Rye Proteins Research Articles

The role of starch digestion in the brewing of gluten-free beers

Celiac disease affects an estimated 1% of the global population, and symptoms are triggered by the consumption of gluten in sensitized individuals. Other gluten sensitivities exist but are poorly characterized or often under-diagnosed, leading a number of people to pursue a gluten-free or gluten-reduced diet even in the absence of a diagnosis. Gluten is a complex suite of proteins from the prolamin fraction of storage proteins in barley, wheat, and rye, meaning that foods or beverages that are produced from these cereals will also contain gluten. Beer is one such beverage that is traditionally produced from malted barley and is thus incompatible with a gluten-free lifestyle. The simplest option to produce a gluten-free beer would seem to be to use alternative, gluten-free, grains such as millet, sorghum, teff, rice, corn or other starchy materials. However, barley has thousands of years of development as a brewing agent behind it and these alternative ingredients cannot be incorporated into the brewing process without substantial alterations. Particularly, these ingredients generally have much lower endogenous starch degrading activity and much higher starch gelatinization temperatures as compared to barley which makes generating sufficient fermentable sugars for yeast fermentation a challenge. In this review we will further elucidate these challenges as well as the solutions that are being developed to bring beer to the gluten-free masses.

The Brachypodium distachyon DREB transcription factor BdDREB-39 confers oxidative stress tolerance in transgenic tobacco.

Key message BdDREB-39 is a DREB/CBF transcription factor, localized in the nucleus with transactivation activity, and BdDREB-39-overexpressing transgenic yeasts and tobacco enhanced the tolerance to oxidative stress.Abstract The DREB/CBF transcription factors are generally recognized to play an important factor in plant growth, development and response to various abiotic stresses. However, the mechanism of DREB/CBFs in oxidative stress response is largely unknown. This study isolated a DREB/CBF gene BdDREB-39 from Brachypodium distachyon (B. distachyon). Multiple sequence alignment and phylogenetic analysis showed that BdDREB-39 was closely related to the DREB proteins of oats, barley, wheat and rye and therefore its study can provide a reference for the excavation and genetic improvement of BdDREB-39 or its homologs in its closely related species. The transcript levels of BdDREB-39 were significantly up-regulated under H2O2 stress. BdDREB-39 was localised in the nucleus and functioned as a transcriptional activator. Overexpression of BdDREB-39 enhanced H2O2 tolerance in yeast. Transgenic tobaccos with BdDREB-39 had higher germination rates, longer root, better growth status, lesser reactive oxygen species (ROS) and malondialdehyde (MDA), and higher superoxide dismutase (SOD) and peroxidase (POD) activities than wild type (WT). The expression levels of ROS-related and stress-related genes were improved by BdDREB-39. In summary, these results revealed that BdDREB-39 can improve the viability of tobacco by regulating the expression of ROS and stress-related genes, allowing transgenic tobacco to accumulate lower levels of ROS and reducing the damage caused by ROS to cells. The BdDREB-39 gene has the potential for developing plant varieties tolerant to stress.

Genome-wide identification of the plant homeodomain-finger family in rye and ScPHD5 functions in cold tolerance and flowering time.

111 PHD genes were newly identified in rye genome and ScPHD5's role in regulating cold tolerance and flowering time was suggested. Plant homeodomain (PHD)-finger proteins regulate the physical properties of chromatin and control plant development and stress tolerance. Although rye (Secale cereale L.) is a major winter crop, PHD-finger proteins in rye have not been studied. Here, we identified 111 PHD genes in the rye genome that exhibited diverse gene and protein sequence structures. Phylogenetic tree analysis revealed that PHDs were genetically close in monocots and diverged from those in dicots. Duplication and synteny analyses demonstrated that ScPHDs have undergone several duplications during evolution and that high synteny is conserved among the Triticeae species. Tissue-specific and abiotic stress-responsive gene expression analyses indicated that ScPHDs were highly expressed in spikelets and developing seeds and were responsive to cold and drought stress. One of these genes, ScPHD5, was selected for further functional characterization. ScPHD5 was highly expressed in the spike tissues and was localized in the nuclei of rye protoplasts and tobacco leaves. ScPHD5-overexpressing Brachypodium was more tolerant to freezing stress than wild-type (WT), with increased CBF and COR gene expression. Additionally, these transgenic plants displayed an extremely early flowering phenotype that flowered more than two weeks earlier than the WT, and vernalization genes, rather than photoperiod genes, were increased in the WT. RNA-seq analysis revealed that diverse stress response genes, including HSPs, HSFs, LEAs, and MADS-box genes, were also upregulated in transgenic plants. Our study will help elucidate the roles of PHD genes in plant development and abiotic stress tolerance in rye.

Genome-wide identification and functional analysis of the TCP gene family in rye (Secale cereale L.)

TEOSINTE BRANCHED1/CYCLOIDEA/PCF (TCP) proteins are plant-specific transcription factors that play significant roles in plant growth, development, and stress response. Rye is a high-value crop with strong resistance to adverse environments. However, the functions of TCP proteins in rye are rarely reported. Based on a genome-wide analysis, the present study identified 26 TCP genes (ScTCPs) in rye. Mapping showed an uneven distribution of the ScTCP genes on the seven rye chromosomes and detected three pairs of tandem duplication genes. Phylogenetic analysis divided these genes into PCF (Proliferrating Cell Factors), CIN (CINCINNATA), and CYC (CYCLOIDEA)/TB1 (Teosinte Branched1) classes, which showed the highest homology between rye and wheat genes. Analysis of miRNA targeting sites indicated that five ScTCP genes were identified as potential targets of miRNA319. Promoter cis-acting elements analysis indicated that ScTCPs were regulated by light signals. Further analysis of the gene expression patterns and functional annotations suggested the role of a few ScTCPs in grain development and stress response. In addition, two TB1 homologous genes (ScTCP9 and ScTCP10) were identified in the ScTCP family. Synteny analysis showed that TB1 orthologous gene pairs existed before the ancestral divergence. Finally, the yeast two-hybrid assay and luciferase complementation imaging assay proved that ScTCP9, localized in the nucleus, interacts with ScFT (Flowering locus T), indicating their role in regulating flowering time. Taken together, this comprehensive study of ScTCPs provides important information for further research on gene function and crop improvement.

Survey of ACCase and ALS resistance in winter annual grasses identifies target-site and nontarget-site imazamox resistance in Secale cereale.

Early detection of herbicide resistance in weeds is crucial for successful implementation of integrated weed management. We conducted a herbicide resistance survey of the winter annual grasses feral rye (Secale cereale), downy brome (Bromus tectorum), and jointed goatgrass (Aegilops cylindrica) from Colorado winter wheat production areas for resistance to imazamox and quizalofop. All samples were susceptible to quizalofop. All downy brome and jointed goatgrass samples were susceptible to imazamox. Out of 314 field collected samples, we identified three feral rye populations (named A, B, and C) that were imazamox resistant. Populations B and C had a target-site mechanism with mutations in the Ser653 residue of the acetolactate synthase (ALS) gene to Asn in B and to Thr in C. Both populations B and C had greatly reduced ALS in vitro enzyme inhibition by imazamox. ALS feral rye protein modeling showed that steric interactions induced by the amino acid substitutions at Ser653 impaired imazamox binding. Individuals from population A had no mutations in the ALS gene. The ALS enzyme from population A was equally sensitive to imazamox as to known susceptible feral rye populations. Imazamox was degraded two times faster in population A compared with a susceptible control. An oxidized imazamox metabolite formed faster in population A and this detoxification reaction was inhibited by malathion. Population A has a nontarget-site mechanism of enhanced imazamox metabolism that may be conferred by cytochrome P450 enzymes. This is the first report of both target-site and metabolism-based imazamox resistance in feral rye. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Development of a Sequence Searchable Database of Celiac Disease-Associated Peptides and Proteins for Risk Assessment of Novel Food Proteins.

Celiac disease (CeD) is an autoimmune enteropathy induced by prolamin and glutelin proteins in wheat, barley, rye, and triticale recognized by genetically restricted major histocompatibility (MHC) receptors. Patients with CeD must avoid consuming these proteins. Regulators in Europe and the United States expect an evaluation of CeD risks from proteins in genetically modified (GM) crops or novel foods for wheat-related proteins. Our database includes evidence-based causative peptides and proteins and two amino acid sequence comparison tools for CeD risk assessment. Sequence entries are based on the review of published studies of specific gluten-reactive T cell activation or intestinal epithelial toxicity. The initial database in 2012 was updated in 2018 and 2022. The current database holds 1,041 causative peptides and 76 representative proteins. The FASTA sequence comparison of 76 representative CeD proteins provides an insurance for possible unreported epitopes. Validation was conducted using protein homologs from Pooideae and non-Pooideae monocots, dicots, and non-plant proteins. Criteria for minimum percent identity and maximum E-scores are guidelines. Exact matches to any of the 1,041 peptides suggest risks, while FASTA alignment to the 76 CeD proteins suggests possible risks. Matched proteins should be tested further by CeD-specific CD4/8+ T cell assays or in vivo challenges before their use in foods.

Beneficial Role of Microbial Transglutaminase in the Pathogenetic Mechanisms of Coeliac Disease.

Coeliac disease (CD) is caused by immunological intolerance to wheat gluten and related proteins of rye and barley. Consequently, gluten-free (GF) products have been developed but technological implementation is required to improve their intrinsic rheological properties. One alternative for increasing the functional properties of GF foodstuff is the incorporation of microbial transglutaminase (mTG), which allows for the cross-linking of proteins that can substitute for the gluten network in the bakery industry. mTG has been, however, suggested to mimic tissue transglutaminase and to be immunogenic in CD patients. Recently, both mTG and gliadin were found to be transported to the endoplasmic reticulum of enterocytes, suggesting cross-presentation and potential interaction with immune cells in CD. Although pathogenetic activity of mTG has not been found to date, these data naturally raise concerns among clinicians and patients about the use of mTG as a food additive. On the contrary, different studies have shown that treatment with mTG was effective in reducing the inflammatory immune response of gluten in CD. In this article, we take advantage of recent advances in gut physiology and CD pathogenesis to revise the literature data on mTG. An updated and unbiased overview of the role of mTG in this pathology allowed us to definitively highlight the beneficial use of this food additive by CD patients.

Impact of condensed tannin interactions with grain proteins and non-starch polysaccharides on batter system properties

Proanthocyanidins (PA) cross-link wheat gluten proteins and dramatically enhance batter viscosity; PA could similarly affect related grains. This study aimed to determine PA effect on viscosity and pasting properties of barley, rye, and oat flours, and the relative contributions of PA interactions with proteins and non-starch polysaccharides (NSP). PA significantly increased batter viscosity, stability, and RVA peak viscosity in rye and barley flours (2.8× and 1.2×, respectively). Interestingly, viscosity peaked distinctively ~75 °C in PA-treated rye and barley flours, and their isolated protein-starch systems, indicating prolamins unravelled and complexed with PA during heating. Oat was largely unaffected by PA, likely because of its protein composition. Furthermore, water-soluble rye NSP and arabinoxylans, but not barley β-glucans, significantly increased starch pasting viscosity with PA; oxidative gelation was not a factor. Thus, rye flour viscosity dramatically increased through interactive effects of PA on rye proteins and NSP, which could expand its food applications.

Functional Properties of Rye Prolamin (Secalin) and Their Improvement by Protein Lipophilization through Capric Acid Covalent Binding.

Secalin (SCL), the prolamin fraction of rye protein, was chemically lipophilized using acylation reaction by treatment with different amounts of capric acid chloride (0, 2, 4, and 6 mmol/g) to enhance its functional properties. It was shown that SCL lipophilization increased the surface hydrophobicity and the hydrophobic interactions, leading to a reduction in protein solubility and water absorption capacity and to a greater oil absorption. In addition, SCL both emulsifying capacity and stability were improved when the protein was treated with low amount of capric acid chloride. Finally, the foaming capacity of SCL markedly increased after its treatment with increasing concentrations of the acylating agent, even though the foam of the modified protein was found to be more stable at the lower level of protein acylation. Technological application of lipophilized SCL as a protein additive in food preparations is suggested.

Gastrointestinal Problems—Autism Toolkit

Gastrointestinal (GI) problems include constipation, diarrhea, reflux, vomiting, belly pain, and feeding problems. Some children with autism spectrum disorder (ASD) have GI problems. These symptoms can add stress to the child and family and may cause behaviors such as aggression or self-injury.As of now, there is no scientific proof that GI problems directly cause ASD. It is possible that some people have genes that cause ASD and also cause GI problems. There is no evidence that children with ASD have abnormalities in their intestines, or overgrowth of yeast, or other organisms that make behaviors worse.A child with ASD who has GI problems should be examined just as any other child would be. Constipation and diarrhea may be caused by diet, so children who are picky eaters may be at higher risk depending on what they eat and drink. Children with ASD may not be able to tell people about GI pain and may therefore show it through other behaviors. For example, if they have stomach pain, they may act out as a way to let parents know because they cannot talk. Children who have trouble toilet training and those who don’t get enough fiber may develop constipation.A pediatrician will talk to you and do a physical examination. She will ask about diet history and weight gain. The doctor may request an evaluation by a pediatric gastroenterologist (a doctor with additional training in GI diseases) or nutritionist or request laboratory tests. As of now, children who do not have signs of GI problems should not have any special tests. The pediatrician might consider specialized evaluations if a child has signs of GI problems. For example, the doctor might use an x-ray to find out more about severe constipation. Some children do not digest milk well and benefit from trying a milk-free diet or enzyme replacement to help digest “milk sugar” (lactose). This is a common problem. Rarely, children will have an immune response to gluten (celiac disease) and need to avoid wheat and other grain products. There is a blood test your child’s pediatrician can do to determine whether your child needs to see a specialist to evaluate for celiac disease. For ongoing GI problems, your child’s doctor might refer your child to a gastroenterologist to get a more in-depth look at your child.If medical testing shows that your child has a GI problem, a pediatrician will recommend treatment for that problem. Treatments may include extra fiber in the diet, stool lubrication or stool softener for constipation, more tests for gastroesophageal reflux, or medications.Sometimes, families take out milk or wheat from their child’s diet to test for relief of GI symptoms with the hope that other behaviors of ASD will improve. This gluten-free/casein-free dietary treatment is popular, but there are little scientific data supporting its general use. Some children might have symptoms of lactose intolerance (intolerance of milk sugar). Getting rid of milk will decrease chance of irritation and harden loose stools in affected children. Celiac disease (a reaction to gluten proteins in barley, rye, and wheat) is found in 1 in 100 people. Children with ASD, especially if they have a family history of celiac disease, should be screened if they have any stomach pain or problems with bowel movements, slow growth, or poor appetite.With ongoing research, doctors will keep learning about GI treatment of children with ASD.The information contained in this resource should not be used as a substitute for the medical care and advice of your pediatrician. There may be variations in treatment that your pediatrician may recommend based on individual facts and circumstances. Original resource included as part of Caring for Children With Autism Spectrum Disorder: A Practical Resource Toolkit for Clinicians, 3rd Edition.Inclusion in this resource does not imply an endorsement by the American Academy of Pediatrics (AAP). The AAP is not responsible for the content of the resources mentioned in this resource. Website addresses are as current as possible but may change at any time.The American Academy of Pediatrics (AAP) does not review or endorse any modifications made to this resource and in no event shall the AAP be liable for any such changes.

The Circulating Midkine in the Newly Diagnosed Celiac Disease: Clinical Implications.

Background:Celiac disease (CeD) is a chronic inflammatory small intestine disorder caused by an abnormal immune response to an array of the epitopes of the wheat gluten and related proteins of rye and barley in genetically susceptible individuals. Midkine (MK) is an angiogenic cytokine, chemotactic in the direction of polymorphonuclear neutrophils and macrophages, and a T-regulatory cell suppressor. So far, a possible relationship with CeD has not yet been explored. Diagnosis of CeD is based on serologic test in a clinical setting suggestive of CeD and confirmatory histologic examination of the duodenal biopsy. Sometimes, genetic testing of human leukocyte antigen (HLA)-DQ2 and HLA-DQ8 may be needed. The objective of this study was to measure and compare the circulating MK in the celiac patients and healthy individuals.Materials and Methods:Twenty newly untreated CeD cases and 20 normal controls were enrolled in this study. The enzyme-linked immunosorbent assay was used to measure the circulating MK in the celiac patients and controls.Results:There was insignificant difference in the circulating MK between the patients and controls (P > 0.05).Conclusions:The study results suggest that the MK marker does not have any diagnostic value in CeD activity to be used at the time of diagnosis or during follow-ups.

Mapping Coeliac Toxic Motifs in the Prolamin Seed Storage Proteins of Barley, Rye, and Oats Using a Curated Sequence Database.

Wheat gluten, and related prolamin proteins in rye, barley and oats cause the immune-mediated gluten intolerance syndrome, coeliac disease. Foods labelled as gluten-free which can be safely consumed by coeliac patients, must not contain gluten above a level of 20 mg/Kg. Current immunoassay methods for detection of gluten can give conflicting results and may underestimate levels of gluten in foods. Mass spectrometry methods have great potential as an orthogonal method, but require curated protein sequence databases to support method development. The GluPro database has been updated to include avenin-like sequences from bread wheat (n = 685; GluPro v1.1) and genes from the sequenced wheat genome (n = 699; GluPro v 1.2) and Triticum turgidum ssp durum (n = 210; GluPro v 2.1). Companion databases have been developed for prolamin sequences from barley (n = 64; GluPro v 3.0), rye (n = 41; GluPro v 4.0), and oats (n = 27; GluPro v 5.0) and combined to provide a complete cereal prolamin database, GluPro v 6.1 comprising 1,041 sequences. Analysis of the coeliac toxic motifs in the curated sequences showed that they were absent from the minor avenin-like proteins in bread and durum wheat and barley, unlike the related avenin proteins from oats. A comparison of prolamin proteins from the different cereal species also showed α- and γ-gliadins in bread and durum wheat, and the sulphur poor prolamins in all cereals had the highest density of coeliac toxic motifs. Analysis of ion-mobility mass spectrometry data for bread wheat (cvs Chinese Spring and Hereward) showed an increased number of identifications when using the GluPro v1.0, 1.1 and 1.2 databases compared to the limited number of verified sequences bread wheat sequences in reviewed UniProt. This family of databases will provide a basis for proteomic profiling of gluten proteins from all the gluten containing cereals and support identification of specific peptide markers for use in development of new methods for gluten quantitation based on coeliac toxic motifs found in all relevant cereal species.

X6: A Novel Antibody for Potential Use in Gluten Quantification.

Gliadin is a fraction of gluten, known to trigger celiac disease in susceptible people. To date, the life-long gluten-free diet is used for the prevention of this disease. Hence, methods for gluten control in foods are of significant importance. Being one of the most-used methods used for this purpose, ELISA should use high-affinity antibodies to gliadin peptides involved into celiac process. This study investigates the characteristics of a novel anti-gliadin antibody X6. We found the QXQPFPXP site to be a recognized epitope that provides specific binding of the antibody to cereal prolamins involved in celiac disease manifestation. A specificity study using immunoblotting shows the recognition of wheat, barley and rye proteins—as well as α-gliadin homologs from non-edible cereals (Dasypyrum villosum). Reactivity to avenin was less pronounced, as this protein does not contain the PFP motif most critical for antibody recognition. The proteins of Zea mays and Setaria italica were not recognized by X6. X6-based ELISA highly correlated with R5 and G12, which are Codex Alimentarius standards in the quantitative assessment of gluten content (Pearson’s R = 0.86 and 0.87, respectively). Qualitative assessment revealed no significant differences between R5 and G12 and X6.

The major constituents of rye (Secale cereale L.) flour and their role in the production of rye bread, a food product to which a multitude of health aspects are ascribed

AbstractBackgroundRye (Secale cereale L.) is, second to wheat (Triticum aestivum L.), the most common cereal used for producing bread. Rye‐based breads are of great interest because of their high dietary fiber and lysine contents and because of their health effects. However, the bread‐making performance of rye flour is inferior to that of wheat flour.FindingsThis review summarizes the current knowledge about the major rye constituents (starch, proteins, and arabinoxylans), their role during bread making, and the potential health implications of rye bread consumption. Rye proteins play only a minor role during bread making, but their exact role remains insufficiently explored. In contrast to wheat proteins, they lack the ability to form a network with good gas‐retaining properties. The rye bread crumb structure is partly attributed to its starch and arabinoxylan polysaccharides. Both bind a lot of water, form networks, and bring about the rye bread structure.ConclusionsNevertheless, a better understanding of the role of rye starch, proteins, and arabinoxylans, their interactions, and their impact on dough structure or product quality still needs further exploration. In terms of health effects, EFSA has acknowledged that rye fiber contributes to normal bowel function. There are also indications that rye bread consumption has a number of positive effects such as reducing the risk of colon cancer and hunger feelings and cholesterol‐lowering effects. However, although beneficial health effects of rye‐bread consumption are shown, more research is needed.Significance and noveltyA literature search with the terms “rye” and “review” in Web of Science in April 2020 resulted in only six manuscripts. Therefore, this review summarizes the current knowledge about the major rye constituents (starch, proteins, and arabinoxylans), their role during bread making, and the potential health implications of rye bread consumption.

A novel approach to produce phage single domain antibody fragments for the detection of gluten in foods

In this study, we demonstrated the feasibility of isolating recombinant phage-antibodies against gluten from a non-immunized library of human single-domain antibodies (dAbs). Phage display technology enabled the selection of affinity probes by successive rounds of biopanning against a biotinylated synthetic peptide comprising repetitive immunogenic gluten motifs. The analysis of a wide representation of heterologous plant species corroborated that two of the isolated clones were specific to wheat, barley and rye proteins. The phage antibody selected as the most appropriate clone for the detection of gluten in foods (dAb8E-phage) was further applied in an indirect ELISA to the analysis of 50 commercial food samples. Although the limit of detection achieved did not improve those of current immunoassays, the proposed methodology could provide promising new pathways for the generation of recombinant antibodies that allow a comprehensive determination of gluten in foods, whilst replacing the need for animal immunization.

Linear and Non-linear Rheology of Bread Doughs Made from Blends of Wheat (Triticum aestivum L.) and Rye (Secale cereale L.) Flour

Strain hardening of wheat flour (WF) proteins during fermentation and baking is at the basis of the excellent quality of wheat breads. Yet, there are good reasons to use rye flour (RF), such as its high content of dietary fiber and other bioactives. However, rye proteins lack the ability to form dough with sufficient strain hardening capacity under extension, which partially explains the inferior quality of rye breads. A promising strategy to make high-quality breads with high nutritional value is the use of WF and RF blends. Despite this, the rheological behavior of WF/RF bread doughs has not yet been investigated. This study examines their rheology by non-linear uniaxial extensional and small-amplitude oscillatory shear measurements, and relates these measurements to the quality of the resulting breads. The strain hardening index (SHI), derived from extensional measurements, is expressed as the transient extensional viscosity at maximum strain [ηe+(εmax)] relative to the linear extensional viscosity extrapolated to maximum strain [ηe0+(εmax)]. Both the SHI and bread volume decreased when the level of RF in the blend increased. As an example, replacing 60% WF by RF led to a relative decrease of the SHI and bread volume of 34 and 64%, respectively. The decrease of the SHI is correlated to an increase in ηe0+(εmax). Comparison of the extensional curves of doughs made with blends of WF and RF and doughs made with blends of WF and wheat starch (WS) further revealed that RF components play a major role in the rheology of the doughs. In conclusion, it was found that WF/RF blends need to contain at least 60% WF to obtain bread of proper quality.

On the immune response to barley in celiac disease: Biased and public T-cell receptor usage to a barley unique and immunodominant gluten epitope.

Celiac disease (CeD) is driven by CD4+ T-cell responses to dietary gluten proteins of wheat, barley, and rye when deamidated gluten epitopes are presented by certain disease-associated HLA-DQ allotypes. About 90% of the CeD patients express HLA-DQ2.5. In such patients, five gluten epitopes dominate the anti-gluten T-cell response; two epitopes unique to wheat, two epitopes present in wheat, barley, and rye and one epitope unique to barley. Despite presence of barley in commonly consumed food and beverages and hence being a prominent source of gluten, knowledge about T-cell responses elicited by barley in CeD is scarce. Therefore, in this study, we explored T-cell response toward the barley unique epitope DQ2.5-hor-3 (PIPEQPQPY) by undertaking HLA-DQ:gluten peptide tetramer staining, single-cell T-cell receptor (TCR) αβ sequencing, T-cell cloning, and T-cell proliferation studies. We demonstrate that majority of the CeD patients generate T-cell response to DQ2.5-hor-3, and this response is characterized by clonal expansion, preferential TCR V-gene usage and public TCR features thus echoing findings previously made for wheat gluten epitopes. The knowledge that biased and public TCRs underpin the T-cell response to all the immunodominant gluten epitopes in CeD suggests that such T cells are promising diagnostic and therapeutic targets.

Assessing the Utility of Multiplexed Liquid Chromatography-Mass Spectrometry for Gluten Detection in Australian Breakfast Food Products.

Coeliac disease (CD) is an autoimmune disorder triggered by the ingestion of gluten that is associated with gastrointestinal issues, including diarrhea, abdominal pain, and malabsorption. Gluten is a general name for a class of cereal storage proteins of wheat, barley, and rye that are notably resistant to gastrointestinal digestion. After ingestion, immunogenic peptides are subsequently recognized by T cells in the gastrointestinal tract. The only treatment for CD is a life-long gluten-free diet. As such, it is critical to detect gluten in diverse food types, including those where one would not expect to find gluten. The utility of liquid chromatography-mass spectrometry (LC-MS) using cereal-specific peptide markers to detect gluten in heavily processed food types was assessed. A range of breakfast products, including breakfast cereals, breakfast bars, milk-based breakfast drinks, powdered drinks, and a savory spread, were tested. No gluten was detected by LC-MS in the food products labeled gluten-free, yet enzyme-linked immunosorbent assay (ELISA) measurement revealed inconsistencies in barley-containing products. In products containing wheat, rye, barley, and oats as labeled ingredients, gluten proteins were readily detected using discovery proteomics. Panels comprising ten cereal-specific peptide markers were analyzed by targeted proteomics, providing evidence that LC-MS could detect and differentiate gluten in complex matrices, including baked goods and milk-based products.

Celiac disease and its histopathology

Introduction: Celiac disease, also known as celiac sprue, non-tropical sprue, gluten-induced enteropathy, or gluten-sensitive enteropathy (GSE), is a chronic immune-mediated disorder of the small intestine characterized by malabsorption after ingestion of wheat gluten or related proteins in rye (secalins) and barley (hordeins) in individuals with a certain genetic background. Materials and Method: The study was conducted in Mahatma Gandhi Medical College and hospital, Jaipur. 53 Duodenal biopsies were taken who have symptoms of diarrhea, iron deficiency anemia, amenorrhea, recurrent mouth ulceration. All biopsies taken from duodenal sitewere brought in 10% buffered formalin. After overnight fixation in formalin, the tissues were processed in automated tissue processor for dehydration, clearing, and paraffin embedding. Sections were cut in rotary microtome 4 micrometer thickness. The section was stained for Hematoxylin and Eosin stain. Results: The present study included 53 Duodenal biopsies. Out of 53 duodenal biopsies, 12 biopsies have shown presence of Celiac disease. Rest 41 biopsies have shown chronic duodenitis. Conclusion: Celiac disease is a common autoimmune condition with mainly intestinal, but also extra-intestinal manifestations. The histologic hallmark of GlutenSensitive Enteropathy is both increased inflammation and architectural derangement in the small intestinal villi.The small intestinal mucosa of untreated persons with celiac disease usually display villous atrophy (Marsh III), or, less commonly, isolated crypt hyperplasia (Marsh II), together with intraepithelial lymphocytic proliferation.

Correlation of Serum Tissue Transglutaminase Antibody Levels and Marsh Grading of Duodenal Biopsy in Patients of Celiac Disease

Background: Celiac disease (CD) is a chronic small intestinal immune-mediated enteropathy precipitated in genetically predisposed individuals by ingestion of wheat gluten and related proteins of barley and rye. It is an under-diagnosed gluten-sensitive enteropathy, usually presenting with atypical/extra-intestinal features. Duodenal biopsy is considered as the gold standard in the diagnosis of CD. Serum anti tissue transglutaminase (anti-tTG) antibodies is auto antibodies directed against tissue transglutaminase (tTG) enzyme. Aims and objective: The aim of the study is to find out the prevalence of CD among clinically suspected cases of malabsorption, to assess the correlation of anti-tTG levels with modified Marsh grading of duodenal biopsy, to find out sensitivity, specificity and predictive values of anti-tTG test and to calculate the cut off values of anti-tTG diagnostic of high grade CD. Methodology: The present study included 118 clinically suspected patients of malabsorption in which both duodenal biopsy and anti-tTG test was done. The mucosal damage graded according to the modified Marsh grading and anti-tTG tests were done in serum using ELISA kit. Results: The prevalence of CD among clinically suspected cases of malabsorption was 12.7%. The incidence was highest in middle aged male patients. The most common clinical symptom was chronic diarrhea and commonest presenting sign was anemia. Anti-tTG levels were found to be proportionately increasing with the severity of CD and showed a positive correlation(r=0.433). Patients with anemia and chronic diarrhea showed statistically significant (p value=0.0242 and 0.0089 respectively) mean anti-tTG levels in Celiac patients in relation to non-Celiac patients. Although the mean levels of anti-tTG was 6.3 times higher than baseline in grade 3 disease, the ROC analysis showed that anti-tTG in isolation had no significance in exactly grading the disease (AUC=0.30). Conclusion: Positive correlation was found between anti-tTG levels and modified Marsh grading. Anti-tTG showed proportionate increase with severity of duodenal damage. Anti-tTG test showed low sensitivity (42.86%) and high specificity (84.34%). Hence it is not a reliable tool in isolation to screen CD but should be used in combination with duodenal biopsy. It can be used to assess the severity of CD and for follow up.