Gut Inflammation in Axial Spondyloarthritis Patients is Characterized by a Marked Type 17 Skewed Mucosal Innate-like T Cell Signature.

OBJECTIVE: Patients with spondyloarthritis (SpA) often present with microscopic signs of gut inflammation, a risk factor for progressive disease. We investigated whether mucosal innate-like T cells are involved in dysregulated interleukin-23 (IL-23)/IL-17 responses in the gut-joint axis in SpA. METHODS: Ileal and colonic intraepithelial lymphocytes (IELs), lamina propria lymphocytes (LPLs), and paired peripheral blood mononuclear cells (PBMCs) were isolated from treatment-naive patients with nonradiographic axial SpA with (n = 11) and without (n = 14) microscopic gut inflammation and healthy controls (n = 15) undergoing ileocolonoscopy. The presence of gut inflammation was assessed histopathologically. Immunophenotyping of innate-like T cells and conventional T cells was performed using intracellular flow cytometry. Unsupervised clustering analysis was done by FlowSOM technology. Serum IL-17A levels were measured via Luminex. RESULTS: Microscopic gut inflammation in nonradiographic axial SpA was characterized by increased ileal intraepithelial γδ-hi T cells, a γδ-T cell subset with elevated γδ-T cell receptor expression. γδ-hi T cells were also increased in PBMCs of patients with nonradiographic axial SpA versus healthy controls and were strongly associated with Ankylosing Spondylitis Disease Activity Score. The abundance of mucosal-associated invariant T cells and invariant natural killer T cells was unaltered. Innate-like T cells in the inflamed gut showed increased RORγt, IL-17A, and IL-22 levels with loss of T-bet, a signature that was less pronounced in conventional T cells. Presence of gut inflammation was associated with higher serum IL-17A levels. In patients treated with tumor necrosis factor blockade, the proportion of γδ-hi cells and RORγt expression in blood was completely restored. CONCLUSION: Intestinal innate-like T cells display marked type 17 skewing in the inflamed gut mucosa of patients with nonradiographic axial SpA. γδ-hi T cells are linked to intestinal inflammation and disease activity in SpA.

Gut microbiota as a driver of the interindividual variability of cardiometabolic effects from tea polyphenols.

Tea polyphenols have been extensively studied for their preventive properties against cardiometabolic diseases. Nevertheless, the evidence of these effects from human intervention studies is not always consistent, mainly because of a large interindividual variability. The bioavailability of tea polyphenols is low, and metabolism of tea polyphenols highly depends on individual gut microbiota. The accompanying reciprocal relationship between tea polyphenols and gut microbiota may result in alterations in the cardiometabolic effects, however, the underlying mechanism of which is little explored. This review summarizes tea polyphenols-microbiota interaction and its contribution to interindividual variability in cardiometabolic effects. Currently, only a few bacteria that can biodegrade tea polyphenols have been identified and generated metabolites and their bioactivities in metabolic pathways are not fully elucidated. A deeper understanding of the role of complex interaction necessitates fully individualized data, the ntegration of multiple-omics platforms and development of polyphenol-centered databases. Knowledge of this microbial contribution will enable the functional stratification of individuals in the gut microbiota profile (metabotypes) to clarify interindividual variability in the health effects of tea polyphenols. This could be used to predict individual responses to tea polyphenols consumption, hence bringing us closer to personalized nutrition with optimal dose and additional supplementation of specific microorganisms.

Oral and Gut Microbial Carbohydrate-Active Enzymes Landscape in Health and Disease.

Inter-individual variability in the microbial gene complement encoding for carbohydrate-active enzymes (CAZymes) can profoundly regulate how the host interacts with diverse carbohydrate sources thereby influencing host health. CAZy-typing, characterizing the microbiota-associated CAZyme-coding genes within a host individual, can be a useful tool to predict carbohydrate pools that the host can metabolize, or identify which CAZyme families are underrepresented requiring supplementation via microbiota transplantation or probiotics. CAZy-typing, moreover, provides a novel framework to search for disease biomarkers. As a proof of concept, we used publicly available metagenomes (935) representing 310 type strain bacterial genomes to establish the link between disease status and CAZymes in the oral and gut microbial ecosystem. The abundance and distribution of 220 recovered CAZyme families in saliva and stool samples from patients with colorectal cancer, rheumatoid arthritis, and type 1 diabetes were compared with healthy subjects. Based on the multivariate discriminant analysis, the disease phenotype did not alter the CAZyme profile suggesting a functional conservation in carbohydrate metabolism in a disease state. When disease and healthy CAZyme profiles were contrasted in differential analysis, CAZyme markers that were underrepresented in type 1 diabetes (15), colorectal cancer (12), and rheumatoid arthritis (5) were identified. Of interest, are the glycosyltransferase which can catalyze the synthesis of glycoconjugates including lipopolysaccharides with the potential to trigger inflammation, a common feature in many diseases. Our analysis has also confirmed the expansive carbohydrate metabolism in the gut as evidenced by the overrepresentation of CAZyme families in the gut compared to the oral site. Nevertheless, each site exhibited specific CAZyme markers. Taken together, our analysis provides an insight into the CAZyme landscape in health and disease and has demonstrated the diversity in carbohydrate metabolism in host-microbiota which can be a sound basis for optimizing the selection of pre, pro, and syn-biotic candidate products.

Versatile human in vitro triple coculture model coincubated with adhered gut microbes reproducibly mimics pro-inflammatory host-microbe interactions in the colon.

The colonic epithelial barrier is vital to preserve gut and host health by maintaining the immune homeostasis between host and microbes. The mechanisms underlying beneficial or harmful host-microbe interactions are poorly understood and impossible to study in vivo given the limited accessibility and ethical constraints. Moreover, existing in vitro models lack the required cellular complexity for the routine, yet profound, analysis of the intricate interplay between different types of host and microbial cells. We developed and characterized a broadly applicable, easy-to-handle in vitro triple coculture model that combines chemically-induced macrophage-like, goblet and epithelial cells covered by a mucus layer, which can be coincubated with complex human-derived gut microbiota samples for 16 h. Comparison with a standard epithelial monolayer model revealed that triple cocultures produce thicker mucus layers, morphologically organize in a network and upon exposure to human-derived gut microbiota samples, respond via pro-inflammatory cytokine production. Both model systems, however, were not suffering from cytotoxic stress or different microbial loads, indicating that the obtained endpoints were caused by the imposed conditions. Addition of the probiotic Lactobacillus rhamnosus GG to assess its immunomodulating capacity in the triple coculture slightly suppressed pro-inflammatory cytokine responses, based on transcriptomic microarray analyses. TNF conditioning of the models prior to microbial exposure did not cause shifts in cytokines, suggesting a strong epithelial barrier in which TNF did not reach the basolateral side. To conclude, the triple coculture model is tolerable towards manipulations and allows to address mechanistic host-microbe research questions in a stable in vitro environment.

Aberrant gut-microbiota-immune-brain axis development in premature neonates with brain damage.

Premature infants are at substantial risk for suffering from perinatal white matter injury. Though the gut microbiota has been implicated in early-life development, a detailed understanding of the gut-microbiota-immune-brain axis in premature neonates is lacking. Here, we profiled the gut microbiota, immunological, and neurophysiological development of 60 extremely premature infants, which received standard hospital care including antibiotics and probiotics. We found that maturation of electrocortical activity is suppressed in infants with severe brain damage. This is accompanied by elevated γδ T cell levels and increased T cell secretion of vascular endothelial growth factor and reduced secretion of neuroprotectants. Notably, Klebsiella overgrowth in the gut is highly predictive for brain damage and is associated with a pro-inflammatory immunological tone. These results suggest that aberrant development of the gut-microbiota-immune-brain axis may drive or exacerbate brain injury in extremely premature neonates and represents a promising target for novel intervention strategies.

Efficacy and safety of spore-forming probiotics in the treatment of functional dyspepsia: a pilot randomised, double-blind, placebo-controlled trial.

BACKGROUND: Current treatments for functional dyspepsia have limited efficacy or present safety issues. We aimed to assess spore-forming probiotics in functional dyspepsia as monotherapy or add-on therapy to long-term treatment with proton-pump inhibitors. METHODS: In this single-centre, randomised, double-blind, placebo-controlled pilot trial that took place at University Hospitals Leuven (Leuven, Belgium), adult patients (≥18 years) with functional dyspepsia (as defined by Rome IV criteria, on proton-pump inhibitors or off proton-pump inhibitors) were randomly assigned (1:1) via computer-generated blocked lists, stratified by proton-pump inhibitor status, to receive 8 weeks of treatment with probiotics (Bacillus coagulans MY01 and Bacillus subtilis MY02, 2·5 × 109 colony-forming units per capsule) or placebo consumed twice per day, followed by an open-label extension phase of 8 weeks. Individuals with a history of abdominal surgery, diabetes, coeliac or inflammatory bowel disease, active psychiatric conditions, and use of immunosuppressant drugs, antibiotics, or probiotics in the past 3 months were excluded. All patients and on-site study personnel were masked to treatment allocation in the first 8 weeks. Symptoms, immune activation, and faecal microbiota were assessed and recorded. The primary endpoint was a decrease of at least 0·7 in the postprandial distress syndrome (PDS) score of the Leuven Postprandial Distress Scale in patients with a baseline PDS score of 1 or greater (at least mild symptoms), assessed in the intention-to-treat population. This study is registered with ClinicalTrials.gov, NCT04030780. FINDINGS: Between June 3, 2019, and March 11, 2020, of 93 individuals assessed for eligibility, we included 68 patients with functional dyspepsia (51 [75%] women, mean age 40·1 years [SD 14·4], 34 [50%] on proton-pump inhibitors). We randomly assigned 32 participants to probiotics and 36 to placebo. The proportion of clinical responders was higher with probiotics (12 [48%] of 25) than placebo (six [20%] of 30; relative risk 1·95 [95% CI 1·07-4·11]; p=0·028). The number of patients with adverse events was similar with probiotics (five [16%] of 32) and placebo (12 [33%] of 36). Two serious adverse events occurring during the open-label phase (appendicitis and syncope in two separate patients) were assessed as unlikely to be related to the study product. INTERPRETATION: In this exploratory study, B coagulans MY01 and B subtilis MY02 were efficacious and safe in the treatment of functional dyspepsia. Participants had potentially beneficial immune and microbial changes, which could provide insights into possible underlying mechanisms as future predictors or treatment targets. FUNDING: MY HEALTH.

Assessment of bioaccessible and dialyzable fractions of nickel in food products and their impact on the chronic exposure of Belgian population to nickel.

This study aimed to investigate bioaccessible/dialyzable fractions of nickel in selected foods and to clarify the impact of the food digestion/absorption on the final exposure of consumers to nickel. In vitro gastrointestinal incubation experiments were conducted to estimate the bioaccessibility of nickel in different foods. For estimation of a dialyzable fraction, dialysis filtration was conducted. Highest bioaccessibility (99.6%) was observed for wheat-based breakfast cereals. Lowest bioaccessibilities was observed for dried-fruits (on average 20.4%). Highest (61.5%) and lowest (24.5%) dialyzable fractions were observed for wheat-based breakfast cereal and chocolate respectively. Bioaccessible/dialyzable fractions based exposure assessments were highlighted the overestimation of exposures calculated based on total nickel concentrations in foods. This is particularly important when exposure values were compared with toxicological thresholds in a risk characterization study. When threshold values have been obtained through animal studies in which nickel was dosed at 100% accessibility/availability, e.g. nickel salts this is even more important.

Spatial and temporal modulation of enterotoxigenic E. coli H10407 pathogenesis and interplay with microbiota in human gut models.

BACKGROUND: Enterotoxigenic Escherichia coli (ETEC) substantially contributes to the burden of diarrheal illnesses in developing countries. With the use of complementary in vitro models of the human digestive environment, TNO gastrointestinal model (TIM-1), and Mucosal Simulator of the Human Intestinal Microbial Ecosystem (M-SHIME), we provided the first detailed report on the spatial-temporal modulation of ETEC H10407 survival, virulence, and its interplay with gut microbiota. These systems integrate the main physicochemical parameters of the human upper digestion (TIM-1) and simulate the ileum vs ascending colon microbial communities and luminal vs mucosal microenvironments, captured from six fecal donors (M-SHIME). RESULTS: A loss of ETEC viability was noticed upon gastric digestion, while a growth renewal was found at the end of jejunal and ileal digestion. The remarkable ETEC mucosal attachment helped to maintain luminal concentrations above 6 log10 mL-1 in the ileum and ascending colon up to 5 days post-infection. Seven ETEC virulence genes were monitored. Most of them were switched on in the stomach and switched off in the TIM-1 ileal effluents and in a late post-infectious stage in the M-SHIME ascending colon. No heat-labile enterotoxin production was measured in the stomach in contrast to the ileum and ascending colon. Using 16S rRNA gene-based amplicon sequencing, ETEC infection modulated the microbial community structure of the ileum mucus and ascending colon lumen. CONCLUSIONS: This study provides a better understanding of the interplay between ETEC and gastrointestinal cues and may serve to complete knowledge on ETEC pathogenesis and inspire novel prophylactic strategies for diarrheal diseases.

Lacticaseibacillus casei AMBR2 modulates the epithelial barrier function and immune response in a donor-derived nasal microbiota manner.

Live biotherapeutic products (LBP) are emerging as alternative treatment strategies for chronic rhinosinusitis. The selection of interesting candidate LBPs often involves model systems that do not include the polymicrobial background (i.e. the host microbiota) in which they will be introduced. Here, we performed a screening in a simplified model system of upper respiratory epithelium to assess the effect of nasal microbiota composition on the ability to attach and grow of a potential LBP, Lacticaseibacillus casei AMBR2, in this polymicrobial background. After selecting the most permissive and least permissive donor, L. casei AMBR2 colonisation in their respective polymicrobial backgrounds was assessed in more physiologically relevant model systems. We examined cytotoxicity, epithelial barrier function, and cytokine secretion, as well as bacterial cell density and phenotypic diversity in differentiated airway epithelium based models, with or without macrophage-like cells. L. casei AMBR2 could colonize in the presence of both selected donor microbiota and increased epithelial barrier resistance in presence of donor-derived nasal bacteria, as well as anti-inflammatory cytokine secretion in the presence of macrophage-like cells. This study highlights the potential of L. casei AMBR2 as LBP and the necessity to employ physiologically relevant model systems to investigate host-microbe interaction in LBP research.

Short-term supplementation of celecoxib-shifted butyrate production on a simulated model of the gut microbial ecosystem and ameliorated in vitro inflammation.

Celecoxib has been effective in the prevention and treatment of chronic inflammatory disorders through inhibition of altered cyclooxygenase-2 (COX-2) pathways. Despite the benefits, continuous administration may increase risk of cardiovascular events. Understanding microbiome-drug-host interactions is fundamental for improving drug disposition and safety responses of colon-targeted formulations, but little information is available on the bidirectional interaction between individual microbiomes and celecoxib. Here, we conducted in vitro batch incubations of human faecal microbiota to obtain a mechanistic proof-of-concept of the short-term impact of celecoxib on activity and composition of colon bacterial communities. Celecoxib-exposed microbiota shifted metabolic activity and community composition, whereas total transcriptionally active bacterial population was not significantly changed. Butyrate production decreased by 50% in a donor-dependent manner, suggesting that celecoxib impacts in vitro fermentation. Microbiota-derived acetate has been associated with inhibition of cancer markers and our results suggest uptake of acetate for bacterial functions when celecoxib was supplied, which potentially favoured bacterial competition for acetyl-CoA. We further assessed whether colon microbiota modulates anti-inflammatory efficacy of celecoxib using a simplified inflammation model, and a novel in vitro simulation of the enterohepatic metabolism. Celecoxib was responsible for only 5% of the variance in bacterial community composition but celecoxib-exposed microbiota preserved barrier function and decreased concentrations of IL-8 and CXCL16 in a donor-dependent manner in our two models simulating gut inflammatory milieu. Our results suggest that celecoxib-microbiome-host interactions may not only elicit adaptations in community composition but also in microbiota functionality, and these may need to be considered for guaranteeing efficient COX-2 inhibition.

Gap analysis of nickel bioaccessibility and bioavailability in different food matrices and its impact on the nickel exposure assessment.

The metal nickel is well known to cause nickel allergy in sensitive humans by prolonged dermal contact to materials releasing (high) amounts of nickel. Oral nickel exposure via water and food intake is of potential concern. Nickel is essential to plants and animals and can be naturally found in food products or contamination may occur across the agro-food chain. This gap analysis is an evaluation of nickel as a potential food safety hazard causing a risk for human health. In the first step, the available data regarding the occurrence of nickel and its contamination in food and drinks have been collected through literature review. Subsequently, a discussion is held on the potential risks associated with this contamination. Elevated nickel concentrations were mostly found in plant-based foods, e.g. legumes and nuts in which nickel of natural origin is expected. However, it was observed that dedicated and systematic screening of foodstuffs for the presence of nickel is currently still lacking. In a next step, published studies on exposure of humans to nickel via foods and drinks were critically evaluated. Not including bioaccessibility and/or bioavailability of the metal may lead to an overestimation of the exposure of the body to nickel via food and drinks. This overestimation may be problematic when the measured nickel level in foods is high and bioaccessibility and/or bioavailability of nickel in these products is low. Therefore, this paper analyzes the outcomes of the existing dietary intake and bioaccessibility/bioavailability studies conducted for nickel. Besides, the available gaps in nickel bioaccessibility and/or bioavailability studies have been clarified in this paper. The reported bioaccessibility and bioavailability percentages for different food and drinks were found to vary between

Supplementation of a propionate-producing consortium improves markers of insulin resistance in an in vitro model of gut-liver axis.

Gut-liver cross talk is an important determinant of human health with profound effects on energy homeostasis. While gut microbes produce a huge range of metabolites, specific compounds such as short-chain fatty acids (SCFAs) can enter the portal circulation and reach the liver (Brandl K, Schnabl B. Curr Opin Gastroenterol 33: 128-133, 2017), a central organ involved in glucose homeostasis and diabetes control. Propionate is a major SCFA involved in activation of intestinal gluconeogenesis (IGN), thereby regulating food intake, enhancing insulin sensitivity, and leading to metabolic homeostasis. Although microbiome-modulating strategies may target the increased microbial production of propionate, it is not clear whether such an effect spreads through to the hepatic cellular level. Here, we designed a propionate-producing consortium using a selection of commensal gut bacteria, and we investigated how their delivered metabolites impact an in vitro enterohepatic model of insulin resistance. Glycogen storage on hepatocyte-like cells and inflammatory markers associated with insulin resistance were evaluated to understand the role of gut metabolites on gut-liver cross talk in a simulated scenario of insulin resistance. The metabolites produced by our consortium increased glycogen synthesis by ~57% and decreased proinflammatory markers such as IL-8 by 12%, thus elucidating the positive effect of our consortium on metabolic function and low-grade inflammation. Our results suggest that microbiota-derived products can be a promising multipurpose strategy to modulate energy homeostasis, with the potential ability to assist in managing metabolic diseases due to their adaptability.

Dual and Triple Epithelial Coculture Model Systems with Donor-Derived Microbiota and THP-1 Macrophages To Mimic Host-Microbe Interactions in the Human Sinonasal Cavities.

The epithelium of the human sinonasal cavities is colonized by a diverse microbial community, modulating epithelial development and immune priming and playing a role in respiratory disease. Here, we present a novel in vitro approach enabling a 3-day coculture of differentiated Calu-3 respiratory epithelial cells with a donor-derived bacterial community, a commensal species (Lactobacillus sakei), or a pathobiont (Staphylococcus aureus). We also assessed how the incorporation of macrophage-like cells could have a steering effect on both epithelial cells and the microbial community. Inoculation of donor-derived microbiota in our experimental setup did not pose cytotoxic stress on the epithelial cell layers, as demonstrated by unaltered cytokine and lactate dehydrogenase release compared to a sterile control. Epithelial integrity of the differentiated Calu-3 cells was maintained as well, with no differences in transepithelial electrical resistance observed between coculture with donor-derived microbiota and a sterile control. Transition of nasal microbiota from in vivo to in vitro conditions maintained phylogenetic richness, and yet a decrease in phylogenetic and phenotypic diversity was noted. Additional inclusion and coculture of THP-1-derived macrophages did not alter phylogenetic diversity, and yet donor-independent shifts toward higher Moraxella and Mycoplasma abundance were observed, while phenotypic diversity was also increased. Our results demonstrate that coculture of differentiated airway epithelial cells with a healthy donor-derived nasal community is a viable strategy to mimic host-microbe interactions in the human upper respiratory tract. Importantly, including an immune component allowed us to study host-microbe interactions in the upper respiratory tract more in depth.IMPORTANCE Despite the relevance of the resident microbiota in sinonasal health and disease and the need for cross talk between immune and epithelial cells in the upper respiratory tract, these parameters have not been combined in a single in vitro model system. We have developed a coculture system of differentiated respiratory epithelium and natural nasal microbiota and incorporated an immune component. As indicated by absence of cytotoxicity and stable cytokine profiles and epithelial integrity, nasal microbiota from human origin appeared to be well tolerated by host cells, while microbial community composition remained representative for that of the human (sino)nasal cavity. Importantly, the introduction of macrophage-like cells enabled us to obtain a differential readout from the epithelial cells dependent on the donor microbial background to which the cells were exposed. We conclude that both model systems offer the means to investigate host-microbe interactions in the upper respiratory tract in a more representative way.

Berry-Enriched Diet in Salt-Sensitive Hypertensive Rats: Metabolic Fate of (Poly)Phenols and the Role of Gut Microbiota.

Diets rich in (poly)phenols are associated with a reduced reduction in the incidence of cardiovascular disorders. While the absorption and metabolism of (poly)phenols has been described, it is not clear how their metabolic fate is affected under pathological conditions. This study evaluated the metabolic fate of berry (poly)phenols in an in vivo model of hypertension as well as the associated microbiota response. Dahl salt-sensitive rats were fed either a low-salt diet (0.26% NaCl) or a high-salt diet (8% NaCl), with or without a berry mixture (blueberries, blackberries, raspberries, Portuguese crowberry and strawberry tree fruit) for 9 weeks. The salt-enriched diet promoted an increase in the urinary excretion of berry (poly)phenol metabolites, while the abundance of these metabolites decreased in faeces, as revealed by UPLC-MS/MS. Moreover, salt and berries modulated gut microbiota composition as demonstrated by 16S rRNA analysis. Some changes in the microbiota composition were associated with the high-salt diet and revealed an expansion of the families Proteobacteria and Erysipelotrichaceae. However, this effect was mitigated by the dietary supplementation with berries. Alterations in the metabolic fate of (poly)phenols occur in parallel with the modulation of gut microbiota in hypertensive rats. Thus, beneficial effects of (poly)phenols could be related with these interlinked modifications, between metabolites and microbiota environments.

Effects of Olive and Pomegranate By-Products on Human Microbiota: A Study Using the SHIME® in Vitro Simulator.

Two by-products containing phenols and polysaccharides, a "pâté" (OP) from the extra virgin olive oil milling process and a decoction of pomegranate mesocarp (PM), were investigated for their effects on human microbiota using the SHIME® system. The ability of these products to modulate the microbial community was studied simulating a daily intake for nine days. Microbial functionality, investigated in terms of short chain fatty acids (SCFA) and NH4+, was stable during the treatment. A significant increase in Lactobacillaceae and Bifidobacteriaceae at nine days was induced by OP mainly in the proximal tract. Polyphenol metabolism indicated the formation of tyrosol from OP mainly in the distal tract, while urolithins C and A were produced from PM, identifying the human donor as a metabotype A. The results confirm the SHIME® system as a suitable in vitro tool to preliminarily investigate interactions between complex botanicals and human microbiota before undertaking more challenging human studies.

Cell line-dependent increase in cellular quercetin accumulation upon stress induced by valinomycin and lipopolysaccharide, but not by TNF-α.

As the interface between the luminal and internal environment, the intestinal epithelium is strongly exposed to food-related, host-related and microbial stress. Furthermore, the endothelial stress response plays an important role in vascular disease development, which may be improved upon consumption of dietary bioactives such as polyphenols. The impact of the latter, however, is largely individual-dependent and effects are, in most cases, only observed under mild diseased conditions. Here, it is hypothesized that the individual's stressor levels may contribute to this variable response. To this end, the impact of the stressors (i) valinomycin (as model for cereulide, food-related microbial metabolite), (ii) TNF-α (host-related) and (iii) lipopolysaccharide (gram-negative bacterial cell related) on flavonoid accumulation was investigated in several intestinal and endothelial cell lines. Flow cytometry, confocal microscopy and an in-house developed, robust and high-throughput spectrofluorometric method, showed that quercetin accumulated in all tested cell lines in a dose-dependent manner. Upon stress induced by valinomycin and to a lesser extent by lipopolysaccharide, but not by TNF-α, an increased quercetin accumulation was observed in proliferating intestinal and endothelial cells and not in differentiated intestinal or quiescent endothelial cells. Therefore, flavonoid accumulation may be a potential cellular stress response mechanism which strongly depends on the applied stressor, flavonoid, cell line and even growth conditions. This opens perspectives for further understanding the mechanisms by which cellular stress may shape the individual's response to bioactive compounds.

The response of five intestinal cell lines to anoxic conditions in vitro.

BACKGROUND INFORMATION: In vivo oxygen levels in tissues range from 1% to 15%, while mechanistic cell culture studies employ an atmospheric oxygen level of 21% to grow cells. These oxygen concentrations are therefore not representative for conditions where the cell response is dependent on oxygen partial pressure. In pathological situation, such as (colon) cancer or chronic inflammation, tissue oxygenation is severely affected, and even under physiological conditions a steep oxygen gradient is present in the large intestine, where epithelial cells co-exist with microbial species, resulting in almost anoxia at the midpoint of the lumen. In these situations, a better characterisation of the essential cellular behaviour under hypoxia or anoxia is required. RESULTS: We have characterised the cellular response of commonly used cell cultures for the study of intestinal epithelial processes and colon cancer development (Caco-2, HT-29, SW480, HCT 116 and LoVo) under conventional normoxic conditions (21% O2 ) and in an anoxic (<0.1% O2 ) environment generated in an anaerobic chamber. In general, anoxic conditions led to lower levels of oxidative stress, a reduction in reduced glutathione/oxidised glutathione (GSH/GSSG) ratio, the shift of the redox status to oxidised glutathione levels, reduced cell proliferation, decreased barrier function and higher glycolysis rates at the expense of oxidative respiration. CONCLUSIONS: Continuous exposure to anoxic conditions, such as occurring at the host-microbe interface in the intestine, may create an adaptive metabolic cellular response of the cells. SIGNIFICANCE: Considering adequate oxygen levels is essential for creating more physiologically relevant models for the study of host-microbe interactions and colon cancer development.

The Safety and Tolerability of a Potential Alginate-Based Iron Chelator; Results of A Healthy Participant Study.

Evidence supporting the ferro-toxic nature of iron in the progression of inflammatory bowel disease (IBD) is becoming well established. A microbial dysbiosis is observed in IBD patients, and intra-luminal colonic-iron is able to support a more pathogenic community of bacteria; whether this is attributed to the development of IBD and how iron could be mediating these microbial changes is still unknown. Dietary fibres are commonly used in pre-biotic supplements to beneficially affect the host by improving the viability of bacterial communities within the colon. Alginates are a class of biopolymers considered as prebiotics due to their fibre-like composition and are able to bind metal cations, in particular, iron. Considering that iron excess is able to negatively alter the microbiome, the use of alginate as a food supplement could be useful in colonic-iron chelation. As such, this first-in-man study aimed to assess whether the use of alginate as a dietary iron chelator was both safe and well tolerated. In addition, the impact of alginate on the microbiome and iron levels was assessed by using an intestinal model SHIME (Simulation of the Human Intestinal Microbial Ecosystem). Alginate was supplemented into the diets (3 g/day) of healthy volunteers (n = 17) for 28 days. Results from this study suggest that daily ingestion of 3 g alginate was well tolerated with very minor side effects. There were no detrimental changes in a variety of haematological parameters or the intestinal microbiome. The bacterial communities within the SHIME model were also not influenced by iron and or alginate; it is possible that alginate may be susceptible to bacterial or enzymatic degradation within the gastro-intestinal tract.

Development of a host-microbiome model of the small intestine.

The intestinal epithelium plays an essential role in the balance between tolerant and protective immune responses to infectious agents. In vitro models do not typically consider the innate immune response and gut microbiome in detail, so these models do not fully mimic the physiologic aspects of the small intestine. We developed and characterized a long-term in vitro model containing enterocyte, goblet, and immune-like cells exposed to a synthetic microbial community representative of commensal inhabitants of the small intestine. This model showed differential responses toward a synthetic microbial community of commensal bacterial inhabitants of the small intestine in the absence or presence of LPS from Escherichia coli O111:B4. Simultaneous exposure to LPS and microbiota induced impaired epithelial barrier function; increased production of IL-8, IL-6, TNF-α, and C-X-C motif chemokine ligand 16; and augmented differentiation and adhesion of macrophage-like cells and the overexpression of dual oxidase 2 and TLR-2 and -4 mRNA. In addition, the model demonstrated the ability to assess the adhesion of specific bacterial strains from the synthetic microbial community-more specifically, Veillonella parvula-to the simulated epithelium. This novel in vitro model may assist in overcoming sampling and retrieval difficulties when studying host-microbiome interactions in the small intestine.-Calatayud, M., Dezutter, O., Hernandez-Sanabria, E., Hidalgo-Martinez, S., Meysman, F. J. R., Van de Wiele, T. Development of a host-microbiome model of the small intestine.