are excreted in the urine and feces, have emerged as promising biomarkers to monitor gluten intake. We have implemented a simple and sensitive label-free point-of-care (POC) device based on surface plasmon resonance for the direct detection of these biomarkers in urine. The assay employs specific monoclonal antibodies and has been optimized for the detection of the 33-mer α2-gliadin, known as the main immunogenic peptide of wheat gluten, and for the detection of GIP. Direct detection in undiluted urine has been accomplished by using biosensing chips containing a robust and stable biorecognition layer, obtained after carefully optimizing the biofunctionalization protocol. Excellent limits of detection have been reached (1.6-4.0 ng mL-1 using mAb G12 and A1, respectively), which ensures the detection of gluten peptides even when the gluten intake is around the maximum tolerable amount in the digestive tract (< 50 mg) for celiac individuals. No sample pretreatment, extraction, or dilution is required, and the analysis takes less than 15 min. The assays have excellent reproducibility' as demonstrated by measuring spiked urine samples containing the same target concentration using different biofunctionalized chips prepared and stored at different periods of time (i.e., CV% of 3.58% and 11.30%, for G12- and A1-based assays, respectively). The assay has been validated with real samples. These features pave the way towards an end-user easy-to-handle biosensor device for the rapid monitoring of gluten-free diet (GFD) and follow-up of the health status in celiac patients.

Prospective longitudinal study: use of faecal gluten immunogenic peptides to monitor children diagnosed with coeliac disease during transition to a gluten-free diet.

BACKGROUND: Treatment for coeliac disease is a lifelong strict gluten-free diet. Although guidelines recommend regular follow-up with dietary interviews and coeliac serology, these methods may be inaccurate. AIM: To evaluate the usefulness of faecal gluten immunogenic peptides to support the diagnosis and to determine the adherence to the gluten-free diet in coeliac children. METHODS: Multicentre prospective observational study including 64 coeliac children. Faecal gluten peptides, and tissue transglutaminase and deamidated gliadin peptide antibodies were analyzed at diagnosis, and 6, 12 and 24 months thereafter. Gluten consumption was estimated from gluten peptide levels. RESULTS: Most children (97%) had detectable gluten peptides at diagnosis. On a gluten-free diet, the rate of gluten peptides increased from 13% at 6 months to 25% at 24 months. Mean estimated gluten exposure dropped from 5543 mg/d at diagnosis to 144 mg/d at 6 months, then increased to 606 mg/d by 24 months. In contrast, deamidated gliadin peptide antibodies normalised and only 20% had elevated tissue transglutaminase antibody by 24 months. The elevation of tissue transglutaminase antibody was more prolonged in patients with detectable gluten peptides (P < 0.05). Nevertheless, absolute levels of tissue transglutaminase antibody had low sensitivity to identify patients with detectable gluten peptides (P > 0.1). Dietitian assessment was only moderately correlated with gluten peptide detection (κ = 0.5). CONCLUSIONS: Faecal gluten peptides testing may guide treatment of coeliac disease prior to diagnosis and during the assessment diet adherence. Further studies could determine if early identification of gluten exposure reduces the need for expensive/invasive investigations for non-responsive coeliac disease. ClinicalTrials.gov Number: NCT02711397.

Gluten Immunogenic Peptides as Standard for the Evaluation of Potential Harmful Prolamin Content in Food and Human Specimen.

Gluten is a complex mixture of storage proteins in cereals like wheat, barley, and rye. Prolamins are the main components of gluten. Their high content in proline and glutamine makes them water-insoluble and difficult to digest in the gastrointestinal tract. Partial digestion generates peptide sequences which trigger immune responses in celiac and gluten-sensitive patients. Gluten detection in food is challenging because of the diversity, in various food matrices, of protein proportions or modifications and the huge number of immunogenic sequences with differential potential immunoactivity. Attempts to develop standard reference materials have been unsuccessful. Recent studies have reported the detection of a limited number of dominant Gluten Immunogenic Peptides (GIP) that share similarities to epitopes presented in the α-gliadin 33-mer, which showed to be highly proteolytic resistant and is considered to be the most immunodominant peptide within gluten in celiac disease (CD). GIP were detectable and quantifiable in very different kind of difficult to analyze food, revealing the potential immunogenicity by detecting T-cell activity of celiac patients. But GIP were also found in stool and urine of celiac patients on a supposedly gluten-free diet (GFD), showing the capacity to resist and be absorbed and excreted from the body, providing the first simple and objective means to assess adherence to the GFD. Methods to specifically and sensitively detect the most active GIP in food and biological fluids are rational candidates may use similar analytical standard references for determination of the immunopathological risk of gluten exposure in gluten-related diseases.

Corrigendum: Fecal Gluten Peptides Reveal Limitations of Serological Tests and Food Questionnaires for Monitoring Gluten-Free Diet in Celiac Disease Patients.

This corrects the article DOI: 10.1038/ajg.2016.439.

Detection of gluten immunogenic peptides in the urine of patients with coeliac disease reveals transgressions in the gluten-free diet and incomplete mucosal healing.

OBJECTIVE: Gluten-free diet (GFD) is the only management for coeliac disease (CD). Available methods to assess GFD compliance are insufficiently sensitive to detect occasional dietary transgressions that may cause gut mucosal damage. We aimed to develop a method to determine gluten intake and monitor GFD compliance in patients with CD and to evaluate its correlation with mucosal damage. DESIGN: Urine samples of 76 healthy subjects and 58 patients with CD subjected to different gluten dietary conditions were collected. A lateral flow test (LFT) with the highly sensitive and specific G12 monoclonal antibody for the most dominant gluten immunogenic peptides (GIP) and a LFT reader were used to quantify GIP in solid-phase extracted urines. RESULTS: GIP were detectable in concentrated urines from healthy individuals previously subjected to GFD as early as 4-6 h after single gluten intake, and remained detectable for 1-2 days. The urine assay revealed infringement of the GFD in about 50% of the patients. Analysis of duodenal biopsies revealed that most of patients with CD (89%) with no villous atrophy had no detectable GIP in urine, while all patients with quantifiable GIP in urine showed incomplete intestinal mucosa recovery. CONCLUSION: GIP are detected in urine after gluten consumption, enabling a new and non-invasive method to monitor GFD compliance and transgressions. The method was sensitive, specific and simple enough to be convenient for clinical monitoring of patients with CD as well as for basic and clinical research applications including drug development. TRIAL REGISTRATION NUMBER: NCT02344758.

Fecal Gluten Peptides Reveal Limitations of Serological Tests and Food Questionnaires for Monitoring Gluten-Free Diet in Celiac Disease Patients.

OBJECTIVES: Treatment for celiac disease (CD) is a lifelong strict gluten-free diet (GFD). Patients should be followed-up with dietary interviews and serology as CD markers to ensure adherence to the diet. However, none of these methods offer an accurate measure of dietary compliance. Our aim was to evaluate the measurement of gluten immunogenic peptides (GIP) in stools as a marker of GFD adherence in CD patients and compare it with traditional methods of GFD monitoring. METHODS: We performed a prospective, nonrandomized, multicenter study including 188 CD patients on GFD and 84 healthy controls. Subjects were given a dietary questionnaire and fecal GIP quantified by enzyme-linked immunosorbent assay (ELISA). Serological anti-tissue transglutaminase (anti-tTG) IgA and anti-deamidated gliadin peptide (anti-DGP) IgA antibodies were measured simultaneously. RESULTS: Of the 188 celiac patients, 56 (29.8%) had detectable GIP levels in stools. There was significant association between age and GIP in stools that revealed increasing dietary transgressions with advancing age (39.2% in subjects ≥13 years old) and with gender in certain age groups (60% in men ≥13 years old). No association was found between fecal GIP and dietary questionnaire or anti-tTG antibodies. However, association was detected between GIP and anti-DGP antibodies, although 46 of the 53 GIP stool-positive patients were negative for anti-DGP. CONCLUSIONS: Detection of gluten peptides in stools reveals limitations of traditional methods for monitoring GFD in celiac patients. The GIP ELISA enables direct and quantitative assessment of gluten exposure early after ingestion and could aid in the diagnosis and clinical management of nonresponsive CD and refractory CD. Trial registration number NCT02711397.

Selective capture of most celiac immunogenic peptides from hydrolyzed gluten proteins.

The available immunomethods for gluten quantitation could underestimate or overestimate the net immunoactivity of foods and beverages if the chosen analytical antibody is not specific to the relevant gluten immunogenic peptides (GIP). Accurate detection of the most active GIP is desirable to assess the potential celiac toxicity of food. We evaluated the capacity of the G12 monoclonal antibody for selectively depleting GIP in samples from two different gluteomes. Samples of hydrolyzed gliadin from wheat and a barley beer were used. The input (starting peptide digest of prolamins), the flow-through (unbound peptides), and the output (captured peptides) were analyzed by G12 and R5 competitive ELISA as well as by stimulation assays of T-cells from celiac patients. Most of the GIP were retained by the G12-agarose and represented the largest part of the immunogenicity of the gluten peptidome. G12 immunodepletion experiments with hydrolyzed gluten showed that this antibody reacted with those with the highest immunoactivity for celiac patients.

Identification and molecular characterization of oat peptides implicated on coeliac immune response.

BACKGROUND: Oats provide important nutritional and pharmacological properties, although their safety in coeliac patients remains controversial. Previous studies have confirmed that the reactivity of the anti-33-mer monoclonal antibody with different oat varieties is proportional to the immune responses in terms of T-cell proliferation. Although the impact of these varieties on the adaptive response has been studied, the role of the dendritic cells (DC) is still poorly understood. The aim of this study is to characterize different oat fractions and to study their effect on DC from coeliac patients. METHODS AND RESULTS: Protein fractions were isolated from oat grains and analyzed by SDS-PAGE. Several proteins were characterized in the prolamin fraction using immunological and proteomic tools, and by Nano-LC-MS/MS. These proteins, analogous to α- and γ-gliadin-like, showed reactive sequences to anti-33-mer antibody suggesting their immunogenic potential. That was further confirmed as some of the newly identified oat peptides had a differential stimulatory capacity on circulating DC from coeliac patients compared with healthy controls. CONCLUSIONS: This is the first time, to our knowledge, where newly identified oat peptides have been shown to elicit a differential stimulatory capacity on circulating DC obtained from coeliac patients, potentially identifying immunogenic properties of these oat peptides.