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The Diverse Potential of Gluten from Different Durum Wheat Varieties in Triggering Celiac Disease: A Multilevel In Vitro, Ex Vivo and In Vivo Approach.

ABSTRACT: The reasons behind the increasing prevalence of celiac disease (CD) worldwide are still not fully understood. This study adopted a multilevel approach (in vitro, ex vivo, in vivo) to assess the potential of gluten from different wheat varieties in triggering CD. Peptides triggering CD were identified and quantified in mixtures generated from simulated gastrointestinal digestion of wheat varieties (n = 82). Multivariate statistics enabled the discrimination of varieties generating low impact on CD (e.g., Saragolla) and high impact (e.g., Cappelli). Enrolled subjects (n = 46) were: 19 healthy subjects included in the control group; 27 celiac patients enrolled for the in vivo phase. Celiacs were divided into a gluten-free diet group (CD-GFD), and a GFD with Saragolla-based pasta group (CD-Sar). The diet was followed for 3 months. Data were compared between CD-Sar and CD-GFD before and after the experimental diet, demonstrating a limited ability of Saragolla to trigger immunity, although not comparable to a GFD. Ex vivo studies showed that Saragolla and Cappelli activated immune responses, although with great variability among patients. The diverse potential of durum wheat varieties in triggering CD immune response was demonstrated. Saragolla is not indicated for celiacs, yet it has a limited potential to trigger adverse immune response.

SUBMITTER: Gaiani F

PROVIDER: S-EPMC7699868 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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The Diverse Potential of Gluten from Different Durum Wheat Varieties in Triggering Celiac Disease: A Multilevel In Vitro, Ex Vivo and In Vivo Approach.

Gaiani Federica F Graziano Sara S Boukid Fatma F Prandi Barbara B Bottarelli Lorena L Barilli Amelia A Dossena Arnaldo A Marmiroli Nelson N Gullì Mariolina M de'Angelis Gian Luigi GL Sforza Stefano S

Nutrients 20201120 11

The reasons behind the increasing prevalence of celiac disease (CD) worldwide are still not fully understood. This study adopted a multilevel approach (in vitro, ex vivo, in vivo) to assess the potential of gluten from different wheat varieties in triggering CD. Peptides triggering CD were identified and quantified in mixtures generated from simulated gastrointestinal digestion of wheat varieties (<i>n</i> = 82). Multivariate statistics enabled the discrimination of varieties generating low impa ...[more]

PMID: 33233787

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Project description:BACKGROUND: Wheat gluten is important for the industrial quality of bread wheat (Triticum aestivum L.) and durum wheat (T. turgidum L.). Gluten proteins are also the source of immunogenic peptides that can trigger a T cell reaction in celiac disease (CD) patients, leading to inflammatory responses in the small intestine. Various peptides with three major T cell epitopes involved in CD are derived from alpha-gliadin fraction of gluten. Alpha-gliadins are encoded by a large multigene family and amino acid variation in the CD epitopes is known to influence the immunogenicity of individual gene family members. Current commercial methods of gluten detection are unable to distinguish between immunogenic and non-immunogenic CD epitope variants and thus to accurately quantify the overall CD epitope load of a given wheat variety. Such quantification is indispensable for correct selection of wheat varieties with low potential to cause CD. RESULTS: A 454 RNA-amplicon sequencing method was developed for alpha-gliadin transcripts encompassing the three major CD epitopes and their variants. The method was used to screen developing grains on plants of 61 different durum wheat cultivars and accessions. A dedicated sequence analysis pipeline returned a total of 304 unique alpha-gliadin transcripts, corresponding to a total of 171 'unique deduced protein fragments' of alpha-gliadins. The numbers of these fragments obtained in each plant were used to calculate quantitative and quantitative differences between the CD epitopes expressed in the endosperm of these wheat plants. A few plants showed a lower fraction of CD epitope-encoding alpha-gliadin transcripts, but none were free of CD epitopes. CONCLUSIONS: The dedicated 454 RNA-amplicon sequencing method enables 1) the grouping of wheat plants according to the genetic variation in alpha-gliadin transcripts, and 2) the screening for plants which are potentially less CD-immunogenic. The resulting alpha-gliadin sequence database will be useful as a reference in proteomics analysis regarding the immunogenic potential of mature wheat grains.

Project description:Patients with non-celiac gluten sensitivity (NCGS) do not have celiac disease, but their symptoms improve after a gluten-free diet (GFD). However, to date, it is uncertain if gluten or other components of wheat are responsible for these symptoms. The aim of this study was to compare the effects of an organic durum wheat variety with those of standard commercial wheat in patients with known NCGS. We performed a double-blind randomized cross-over trial of 42 patients (mean age 45 years, 8 men) with NCGS diagnosed according to the Salerno criteria and adherence to GFD for at least 12 weeks from screening. Enrolled subjects were randomly assigned to one the following groups of treatment: (A) a two-week diet with Senatore Cappelli wheat variety pasta; (B) a two-week diet with standard commercial pasta. Then, after a two-week washout period on gluten-free diet, each patient crossed over to the other treatment group. Symptoms were assessed through a modified version of the Gastrointestinal Symptom Rating Scale (GSRS), tailored on NCGS. Between April 2018 and July 2018, 42 patients with NCGS were enrolled in the study (70.6% females), and 34 patients completed the study. Patients reported lower overall symptoms scores after eating Senatore Cappelli pasta than standard pasta (p = 0.03) and also significantly lower scores in several specific gastrointestinal and extra-intestinal symptoms after eating Senatore Cappelli pasta than standard pasta, specifically, bloating (p = 0.04), abdominal distention (p = 0.004), eructation (p = 0.01), flatus (p = 0.02), feeling of incomplete evacuation (p = 0.001), dermatitis (p = 0.01), and limb numbness (p = 0.03). In our study, patients with NCGS experienced lower gastrointestinal and extra-intestinal symptom scores after eating the Senatore Cappelli wheat variety than a standard commercial wheat. Should our preliminary results be confirmed by further studies, new dietary alternatives may be available to patients with NCGS, with consequent health, economic, and social benefits.

Project description:In this investigation, we reported the production of prototype breads from the processed flours of three specific Triticum turgidum wheat genotypes that were selected in our previous investigation for their potential low toxic/immunogenic activity for celiac disease (CD) patients. The flours were subjected to sourdough fermentation with a mixture of selected Lactobacillus strains, and in presence of fungal endoproteases. The breads were characterized by R5 competitive enzyme linked immunosorbent assay in order to quantify the residual gluten, and the differential efficacy in gluten degradation was assessed. In particular, two of them were classified as gluten-free (<20 ppm) and very low-gluten content (<100 ppm) breads, respectively, whereas the third monovarietal prototype retained a gluten content that was well above the safety threshold prescribed for direct consumption by CD patients. In order to investigate such a genotype-dependent efficiency of the detoxification method applied, an advanced proteomic characterization by high-resolution tandem mass spectrometry was performed. Notably, to the best of our knowledge, this is the first proteomic investigation which benefitted, for protein identification, from the full sequencing of the Triticum turgidum ssp. durum genome. The differences of the proteins' primary structures affecting their susceptibility to hydrolysis were investigated. As a confirmation of the previous immunoassay-based results, two out of the three breads made with the processed flours presented an exhaustive degradation of the epitopic sequences that are relevant for CD immune stimulatory activity. The list of the detected epitopes was analyzed and critically discussed in light of their susceptibility to the detoxification strategy applied. Finally, in-vitro experiments of human gastroduodenal digestion were carried out in order to assess, in-silico, the toxicity risk of the prototype breads under investigation for direct consumption by CD patients. This approach allowed us to confirm the total degradation of the epitopic sequences upon gastro-duodenal digestion.

Project description:Gluten strength is one of the factors that determine the end-use quality of durum wheat and is an important breeding target for this crop. To characterize the quantitative trait loci (QTL) controlling gluten strength in Canadian durum wheat cultivars, a population of 162 doubled haploid (DH) lines segregating for gluten strength and derived from cv. Pelissier × cv. Strongfield was used in this study. The DH lines, parents, and controls were grown in 3 years and two seeding dates in each year and gluten strength of grain samples was measured by sodium dodecyl sulfate (SDS)-sedimentation volume (SV). With a genetic map created by genotyping the DH lines using the Illumina Infinium iSelect Wheat 90K SNP (single nucleotide polymorphism) chip, QTL contributing to gluten strength were detected on chromosome 1A, 1B, 2B, and 3A. Two major and stable QTL detected on chromosome 1A (QGlu.spa-1A) and 1B (QGlu.spa-1B.1) explaining 13.7-18.7% and 25.4-40.1% of the gluten strength variability respectively were consistently detected over 3 years, with the trait increasing alleles derived from Strongfield. Putative candidate genes underlying the major QTL were identified. Two novel minor QTL (QGlu.spa-3A.1 and QGlu.spa-3A.2) with the trait increasing allele derived from Pelissier were mapped on chromosome 3A explaining up to 8.9% of the phenotypic variance; another three minor QTL (QGlu.spa-2B.1, QGlu.spa-2B.2, and QGlu.spa-2B.3) located on chromosome 2B explained up to 8.7% of the phenotypic variance with the trait increasing allele derived from Pelissier. QGlu.spa-2B.1 is a new QTL and has not been reported in the literature. Multi-environment analysis revealed genetic (QTL) × environment interaction due to the difference of effect in magnitude rather than the direction of the QTL. Eleven pairs of digenic epistatic QTL were identified, with an epistatic effect between the two major QTL of QGlu.spa-1A and QGlu.spa-1B.1 detected in four out of six environments. The peak SNPs and SNPs flanking the QTL interval of QGlu.spa-1A and QGlu.spa-1B.1 were converted to Kompetitive Allele Specific PCR (KASP) markers, which can be deployed in marker-assisted breeding to increase the efficiency and accuracy of phenotypic selection for gluten strength in durum wheat. The QTL that were expressed consistently across environments are of great importance to maintain the gluten strength of Canadian durum wheat to current market standards during the genetic improvement.

Dataset Information

The Diverse Potential of Gluten from Different Durum Wheat Varieties in Triggering Celiac Disease: A Multilevel In Vitro, Ex Vivo and In Vivo Approach.

Publications

The Diverse Potential of Gluten from Different Durum Wheat Varieties in Triggering Celiac Disease: A Multilevel In Vitro, Ex Vivo and In Vivo Approach.

Similar Datasets

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