The gut microbiota has been implicated in onset and progression of ulcerative colitis (UC). Here, we assess potential causal involvement of the microbiota and -associated fecal water (FW) metabolome in altering key functional parameters of the colonic epithelium. Fecal samples were collected from N = 51 healthy controls (HC), N = 36 patients with active UC (UC-A), and N = 41 subjects in remission N = 41 (UC-R). Using in vitro incubation experiments, the FW metabolome's impact on butyrate oxidation rates/gene expression and cell death (cytotoxicity) of HT-29 cells, cytokine production by PBMC, and barrier integrity of Caco2 monolayers was evaluated. The FW metabolome from patients and individuals hosting the Bacteroides 2 (Bact2) enterotype (69% of UC-A, 31% of UC-R, 3% of HC), characterized by lower levels of median- and short-chain fatty acids and furan compounds, left butyrate oxidation rates unaltered but affected associated gene expression profiles. UC patients/Bact2-carriers' FW lowered PBMC IL-8 production and increased IL-1 beta production. Patients' FW increased cytotoxicity, associated with sulfide compound levels. Bact2 carriers' FW, displaying higher levels of bile acids, lowered barrier function upon incubation of monolayers. The FW metabolome of patients and individuals hosting a dysbiotic microbiota could contribute to the disruption of functional processes of the colonic epithelium as observed in UC.

Background: The breast-milk composition in the first 6 wk postpartum of women who have undergone bariatric surgery (BS) is unknown. Objective: The aim of this study was to examine 1) the breast-milk macronutrient and vitamin A composition in women who had and who had not undergone BS and 2) the impact of maternal diet on the breast-milk composition. We hypothesized that the milk of women who had undergone BS would be less energy dense and have a lower vitamin A concentration than that of other women. Methods: A multicenter prospective substudy was conducted at 2 university hospitals. Breast-milk samples were collected from 24 normal-weight [NW; mean +/- SD body mass index (BMI; kg/m(2)): 21.5 +/- 1.7; mean +/- SD age: 29 +/- 6 y], 39 overweight (OW; BMI: 26.9 +/- 1.5; aged 29 +/- 5 y), and 12 obese women (BMI: 35.0 +/- 5.7; aged 29 +/- 5 y) as well as from 11 women who had undergone BS (BMI: 28.0 +/- 4.4; aged 30 +/- 4 y) from day 3 until week 6 of lactation. Milk energy and macronutrients (Human Milk Analyzer; Miris) and vitamin A concentrations (iCheck Fluoro; BioAnalyt) were determined at the end of each week. Maternal diet (food-frequency questionnaire) and physical activity (Kaiser Physical Activity Survey) were measured during the third trimester of pregnancy and on day 3 or 4 and during week 6 of lactation. Statistical analyses include 1-factor ANOVA, Spearman and Pearson correlations, and multiple linear regression. Results: In all women, a weekly increase in milk energy, total fat, and total carbohydrates was seen, whereas a weekly decrease in proteins and vitamin A was found during the first 2 wk of lactation, followed by a stable concentration of all nutrients. At week 4, milk protein concentrations were higher in women who had undergone BS (14 g/L) compared with NW (8 g/L; P = 0.005) and OW (9 g/L; P = 0.019) women. At week 5, milk carbohydrate concentrations were higher in women who had undergone BS (74 g/L) compared with NW women (68 g/L; P = 0.042). Conclusions: Breast milk of women who have undergone BS appears to be adequate in energy, macronutrients, and vitamin A during the first 6 wk of lactation. This supports the conclusion that breast feeding should not be discouraged in this group of women.

Key points The short‐chain fatty acids (SCFAs) are bacterial metabolites produced during the colonic fermentation of undigested carbohydrates, such as dietary fibre and prebiotics, and can mediate the interaction between the diet, the microbiota and the host. We quantified the fraction of colonic administered SCFAs that could be recovered in the systemic circulation, the fraction that was excreted via the breath and urine, and the fraction that was used as a precursor for glucose, cholesterol and fatty acids. This information is essential for understanding the molecular mechanisms by which SCFAs beneficially affect physiological functions such as glucose and lipid metabolism and immune function. Abstract The short‐chain fatty acids (SCFAs), acetate, propionate and butyrate, are bacterial metabolites that mediate the interaction between the diet, the microbiota and the host. In the present study, the systemic availability of SCFAs and their incorporation into biologically relevant molecules was quantified. Known amounts of 13 C‐labelled acetate, propionate and butyrate were introduced in the colon of 12 healthy subjects using colon delivery capsules and plasma levels of 13 C‐SCFAs 13 C‐glucose, 13 C‐cholesterol and 13 C‐fatty acids were measured. The butyrate‐producing capacity of the intestinal microbiota was also quantified. Systemic availability of colonic‐administered acetate, propionate and butyrate was 36%, 9% and 2%, respectively. Conversion of acetate into butyrate (24%) was the most prevalent interconversion by the colonic microbiota and was not related to the butyrate‐producing capacity in the faecal samples. Less than 1% of administered acetate was incorporated into cholesterol and <15% in fatty acids. On average, 6% of colonic propionate was incorporated into glucose. The SCFAs were mainly excreted via the lungs after oxidation to 13 CO 2 , whereas less than 0.05% of the SCFAs were excreted into urine. These results will allow future evaluation and quantification of SCFA production from 13 C‐labelled fibres in the human colon by measurement of 13 C‐labelled SCFA concentrations in blood.

There is increasing interest in the colonic microbiota as a relevant source of uremic retention solutes accumulating in CKD. Renal disease can also profoundly affect the colonic microenvironment and has been associated with a distinct colonic microbial composition. However, the influence of CKD on the colonic microbial metabolism is largely unknown. Therefore, we studied fecal metabolite profiles of hemodialysis patients and healthy controls using a gas chromatography-mass spectrometry method. We observed a clear discrimination between both groups, with 81 fecal volatile organic compounds detected at significantly different levels in hemodialysis patients and healthy controls. To further explore the differential impact of renal function loss per se versus the effect of dietary and other CKD-related factors, we also compared fecal metabolite profiles between patients on hemodialysis and household contacts on the same diet, which revealed a close resemblance. In contrast, significant differences were noted between the fecal samples of rats 6 weeks after 5/6th nephrectomy and those of sham-operated rats, still suggesting an independent influence of renal function loss. Thus, CKD associates with a distinct colonic microbial metabolism, although the effect of renal function loss per se in humans may be inferior to the effects of dietary and other CKD-related factors. The potential beneficial effect of therapeutics targeting colonic microbiota in patients with CKD remains to be examined.

Background: Crohn's disease and ulcerative colitis represent the 2 major phenotypes of inflammatory bowel disease (IBD) that are characterized by chronic inflammation of all or parts of the gastrointestinal tract. The pathogenesis of both diseases is influenced by genetic predispositions as well as microbial and environmental factors. Currently, there is an emerging consensus hypothesis that a microbial dysbiosis is involved in initiating the disease or in maintaining it. These compositional alterations may be reflected in altered metabolic activities of the gut microbiota and has led to the use of ‘omic' profiling to improve the understanding of the pathophysiology of IBD. Key Messages: In the past few years, a metabolic approach has increasingly been applied in a number of studies of experimental and human IBD which were mostly focused on exploring disease-related metabolites to gain more insight into metabolic pathways. Metabolomics involves the high throughput identification, characterization and quantification of small molecule metabolites by different analytical techniques and has been performed in different biofluids such as serum/plasma, urine or fecal samples. The application of such a metabolite profiling technique has revealed different metabolites that allow the discrimination of IBD patients from healthy controls. In addition, separate IBD subtypes could be differentiated. Some of these metabolic changes were directly associated to alterations of specific gut microbial populations, implying a perturbation in the gut microbiome in the development of IBD. Conclusions: This review covers the emerging contribution of metabolomics for the discovery of an IBD signature and to identify biomarkers linked with a metabolic imbalance. For the implementation of metabolomics as a diagnostic tool in IBD, large prospective cohort studies are necessary.

Background: Many studies have focused on the fecal microbiota however it is now widely accepted that the gastrointestinal (GI) mucosa associated microbiota, rather than luminal content, is critically important in host-microbe interactions linked to health and disease.Mucosal biopsy is the most common sampling technique used to assess the mucosa associated microbiota.However, normal biopsy devices are designed to take samples for histologic assessment.Thus when biopsies are taken via working channels that are also used to aspirate luminal content it is highly likely that cross contamination occurs, that ultimately questions the validity of these samples.In view of this difficulty we have developed a novel device that allows targeted biopsies to be taken in any segment of the GI tract without cross contamination in an aseptic manner.Methods: Six patients undergoing upper GI endoscopy for iron deficiency were recruited with consent and ethical approval.In pilot experiments various prototypes of a sampling device were tested in vivo.Two final configurations of a sheath that covered miniature biopsy forceps were tested and compared with the normal single-use biopsy forceps (18 samples).Matched duodenal biopsy samples were collected from each patient, transferred individually into RNA later, and subject to gDNA extraction.Amplicon libraries spanning the V6-V8 region of the 16S rRNA gene were constructed, sequenced using the Illumina MiSeq platform, and analysed via the QIIME pipeline.Results: Microbial DNA, representing a diverse community, was observed in all duodenal samples obtained using the aseptic device, indicating this device is effective in sampling mucosa associated organisms present in the duodenum (Chao 1 estimated richness = 386, Good's coverage 99.8%).Assessment of the duodenal microbiota revealed a community dominated by the genera Streptococcus, Prevotella, Veillonella, Neisseria and Porphyromonas.There were substantial differences in the microbiota of samples obtained using the aseptic device and samples obtained with standard biopsy forceps.Only a low level of correlation (Pearson's r<0.6) between the matched biopsy pairs was seen across all bacterial phyla observed.Phylogenetic based (UniFrac) evaluation of beta-diversity revealed the aseptic samples clustered together and showed reduced inter-individual variability compared to those obtained with standard forceps.Conclusions: Sampling of duodenal biopsies with routine biopsy forceps resulted in greater microbial diversity as compared to samples acquired under aseptic conditions.This suggests cross contamination from other sites of the upper GI tract when utilising standard forceps.Based upon our data, aseptic techniques with a specific sampling device should be used to collect samples for studies that aim to define the mucosa associated microbiota.

Background: Many studies have focused on the fecal microbiota however it is now widely accepted that the gastrointestinal (GI) mucosa associated microbiota, rather than luminal content, is critically important in host-microbe interactions linked to health and disease. Mucosal biopsy is the most common sampling technique used to assess the mucosa associated microbiota. However, normal biopsy devices are designed to take samples for histologic assessment. Thus when biopsies are taken via working channels that are also used to aspirate luminal content it is highly likely that cross contamination occurs, that ultimately questions the validity of these samples. In view of this difficulty we have developed a novel device that allows targeted biopsies to be taken in any segment of the GI tract without cross contamination in an aseptic manner. Methods: Six patients undergoing upper GI endoscopy for iron deficiency were recruited with consent and ethical approval. In pilot experiments various prototypes of a sampling device were tested in vivo. Two final configurations of a sheath that covered miniature biopsy forceps were tested and compared with the normal single-use biopsy forceps (18 samples). Matched duodenal biopsy samples were collected from each patient, transferred individually into RNA later, and subject to gDNA extraction. Amplicon libraries spanning the V6-V8 region of the 16S rRNA gene were constructed, sequenced using the Illumina MiSeq platform, and analysed via the QIIME pipeline. Results: Microbial DNA, representing a diverse community, was observed in all duodenal samples obtained using the aseptic device, indicating this device is effective in sampling mucosa associated organisms present in the duodenum (Chao 1 estimated richness = 386, Good's coverage 99.8%). Assessment of the duodenal microbiota revealed a community dominated by the genera Streptococcus, Prevotella, Veillonella, Neisseria and Porphyromonas. There were substantial differences in the microbiota of samples obtained using the aseptic device and samples obtained with standard biopsy forceps. Only a low level of correlation (Pearson's r<0.6) between the matched biopsy pairs was seen across all bacterial phyla observed. Phylogenetic based (UniFrac) evaluation of beta-diversity revealed the aseptic samples clustered together and showed reduced inter-individual variability compared to those obtained with standard forceps. Conclusions: Sampling of duodenal biopsies with routine biopsy forceps resulted in greater microbial diversity as compared to samples acquired under aseptic conditions. This suggests cross contamination from other sites of the upper GI tract when utilising standard forceps. Based upon our data, aseptic techniques with a specific sampling device should be used to collect samples for studies that aim to define the mucosa associated microbiota.

The lactose hydrogen breath test is a commonly used, non-invasive method for the detection of lactose malabsorption and is based on an abnormal increase in breath hydrogen (H2) excretion after an oral dose of lactose. We use a combined 13C/H2 lactose breath test that measures breath 13CO2 as a measure of lactose digestion in addition to H2 and that has a better sensitivity and specificity than the standard test. The present retrospective study evaluated the results of 1051 13C/H2 lactose breath tests to assess the impact on the diagnostic accuracy of measuring breath CH4 in addition to H2 and 13CO2. Based on the 13C/H2 breath test, 314 patients were diagnosed with lactase deficiency, 138 with lactose malabsorption or small bowel bacterial overgrowth (SIBO), and 599 with normal lactose digestion. Additional measurement of CH4 further improved the accuracy of the test as 16% subjects with normal lactose digestion and no H2-excretion were found to excrete CH4. These subjects should have been classified as subjects with lactose malabsorption or SIBO. In conclusion, measuring CH4-concentrations has an added value to the 13C/H2 breath test to identify methanogenic subjects with lactose malabsorption or SIBO.

the smooth muscle.Rhythmic calcium transients occuring simultaneously in many Dogiel Type I and II neurons, across many rows of myenteric ganglia, were found to cause timelocked calcium action potentials arising from EJPs in neigbouring smooth muscle layers, that lead to muscle contraction underlying each CMMC.All synchronized calcium transients across multiple ganglia were abolished by hexamethonium (300 micromolar; N=6) or tetrodotoxin (1 micromolar; N=4).This is the first demonstration of a rhythmic neuronal firing pattern in mammalian ENS that is responsible for rhythmic cholinergic EJPs in the smooth muscle during a complex intestinal motor pattern.Of particular interest was the discovery that both Dogiel Type II (intrinsic sensory neurons) and Type I neurons (interneurons and motor neurons) generated simultaneous and rhythmic calcium transients during each CMMC, over large spatial fields of ENS.

Owing to the recent developments in analytical techniques to analyse the composition of complex microbial ecosystems, our understanding of the intestinal microbiota has tremendously increased. Several disorders have been associated with an altered composition of the gut bacteria. As a consequence, the micro biota is increasingly recognized as a therapeutic target to improve health. Besides having a trophic and protective function, the microbiota is a metabolically very active ecosystem. Amongst the wide variety of metabolites produced, short-chain fatty acids (SCFA) constitute the most relevant compounds in relation to health. As far as we know to date, administration of prebiotics selectively modifies the composition of the intestinal microbiota through several mechanisms and favours the saccharolytic fermentation resulting in increased SCFA production. These SCFA play a pivotal role in the health benefits associated with prebiotic intake as they acidify the colonic lumen, which influences metabolic pathways and inhibits pathogens, and act as signalling molecules on specific receptors. In the future, more detailed information on the exact role of each individual SCFA and on the proportion of the SCFA produced from different prebiotic substrates will be essential to further exploit the benefits of prebiotic use.

Recently, low-residue diets were removed from the American Academy of Nutrition and Dietetics' Nutrition Care Manual due to the lack of a scientifically accepted quantitative definition and the unavailability of a method to estimate the amount of food residue produced. This narrative review focuses on defining the similarities and/or discrepancies between low-residue and low-fiber diets and on the diagnostic and therapeutic values of these diets in gastrointestinal disease management. Diagnostically, a low-fiber/low-residue diet is used in bowel preparation. A bowel preparation is a cleansing of the intestines of fecal matter and secretions conducted before a diagnostic procedure. Therapeutically, a low-fiber/low-residue diet is part of the treatment of acute relapses in different bowel diseases. The available evidence on low-residue and low-fiber diets is summarized. The main findings showed that within human disease research, the terms "low residue" and "low fiber" are used interchangeably, and information related to the quantity of residue in the diet usually refers to the amount of fiber. Low-fiber/low-residue diets are further explored in both diagnostic and therapeutic situations. On the basis of this literature review, the authors suggest redefining a low-residue diet as a low-fiber diet and to quantitatively define a low-fiber diet as a diet with a maximum of 10 g fiber/d. A low-fiber diet instead of a low-residue diet is recommended as a diagnostic value or as specific therapy for gastrointestinal conditions.

Systems biology represents an integrative research strategy that studies the interactions between DNA, mRNA, protein, and metabolite level in an organism, thereby including the interactions with the physical environment and other organisms. The application of metabonomics, or the quantitative study of metabolites in biological systems, in systems biology is currently an emerging area of research, which can contribute to the discovery of (disease) signatures, drug targeting and design, and the further elucidation of basic and more complex biochemical principles. This chapter covers the contribution of metabonomics in advancing our understanding in systems biology.

In ulcerative colitis (UC) the butyrate metabolism is impaired, leading to energy‐deficiency in the colonic cells. The effect of inflammation on the butyrate metabolism was investigated. HT‐29 cells were incubated with pro‐inflammatory cytokines (TNF‐α and/or IFN‐γ) for 1 and 24 h. Cells were additionally stimulated with butyrate to investigate its anti‐inflammatory potential. Butyrate uptake and oxidation were measured using 14C‐labeled butyrate. Gene expression of the butyrate metabolism enzymes, interleukin 8 (IL‐8; inflammatory marker) and villin‐1 (VIL‐1; epithelial cell damage marker) was measured via quantitative RT‐PCR. Significantly increased IL‐8 expression and decreased VIL‐1 expression after 24 h incubation with TNF‐α and/or IFN‐γ confirmed the presence of inflammation. These conditions induced a decrease of both butyrate uptake and oxidation, whereas the gene expression was not reduced. Simultaneous incubation with butyrate counteracted the reduced butyrate oxidation. In contrast, 1 h incubation with TNF‐α induced a significant increased IL‐8 expression and decreased butyrate uptake. Incubation with TNF‐α and/or IFN‐γ for 1 h did not induce cell damage nor influence butyrate oxidation. The inflammation‐induced downregulation of the butyrate metabolism was not caused by a reduced gene expression, but appeared consequential to a decreased butyrate uptake. Increasing the luminal butyrate levels might have therapeutic potential in UC. J. Cell. Physiol. 230: 418–426, 2015. © 2014 Wiley Periodicals, Inc.

Introduction and Aims: Although it is reported that gut microbiota and colon barrier function were deteriorated in CKD state, the detailed mechanism or pathological relevance has not been elucidated. Oral activated charcoal adsorbent (AST-120) has been reported to delay the progression of CKD by adsorbing uremic toxin from the intestine. However, the effects of AST on the gut environments in CKD and its impact on CKD progression have not been fully elucidated. Methods: Six-week-old spontaneously hypertensive rats (SHR) were rendered CKD by 5/6 nephrectomy (Nx). The SHRs were divided into four groups; group 1, sham-operated SHR (SHR, n=10); group 2, SHR given AST-120 (SHR+AST, n=10); group 3, 5/6 nephrectomized SHR (Nx, n=10); group 4, 5/6 nephrectomized SHR given AST-120 (Nx+AST, n=10). AST-120 was orally given at a dose of 4g/kg/day by mixing with regular chow. After 12 weeks, rats were sacrificed and biochemical parameters, histological changes in the kidney and colon, and molecular changes were compared among the groups. The distribution of the intestinal flora was examined by T-RFLP (Terminal Restriction Fragment Length Polymorphism) and real time PCR. Human colon cell line, Caco-2 cell were treated with uremic toxin precursor, indole. Results: Serum level of indoxylsulfate (IS), one of the major uremic toxin was increased in Nx, which was significantly reduced in Nx+AST. Serum creatinine levels and urinary protein excretion were increased in Nx, which were attenuated in Nx+AST. Glomerular sclerosis and tissue fibrosis evaluated by Masson-trichome staining were increased in Nx, which were also attenuated in Nx+AST. The histological analysis of the colon tissue revealed that the number of goblet cell and the protein expression of mucin-2 were decreased in Nx. These decreases were restored by AST. Although mRNA levels were not altered, the expression levels of tight junction protein ZO-1 and Claudin-1 were decreased in Nx, which were mitigated in Nx+AST. The analysis of the gut flora showed the decrease of the Lactobacillus in Nx. This decrease was also reversed in Nx+AST. Regression analysis revealed that the number of Lactobacillus in colon was significantly associated with serum creatinine levels. In Caco-2 cells, treatment with indole, precursor of IS produced in the colon downregulated mucin-2 expression as well as Occludin and ZO-1 expressions. Conclusions: In the previous reports, tight junction proteins were downregulated and intestinal barrier function was impaired in mucin-2 deficient mice. It has been also reported that mucin-2 enhanced the growth of Lactobacillus by inducing their adhesion to the colon. Our data demonstrated that AST improved the gut environment favorable to Lactobacillus which affected the expressions of the tight junction proteins. And indole directly influences the tight junction and mucin-2. These effects provide a novel mechanism whereby AST120 or probiotic therapy improves renal function through modulating the gut environment.

s presented at the 19th Annual Congress of the Belgian Society of Internal Medicine 12-13 December 2014 – Auditorium Brouwer, Medical Campus Vrije Universiteit Brussel, Laarbeeklaan 103, Brussels