RESUMO
Beer is a beverage that contains gluten and cannot be consumed by people with celiac disease. In this context, the enzyme prolyl endoprotease (PEP) can be used to reduce the gluten content in beer. The present study aimed to produce the PEP from Aspergillus sp. FSDE 16 using solid-state fermentation with 5 conditions and comparing with a similar commercial enzyme produced from Aspergillus niger in the production of a gluten-free beer. The results of the performed cultures showed that during the culture, the most increased protease activity (54.46 U/mL) occurred on the 4th day. In contrast, for PEP, the highest activity (0.0356 U/mL) was obtained on the 3rd day of culture in condition. Regarding beer production, cell growth, pH, and total soluble solids showed similar behavior over the 7 days for beers produced without enzyme addition or with the addition of commercial enzyme and with the addition of the enzyme extract produced. The addition of the enzyme and the enzyme extract did not promote changes, and all the beers produced showed similar and satisfactory results, with acid pH between 4 and 5, total soluble solids ranging from 4.80 to 5.05, alcohol content ranging from 2.83% to 3.08%, and all beers having a dark character with deep amber and light copper color. Gluten removal was effectively using the commercial enzyme and the enzyme produced according to condition (v) reaching gluten concentrations equal to 17 ± 5.31 and 21.19 ± 11.28 ppm, respectively. In this way, the production of the enzyme by SSF and its application in the removal of gluten in beer was efficient.
Assuntos
Cerveja , Serina Endopeptidases , Humanos , Cerveja/análise , Serina Endopeptidases/química , Serina Endopeptidases/metabolismo , Prolil Oligopeptidases , Fermentação , Glutens/análise , Glutens/metabolismo , Aspergillus niger , Extratos VegetaisRESUMO
Non-alcoholic fatty liver disease (NAFLD) is the most prevalent chronic liver disorder in the world. We have seen that gluten intake exacerbated obesity and atherosclerosis in apolipoprotein E knockout (ApoE-/-) mice. In this study, we investigated the effect of gluten consumption on inflammation and oxidative stress in the liver of mice with NAFLD. Male ApoE-/- mice were fed a gluten-free (GF-HFD) or gluten-containing (G-HFD) high-fat diet for 10 weeks. Blood, liver, and spleen were collected to perform the analyses. The animals of the gluten group had increased hepatic steatosis, followed by increased serum AST and ALT. Gluten intake increased hepatic infiltration of neutrophils, macrophages, and eosinophils, as well as the levels of chemotaxis-related factors CCL2, Cxcl2, and Cxcr3. The production of the TNF, IL-1ß, IFNγ, and IL-4 cytokines in the liver was also increased by gluten intake. Furthermore, gluten exacerbated the hepatic lipid peroxidation and nitrotyrosine deposition, which were associated with increased production of ROS and nitric oxide. These effects were related to increased expression of NADPH oxidase and iNOS, as well as decreased activity of superoxide dismutase and catalase enzymes. There was an increased hepatic expression of the NF-κB and AP1 transcription factors, corroborating the worsening effect of gluten on inflammation and oxidative stress. Finally, we found an increased frequency of CD4+FOXP3+ lymphocytes in the spleen and increased gene expression of Foxp3 in the livers of the G-HFD group. In conclusion, dietary gluten aggravates NAFLD, exacerbating hepatic inflammation and oxidative stress in obese ApoE-deficient mice.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Camundongos , Masculino , Animais , Hepatopatia Gordurosa não Alcoólica/etiologia , Hepatopatia Gordurosa não Alcoólica/metabolismo , Dieta Hiperlipídica/efeitos adversos , Glutens/metabolismo , Camundongos Knockout para ApoE , Fígado/metabolismo , Inflamação/metabolismo , Estresse Oxidativo , Apolipoproteínas E/genética , Fatores de Transcrição Forkhead/metabolismo , Camundongos Endogâmicos C57BLRESUMO
Callosobruchus maculatus is the main pest cowpea (Vigna unguiculata). Given its relevance as an insect pest, studies have focused in finding toxic compounds which could prevent its predatory action towards the seeds. Clitoria fairchildiana is a native Amazon species, whose seeds are refractory to insect predation. This characteristic was the basis of our interest in evaluating the toxicity of its seed proteins to C. maculatus larvae. Seed proteins were fractioned, according to their solubility, to albumins (F1), globulins (F2), kaphyrins (F3), glutelins (F4), linked kaphyrins (F5) and cross-linked glutelins (F6). The fractionated proteins were quantified, analysed by tricine-SDS-PAGE and inserted into the diet of this insect pest in order to evaluate their insecticidal potential. The most toxic fraction to C. maculatus, the propanol soluble F3, was submitted to molecular exclusion chromatography and all of the peaks obtained, F3P1, F3P2, F3P3, caused a reduction of larval mass, especially F3P1, seen as a major ~12 kDa electrophoretic band. This protein was identified as a vicilin-like protein by mass spectrometry and BLAST analysis. The alignment of the Cfvic (C. fairchildiana vicilin) peptides with a V. unguiculata vicilin sequence, revealed that Cfvic has at least five peptides (ALLTLVNPDGR, AILTLVNPDGR, NFLAGGKDNV, ISDINSAMDR, NFLAGEK) which lined up with two chitin binding sites (ChBS). This finding was corroborated by chitin affinity chromatography and molecular docking of chitin-binding domains for N-Acetyl-D-glucosamine and by the reduction of Cfvic chitin affinity after chemical modification of its Lys residues. In conclusion, Cfvic is a 12 kDa vicilin-like protein, highly toxic to C. maculatus, acting as an insect toxin through its ability to bind to chitin structures present in the insect midgut.
Assuntos
Clitoria , Besouros , Animais , Quitina/metabolismo , Clitoria/metabolismo , Besouros/metabolismo , Cotilédone/metabolismo , Glutens/análise , Glutens/metabolismo , Larva/metabolismo , Simulação de Acoplamento Molecular , Proteínas de Armazenamento de Sementes , Sementes/químicaRESUMO
Gluten-related disorders (GRDs) are a group of diseases that involve the activation of the immune system triggered by the ingestion of gluten, with a worldwide prevalence of 5%. Among them, Celiac disease (CeD) is a T-cell-mediated autoimmune disease causing a plethora of symptoms from diarrhea and malabsorption to lymphoma. Even though GRDs have been intensively studied, the environmental triggers promoting the diverse reactions to gluten proteins in susceptible individuals remain elusive. It has been proposed that pathogens could act as disease-causing environmental triggers of CeD by molecular mimicry mechanisms. Additionally, it could also be possible that unrecognized molecular, structural, and physical parallels between gluten and pathogens have a relevant role. Herein, we report sequence, structural and physical similarities of the two most relevant gluten peptides, the 33-mer and p31-43 gliadin peptides, with bacterial pathogens using bioinformatics going beyond the molecular mimicry hypothesis. First, a stringent BLASTp search using the two gliadin peptides identified high sequence similarity regions within pathogen-derived proteins, e.g., extracellular proteins from Streptococcus pneumoniae and Granulicatella sp. Second, molecular dynamics calculations of an updated α-2-gliadin model revealed close spatial localization and solvent-exposure of the 33-mer and p31-43 peptide, which was compared with the pathogen-related proteins by homology models and localization predictors. We found putative functions of the identified pathogen-derived sequence by identifying T-cell epitopes and SH3/WW-binding domains. Finally, shape and size parallels between the pathogens and the superstructures of gliadin peptides gave rise to novel hypotheses about activation of innate immunity and dysbiosis. Based on our structural findings and the similarities with the bacterial pathogens, evidence emerges that these pathologically relevant gluten-derived peptides could behave as non-replicating pathogens opening new research questions in the interface of innate immunity, microbiome, and food research.
Assuntos
Doença Celíaca/imunologia , Epitopos de Linfócito T , Gliadina/metabolismo , Glutens/metabolismo , Mimetismo Molecular , Fragmentos de Peptídeos/metabolismo , Carnobacteriaceae/metabolismo , Biologia Computacional , Gliadina/química , Gliadina/imunologia , Glutens/química , Glutens/imunologia , Humanos , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/imunologia , Streptococcus pneumoniae/metabolismoRESUMO
Grain protein composition is important in wheat quality and may influence the amino acidic sequence of bioactive peptides obtained from this feedstock. However, the genetic basis modulating the amino acid profile in durum wheat is not well-understood. Therefore, strong and weak gluten strength durum wheat genotypes were evaluated for their amino acid composition along grain filling. Strong gluten strength lines showed higher expression levels of low-molecular-weight glutenin-related genes between 21 and 35 days post-anthesis (DPA) and exhibited up to 43.5% more alanine than the weak lines at 42 DPA, which was supported by the higher expression levels of putative alanine amino transferase genes in strong genotypes. Therefore, with the involvement of chemistry and molecular biology, the results present here may influence the science of wheat.
Assuntos
Aminoácidos/metabolismo , Regulação da Expressão Gênica de Plantas , Glutens/metabolismo , Sementes/química , Triticum/genética , Aminoácidos/química , Regulação da Expressão Gênica no Desenvolvimento , Glutens/química , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Sementes/genética , Sementes/crescimento & desenvolvimento , Sementes/metabolismo , Triticum/química , Triticum/crescimento & desenvolvimento , Triticum/metabolismoRESUMO
Celiac disease (CD) is a chronic illness characterized by an inflammatory process triggered by gluten protein intake. Recent evidence has suggested that the lower relative abundance of bifidobacteria in the intestinal lumen may be associated with CD. Herein, we assessed the effect of the Bifidobacterium species Bifidobacterium bifidum, Bifidobacterium longum, Bembidion breve, Bifidobacterium animalis alone, and also a Bifidobacterium consortium on the digestion of intact gluten proteins (gliadins and glutenins) and the associated immunomodulatory responses elicited by the resulting peptides. The cytotoxicity and proinflammatory responses were evaluated through the activation of NF-kB p65 and the expression of cytokines TNF-α and IL-1ß in Caco-2 cell cultures exposed to gluten-derived peptides. The peptides induced a clear reduction in cytotoxic responses and proinflammatory marker levels compared to the gluten fragments generated during noninoculated gastrointestinal digestion. These results highlight the possible use of probiotics based on bifidobacteria as a prospective treatment for CD.
Assuntos
Bifidobacterium/metabolismo , Gliadina/metabolismo , Glutens/metabolismo , Biotransformação , Células CACO-2 , Doença Celíaca/tratamento farmacológico , Doença Celíaca/genética , Doença Celíaca/imunologia , Gliadina/química , Gliadina/imunologia , Glutens/imunologia , Humanos , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Probióticos/administração & dosagem , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologiaRESUMO
The gut microbiota plays a relevant role in determining an individual's health status, and the diet is a major factor in modulating the composition and function of gut microbiota. Gluten constitutes an essential dietary component in Western societies and is the environmental trigger of celiac disease. The presence/absence of gluten in the diet can change the diversity and proportions of the microbial communities constituting the gut microbiota. There is an intimate relation between gluten metabolism and celiac disease pathophysiology and gut microbiota; their interrelation defines intestinal health and homeostasis. Environmental factors modify the intestinal microbiota and, in turn, its changes modulate the mucosal and immune responses. Current evidence from studies of young and adult patients with celiac disease increasingly supports that dysbiosis (i.e., compositional and functional alterations of the gut microbiome) is present in celiac disease, but to what extent this is a cause or consequence of the disease and whether the different intestinal diseases (celiac disease, ulcerative colitis, Crohn disease) have specific change patterns is not yet clear. The use of bacterial-origin enzymes that help completion of gluten digestion is of interest because of the potential application as coadjuvant in the current treatment of celiac disease. In this narrative review, we address the current knowledge on the complex interaction between gluten digestion and metabolism, celiac disease, and the intestinal microbiota.
Assuntos
Bactérias/crescimento & desenvolvimento , Doença Celíaca/microbiologia , Dieta , Disbiose , Microbioma Gastrointestinal , Glutens , Intestinos/microbiologia , Proteínas de Bactérias/uso terapêutico , Doença Celíaca/tratamento farmacológico , Dieta Livre de Glúten , Digestão , Disbiose/complicações , Disbiose/microbiologia , Glutens/metabolismo , Glutens/farmacologia , Humanos , Mucosa Intestinal/microbiologia , Probióticos/uso terapêuticoRESUMO
The callitrichids are non-human primates that feed on insects and plant matter in nature, but in captivity, they are fed mostly an artificial diet containing amounts of gluten, in their toxic forms in items such as wheat, barley and rye. The aim of this research was to estimate the blood ß-defensin and Toll like receptor 5 (TLR5) gene expressions and to analyze the stool consistency (firm, soft, diarrheic) in Leontocebus fuscicollis raised in captivity. Blood samples of animals under gluten-free and gluten diets were collected and their fecal output quality was periodically monitored and classified during the course of the study. Gene expression was evaluated using real-time PCR. The stool consistencies of individuals fed a gluten diet were most frequently soft or diarrheic, while it was mostly normal in individuals fed a gluten-free diet. ß-Defensin expression increased in individuals fed a gluten diet, but decreased after 15 days. Expression normalized between 30 and 45 days on a gluten-free diet. However, expression of the TLR5 gene did not change under a gluten diet. A gluten diet affects stool quality, and brings about an immediate increase in blood ß-defensin expression in the beginning but decreases after 15 days.
Assuntos
Dieta Livre de Glúten , Expressão Gênica/imunologia , Glutens/metabolismo , Animais , Callitrichinae , Diarreia , Fezes , Imunidade Inata , Inflamação , Receptor 5 Toll-Like/sangue , beta-Defensinas/sangueRESUMO
BACKGROUND: Life-long removal of gluten from the diet is currently the only way to manage celiac disease (CeD). Until now, no objective test has proven useful to objectively detect ingested gluten in clinical practice. Recently, tests that determine consumption of gluten by assessing excretion of gluten immunogenic peptides (GIP) in stool and urine have been developed. Their utility, in comparison with conventional dietary and analytical follow-up strategies, has not been fully established. AIM: To assess the performance of enzyme-linked immunosorbent assay (ELISA) and point-of-care tests (PoCTs) for GIP excretion in CeD patients on gluten-free diet (GFD). METHODS: We conducted an observational, prospective, cross-sectional study in patients following a GFD for at least two years. Using the Gastrointestinal Symptom Rating Scale questionnaire, patients were classified at enrollment as asymptomatic or symptomatic. Gluten consumption was assessed twice by 3-d dietary recall and GIP excretion (by ELISA in stool and PoCTs (commercial kits for stool and urine) in two consecutive samples. These samples and dietary reports were obtained 10 day apart one from the other. Patients were encouraged to follow their usual GFD during the study period. RESULTS: Forty-four patients were enrolled, of which 19 (43.2%) were symptomatic despite being on a GFD. Overall, 83 sets of stool and/or urine samples were collected. Eleven out of 44 patients (25.0%) had at least one positive GIP test. The occurrence of at least one positive test was 32% in asymptomatic patients compared with 15.8% in symptomatic patients. GIP was concordant with dietary reports in 65.9% of cases (Cohen´s kappa: 0.317). PoCT detected dietary indiscretions. Both ELISA and PoCT in stool were concordant (concomitantly positive or negative) in 67 out of 74 (90.5%) samples. Excretion of GIP was detected in 7 (8.4%) stool and/or urine samples from patients considered to be strictly compliant with the GFD by dietary reports. CONCLUSION: GIP detects dietary transgressions in patients on long-term GFD, irrespective of the presence of symptoms. PoCT for GIP detection constitutes a simple home-based method for self-assessment of dietary indiscretions.
Assuntos
Doença Celíaca/dietoterapia , Dieta Livre de Glúten , Glutens/análise , Cooperação do Paciente , Peptídeos/análise , Adulto , Idoso , Idoso de 80 Anos ou mais , Doenças Assintomáticas , Doença Celíaca/diagnóstico , Doença Celíaca/imunologia , Doença Celíaca/urina , Estudos Transversais , Autoavaliação Diagnóstica , Ensaio de Imunoadsorção Enzimática , Fezes/química , Feminino , Glutens/química , Glutens/imunologia , Glutens/metabolismo , Humanos , Eliminação Intestinal , Masculino , Pessoa de Meia-Idade , Peptídeos/química , Peptídeos/imunologia , Peptídeos/metabolismo , Testes Imediatos , Estudos Prospectivos , Inquéritos e QuestionáriosRESUMO
Germination in the presence of selenium (Se) is an alternative to increase the healthy properties of seeds. This study aimed to compare the Se accumulation in different protein fractions from germinated chickpea (Cicer arietinum L.) and the effect on digestibility and cellular antioxidant activity (CAA) of protein hydrolysates. Chickpeas were germinated during four days after soaking with sodium selenite (0, 1, or 2â¯mg/100â¯g seeds). Total protein (TP) and glutelin (Glu), albumin (Alb) and globulin (Glo) fractions were digested and ultrafiltrated through a 10â¯kDa membrane. Se accumulated in the order of Gluâ¯>â¯Albâ¯>â¯Glo. Ultrafiltrated Glu hydrolysate of four days germinated chickpeas treated with 2â¯mg Na2SeO3/100â¯g increased CAA (51.47%), demonstrating the potential health benefits of selenization. The intensity of vicilin bands (34-37â¯kDa) increased from the second to the fourth day compared with the control samples. Glo digestibility was higher in selenized chickpea sprouts.