Esther Nistal

Differences of small intestinal bacteria populations in adults and children with/without celiac disease: Effect of age, gluten diet, and disease

Background: Scientific evidence has revealed microecological changes in the intestinal tract of celiac infants. The objective of this work is the study of bacterial differences in the upper small intestine in both adults (healthy, untreated celiac disease [CD], and CD treated with a gluten‐free diet) and children (healthy and untreated CD). Methods: Intestinal bacterial communities were identified by 16S rRNA gene sequencing of DNA extracted from duodenal biopsies. Results: Analysis of the sequences from adults and children showed that this niche was colonized by bacteria affiliated mainly with three phyla: Firmicutes, Proteobacteria, and Bacteroidetes. In total, 89 different genera were identified in adults and 46 in children. Bacterial richness was significantly lower in the children than in the adults. A global principal component analysis of the bacterial communities of both healthy and untreated CD patient groups (including both children and adults) revealed a strong effect of age in principal component 1—clustering all adults and children separately—and a possible effect of the disease in adults with untreated patients clustering separately. Conclusions: There are bacterial differences in the upper small intestine between untreated children CD patients and untreated CD adults due to age. There are bacterial differences in the upper small bacteria microbiota between treated and untreated CD adults due to treatment with a gluten‐free diet. (Inflamm Bowel Dis 2011;)

Monitoring of gluten-free diet compliance in celiac patients by assessment of gliadin 33-mer equivalent epitopes in feces

Background: Certain immunotoxic peptides from gluten are resistant to gastrointestinal digestion and can interact with celiac-patient factors to trigger an immunologic response. A gluten-free diet (GFD) is the only effective treatment for celiac disease (CD), and its compliance should be monitored to avoid cumulative damage. However, practical methods to monitor diet compliance and to detect the origin of an outbreak of celiac clinical symptoms are not available. Objective: We assessed the capacity to determine the gluten ingestion and monitor GFD compliance in celiac patients by the detection of gluten and gliadin 33-mer equivalent peptidic epitopes (33EPs) in human feces. Design: Fecal samples were obtained from healthy subjects, celiac patients, and subjects with other intestinal pathologies with different diet conditions. Gluten and 33EPs were analyzed by using immunochromatography and competitive ELISA with a highly sensitive antigliadin 33-mer monoclonal antibody. Results: The resistance of a significant part of 33EPs to gastrointestinal digestion was shown in vitro and in vivo. We were able to detect gluten peptides in feces of healthy individuals after consumption of a normal gluten-containing diet, after consumption of a GFD combined with controlled ingestion of a fixed amount of gluten, and after ingestion of <100 mg gluten/d. These methods also allowed us to detect GFD infringement in CD patients. Conclusions: Gluten-derived peptides could be sensitively detected in human feces in positive correlation with the amount of gluten intake. These techniques may serve to show GFD compliance or infringement and be used in clinical research in strategies to eliminate gluten immunotoxic peptides during digestion. This trial was registered at clinicaltrials.gov as NCT01478867.

Diversity of the cultivable human gut microbiome involved in gluten metabolism: isolation of microorganisms with potential interest for coeliac disease

Gluten, a common component in the human diet, is capable of triggering coeliac disease pathogenesis in genetically predisposed individuals. Although the function of human digestive proteases in gluten proteins is quite well known, the role of intestinal microbiota in the metabolism of proteins is frequently underestimated. The aim of this study was the isolation and characterisation of the human gut bacteria involved in the metabolism of gluten proteins. Twenty-two human faecal samples were cultured with gluten as the principal nitrogen source, and 144 strains belonging to 35 bacterial species that may be involved in gluten metabolism in the human gut were isolated. Interestingly, 94 strains were able to metabolise gluten, 61 strains showed an extracellular proteolytic activity against gluten proteins, and several strains showed a peptidasic activity towards the 33-mer peptide, an immunogenic peptide in patients with coeliac disease. Most of the strains were classified within the phyla Firmicutes and Actinobacteria, mainly from the genera Lactobacillus, Streptococcus, Staphylococcus, Clostridium and Bifidobacterium. In conclusion, the human intestine exhibits a large variety of bacteria capable of utilising gluten proteins and peptides as nutrients. These bacteria could have an important role in gluten metabolism and could offer promising new treatment modalities for coeliac disease.

Protective effect of quercetin on high-fat diet-induced non-alcoholic fatty liver disease in mice is mediated by modulating intestinal microbiota imbalance and related gut-liver axis activation

Abstract Gut microbiota is involved in obesity, metabolic syndrome and the progression of nonalcoholic fatty liver disease (NAFLD). It has been recently suggested that the flavonoid quercetin may have the ability to modulate the intestinal microbiota composition, suggesting a prebiotic capacity which highlights a great therapeutic potential in NAFLD. The present study aims to investigate benefits of experimental treatment with quercetin on gut microbial balance and related gut‐liver axis activation in a nutritional animal model of NAFLD associated to obesity. C57BL/6J mice were challenged with high fat diet (HFD) supplemented or not with quercetin for 16 weeks. HFD induced obesity, metabolic syndrome and the development of hepatic steatosis as main hepatic histological finding. Increased accumulation of intrahepatic lipids was associated with altered gene expression related to lipid metabolism, as a result of deregulation of their major modulators. Quercetin supplementation decreased insulin resistance and NAFLD activity score, by reducing the intrahepatic lipid accumulation through its ability to modulate lipid metabolism gene expression, cytochrome P450 2E1 (CYP2E1)‐dependent lipoperoxidation and related lipotoxicity. Microbiota composition was determined via 16S ribosomal RNA Illumina next‐generation sequencing. Metagenomic studies revealed HFD‐dependent differences at phylum, class and genus levels leading to dysbiosis, characterized by an increase in Firmicutes/Bacteroidetes ratio and in Gram‐negative bacteria, and a dramatically increased detection of Helicobacter genus. Dysbiosis was accompanied by endotoxemia, intestinal barrier dysfunction and gut‐liver axis alteration and subsequent inflammatory gene overexpression. Dysbiosis‐mediated toll‐like receptor 4 (TLR‐4)‐NF‐&kgr;B signaling pathway activation was associated with inflammasome initiation response and reticulum stress pathway induction. Quercetin reverted gut microbiota imbalance and related endotoxemia‐mediated TLR‐4 pathway induction, with subsequent inhibition of inflammasome response and reticulum stress pathway activation, leading to the blockage of lipid metabolism gene expression deregulation. Our results support the suitability of quercetin as a therapeutic approach for obesity‐associated NAFLD via its anti‐inflammatory, antioxidant and prebiotic integrative response. HighlightsDysbiosis is accompanied by gut‐liver axis alteration in HFD‐induced NAFLD.Quercetin prevents dysbiosis‐induced TLR4‐mediated inflammation and lipotoxicity.Quercetin counteracts inflammasome and reticulum stress pathway activation.Modulatory effects displayed by quercetin counteract lipid metabolism deregulation.Quercetin improves NAFLD via an integrative response including its prebiotic effect.

Factors Determining Colorectal Cancer: The Role of the Intestinal Microbiota.

The gastrointestinal tract, in particular the colon, holds a complex community of microorganisms, which are essential for maintaining homeostasis. However, in recent years, many studies have implicated microbiota in the development of colorectal cancer (CRC), with this disease considered a major cause of death in the western world. The mechanisms underlying bacterial contribution in its development are complex and are not yet fully understood. However, there is increasing evidence showing a connection between intestinal microbiota and CRC. Intestinal microorganisms cause the onset and progression of CRC using different mechanisms, such as the induction of a chronic inflammation state, the biosynthesis of genotoxins that interfere with cell cycle regulation, the production of toxic metabolites, or heterocyclic amine activation of pro-diet carcinogenic compounds. Despite these advances, additional studies in humans and animal models will further decipher the relationship between microbiota and CRC, and aid in developing alternate therapies based on microbiota manipulation.

Study of duodenal bacterial communities by 16S rRNA gene analysis in adults with active celiac disease vs non-celiac disease controls.

Several studies have suggested that abnormalities in the small‐intestinal microbiota might be involved in the development or the pathogenesis of celiac disease (CD). The objective of this study was to characterize and compare the composition of the duodenal microbiota between CD patients and non‐CD controls.

Differences in gluten metabolism among healthy volunteers, coeliac disease patients and first-degree relatives.

Coeliac disease (CD) is an immune-mediated enteropathy resulting from exposure to gluten in genetically predisposed individuals. Gluten proteins are partially digested by human proteases generating immunogenic peptides that cause inflammation in patients carrying HLA-DQ2 and DQ8 genes. Although intestinal dysbiosis has been associated with patients with CD, bacterial metabolism of gluten has not been studied in depth thus far. The aim of this study was to analyse the metabolic activity of intestinal bacteria associated with gluten intake in healthy individuals, CD patients and first-degree relatives of CD patients. Faecal samples belonging to twenty-two untreated CD patients, twenty treated CD patients, sixteen healthy volunteers on normal diet, eleven healthy volunteers on gluten-free diet (GFD), seventy-one relatives of CD patients on normal diet and sixty-nine relatives on GFD were tested for several proteolytic activities, cultivable bacteria involved in gluten metabolism, SCFA and the amount of gluten in faeces. We detected faecal peptidasic activity against the gluten-derived peptide 33-mer. CD patients showed differences in faecal glutenasic activity (FGA), faecal tryptic activity (FTA), SCFA and faecal gluten content with respect to healthy volunteers. Alterations in specific bacterial groups metabolising gluten such as Clostridium or Lactobacillus were reported in CD patients. Relatives showed similar parameters to CD patients (SCFA) and healthy volunteers (FTA and FGA). Our data support the fact that commensal microbial activity is an important factor in the metabolism of gluten proteins and that this activity is altered in CD patients.

Gluten Metabolism in Humans: Involvement of the Gut Microbiota

Abstract Gluten proteins are the major storage proteins that are deposited in the starchy endosperm cells of developing wheat grain. These proteins have the capacity to form a viscoelastic network, and thus wheat is used in numerous processed foods. Therefore, a large amount of gluten protein is ingested by humans. However, because of their high proline and glutamine content, gluten peptides are relatively resistant to complete digestion by human digestive proteases because those enzymes are deficient in prolyl endopeptidasic activity. The incomplete digestion of gluten proteins generates high molecular weight oligopeptides that remain in the lumen of the small intestine; some of these are capable of triggering the inflammatory process associated with celiac disease (CD). Nevertheless, there are several reasons why gut microbiota should be taken into account when considering the metabolism of proteins in the human intestine. For example, there are bacteria in the oral cavity that have the ability to hydrolyze gluten peptides, and there are bacteria in the large intestine with the ability to digest gliadin peptides. These bacteria could generate different digestion processes for gluten proteins in CD patients and in healthy people. Therefore, this review examines gluten metabolism throughout the gastrointestinal tract, and the role of the gut microbiota in this process.Gluten proteins are the major storage proteins that are deposited in the starchy endosperm cells of developing wheat grain. These proteins have the capacity to form a viscoelastic network, and thus wheat is used in numerous processed foods. Therefore, a large amount of gluten protein is ingested by humans. However, because of their high proline and glutamine content, gluten peptides are relatively resistant to complete digestion by human digestive proteases because those enzymes are deficient in prolyl endopeptidasic activity. The incomplete digestion of gluten proteins generates high molecular weight oligopeptides that remain in the lumen of the small intestine; some of these are capable of triggering the inflammatory process associated with celiac disease (CD). Nevertheless, there are several reasons why gut microbiota should be taken into account when considering the metabolism of proteins in the human intestine. For example, there are bacteria in the oral cavity that have the ability to hydrolyze gluten peptides, and there are bacteria in the large intestine with the ability to digest gliadin peptides. These bacteria could generate different digestion processes for gluten proteins in CD patients and in healthy people. Therefore, this review examines gluten metabolism throughout the gastrointestinal tract, and the role of the gut microbiota in this process.