BACKGROUND:Alterations in the intestinal microbiota contribute to the pathogenesis of various cardiovascular disorders, but how they affect the development of Kawasaki disease (KD) an acute pediatric vasculitis, remains unclear. METHODS:We used the Lactobacillus casei cell wall extract (LCWE) murine model of KD vasculitis to assess the contribution of the intestinal microbiota to the development of vascular inflammation. We evaluated the severity of vasculitis in microbiota-depleted mice. 16S rRNA gene sequencing was used to characterize the fecal microbiome composition of LCWE-injected mice. Some groups of mice were orally treated with selected live or pasteurized bacteria, short-chain fatty acids, or Amuc_1100, the Toll-like receptor 2 signaling outer membrane protein from Akkermansia muciniphila, and their impact on vasculitis development was assessed. RESULTS:We report that depleting the gut microbiota reduces the development of cardiovascular inflammation in a murine model mimicking KD vasculitis. The development of cardiovascular lesions was associated with alterations in the intestinal microbiota composition and, notably, a decreased abundance of Akkermansia muciniphila and Faecalibacterium prausnitzii. Oral supplementation with either of these live or pasteurized individual bacteria or with short-chain fatty acids produced by them attenuated cardiovascular inflammation, as reflected by decreased local immune cell infiltrations. Treatment with Amuc_1100 also reduced the severity of vascular inflammation. CONCLUSIONS:This study reveals an underappreciated gut microbiota-cardiovascular inflammation axis in KD vasculitis pathogenesis and identifies specific intestinal commensals that regulate vasculitis in mice by producing metabolites or via extracellular proteins capable of enhancing and supporting gut barrier function.

Evidence suggests that a significant interplay exists between the host gut microbiota and both the transmission and severity of malaria. Therefore, we explored the association between malaria and the gut microbiota across various geographic regions, considering host's nutritional habits, helminth coinfections and age. This observational study was conducted in 3 malaria-endemic provinces of Rwanda: West, South and East. Demographic data, blood and fecal samples were collected from 169 participants (85 females and 84 males) aged between 2-78 years. We used questionnaire-derived qualitative data based on geographic regions, age, and nutrition. Malaria and soil-transmitted helminth diagnosis was assessed by microscopy. The gut microbial composition was analyzed based on bacterial 16S rRNA gene amplicon sequencing. We observed that preschool children had a significantly lower microbiota diversity compared to both school children (q = 0.027, K-Wallis) and adults (q = 0.011, K-Wallis). Unlike age, infection status (uninfected, malaria alone, soil-transmitted helminth alone or coinfection) was not significantly associated with the gut microbiota. However, using Bray-Curtis distances, we found a significantly differential gut microbial beta-diversity with a convergent distribution in the Western province compared to the other provinces (q = 0.0045, pairwise PERMANOVA). This geographic difference was not explained by any change in energy intake, protein, lipids, or carbohydrates consumption but was likely due to lower dietary fibre intake in the West compared to the South (q < 0.0001, ANOVA) and the East (q = 0.07, ANOVA). In conclusion, we have not found significant links between infection and gut microbiota. However, we showed a significant difference in the gut microbiota composition of people living in different geographic locations in Rwanda, possibly due to their nutritional habits.

Human milk oligosaccharides (HMOs) promote the development of the infant gut microbiota, notably through the growth of beneficial bacteria.

Obesity is associated with alterations in the gut microbiome that may contribute to metabolic and mental health disturbances. Fecal microbiota transplantation (FMT) from humans to mice is a model proposed to study human microbiota-associated disorders. In this study, we investigated whether gut microbiota from human donors with obesity could affect behavior and metabolomic profiles of mice. Stools from donors with obesity and from lean donors were inoculated to antibiotic-pretreated mice fed a standard low-fat diet throughout the experiment. Obese-recipient mice exhibited a lower bacterial alpha-diversity and limited changes in specific taxa (e.g., an increase in Eubacterium) but were similar to lean-recipient mice in terms of dietary intake, body weight, fat mass, anxiety/depression-like behavior and glucose homeostasis. Non-targeted LC-MS metabolomic analysis revealed no change in the portal and cava serum samples. However, 1-methylnicotinamide, indole-3-acetic acid (I3A) and methyllysine were increased in the cecal content of obese-recipient compared to lean-recipient mice. Microbial metabolites derived from amino acids were positively correlated with Eubacterium. These results indicate that FMT from donors with obesity to mice fed chow diet (low in lipids) leads to minor but persistent change in intestinal microbial-derived metabolites, without recapitulating the metabolic and behavioral alterations of obesity.

The global type 2 diabetes epidemic is a major health crisis. Although the microbiome has roles in the onset of insulin resistance (IR), low-grade inflammation and diabetes, the microbial compounds controlling these processes remain to be discovered. Here, we show that the microbial metabolite trimethylamine (TMA) decouples inflammation and IR from diet-induced obesity by inhibiting interleukin-1 receptor-associated kinase 4 (IRAK4), a central kinase in the Toll-like receptor pathway sensing danger signals. TMA blunts TLR4 signalling in primary human hepatocytes and peripheral blood monocytic cells and rescues mouse survival after lipopolysaccharide-induced septic shock. Genetic deletion and chemical inhibition of IRAK4 result in metabolic and immune improvements in high-fat diets. Remarkably, our results suggest that TMA—unlike its liver co-metabolite trimethylamine N-oxide, which is associated with cardiovascular disease—improves immune tone and glycemic control in diet-induced obesity. Altogether, this study supports the emerging role of the kinome in the microbial–mammalian chemical crosstalk. The microbial metabolite trimethylamine (TMA), the precursor of TMAO, which is associated with adverse cardiometabolic outcomes, is shown to have beneficial metabolic and anti-inflammatory effects in the host in the context of obesity.

Obesity is reaching epidemic proportions worldwide. This excessive increase of adipose tissue is a risk factor for the development of multiple diseases and premature death. Amongst associated diseases, metabolic syndrome is one of the main comorbidities of obesity. In this context, the gut microbiota has been recognized as both shaping and responding to host energy metabolism. Recently metabolomics has emerged as a powerful tool to capture a snapshot of the metabolites present in a specific tissue, providing new insights into host-microbiota interactions. Integrating metabolomics with gut microbiota studies could help us better understand how specific species impact on host metabolomic profile. Dysosmobacter welbionis has been identified as a promising next generation beneficial bacteria with potential effects on fat mass and glucose metabolism in mice, and fecal Dysosmobacter spp. concentration was inversely correlated to body mass index fasting glucose and plasmatic HbA1c in humans. Concentration of Dysosmobacter spp. was quantified by qPCR in the stools of insulin resistant overweight/obese participants with a metabolic syndrome and plasma metabolites were analyzed using untargeted metabolomics. Correlations between Dysosmobacter spp. fecal abundance and the 1169 identified plasma metabolites were uncovered using Spearman correlations followed by a false discovery rate correction. Interestingly, among the detected metabolites, Dysosmobacter spp. was exclusively associated with lipid molecules. Fecal concentration of Dysosmobacter spp. was positively associated with plasmatic levels of five phosphatidylcholines, arachidonate, two monoacylglycerols, twelve diacylglycerols, three lysophosphatidylethanolamines, one phosphatidylinositol and three lysophosphatidylinositols, as well as glycerophosphoethanolamines, glycerophosphatidylcholine and PC(P-16:0). The correlation was particularly interesting with acylcholine and lysophosphatidylcholine metabolites as, respectively, 6/8 and 8/10 detected molecules were positively associated with Dysosmobacter spp. These results suggest that Dysosmobacter spp. plays a specific role in host lipid metabolism. This finding aligns with previous in vivo studies highlighting lipid profile alterations in multiple tissues of mice treated with this bacterium. Further studies are needed to elucidate the underlying mechanisms and assess its potential therapeutic applications.

Undernutrition and malaria remain major global public health challenges. The relationship between nutritional status and malaria infection is complex. Better understanding of their association is needed to improve prevention and control of these health conditions. Undernutrition can be assessed by evaluating nutrient intake (macronutrient and micronutrient intake) measured through Food Frequency Questionnaire or by using indicators for chronic undernutrition, including the stunting parameter defined by the World Health Organization as child being too short for age. This study aims to investigate associations between inadequate nutrient intake or indicators for undernutrition and malaria infection. The analysis compares malaria cases diagnosed by a positive blood smear against controls composed of other malaria free people living in the same households with the cases (same household grouping). Data collection was conducted between November 2021 and December 2023 across 9 endemic districts located in all four provinces and Kigali City. Regression models were developed to investigate the association between undernutrition (i.e. inadequate nutrient intake or stunting) and malaria infection. Despite numerous nutrition interventions aimed at reducing the burden of undernutrition, the dietary patterns observed in this study remain predominantly imbalanced. The food composition was predominantly made up of starchy staples, accounting for 56.7

Anxiety is a common co-morbidity with obesity and metabolic disease, and can lead to a significant impact on quality of life. The vast differences in the gut microbiota between obese and control individuals provide a potential avenue for therapeutic intervention. A high-fat diet (HFD) in rodent models have been shown to induce anxiety-like behaviour and has been tested through an array of distinct behavioural tests such as the elevated plus maze test, light-dark test and open field test. Despite differences in testing and assessment parameters, the behavioural outcomes have previously yielded similar results. Recent evidence suggests that HFD has an anxiolytic effect on mice, complicating the model. Here, we aimed to confirm whether HFD-fed mice are more susceptible to presenting anxiety-like behaviours. Our findings showed no significant differences in behaviour, plasma corticosterone and inflammation markers between HFD and control diet (CTD) mice, despite considerable differences in adiposity and faecal microbial communities. Additionally, daily oral gavage is one of the most common methods for testing bacterial probiotics in rodent models, but this handling could potentially also cause stress to the mice. Thus, we investigated if daily oral gavage could mask differences in HFD and CTD mice. We found no significant differences in weight, fat mass or anxiety-like behaviour in CTD-fed mice with or without daily oral gavage.

Inulin-type fructans (ITF) are fermentable dietary fibres (DF) that can confer beneficial metabolic health effects through changes in the gut microbiota. Many papers suggest that complex food rich in DF could be more relevant than purified DF in terms of health effect. We compared the prebiotic effect of natural source of inulin (scorzonera) versus native inulin extracted from chicory root in a model of obesity. Mice were fed during 6 weeks a low-fat (LF), high-fat (HF) or high-fat diet enriched with either purified inulin from chicory root (Inu) or lyophilized scorzonera (Sco), with the same amount of ITF intake (10

Dolutegravir and bictegravir are second-generation HIV integrase strand transfer inhibitors (INSTIs) that were previously associated with abnormal weight gain. This monocentric cross-sectional study investigates associations between weight gain during the first year after initiation of dolutegravir, bictegravir or other anchor drugs and gut microbiome diversity as well as taxa composition. The study enrolled 79 participants receiving dolutegravir, 32 receiving bictegravir and 10 receiving non-INSTI based regimens. Most of them were treatment experienced at initiation of those anchor drugs agents. Although weight gain was not linked to overall bacterial diversity, strong associations with specific taxa were demonstrated (FDR q < 0.01). Using multiple linear regression, we identified 4 distinct groups of bacteria associated with either dolutegravir, bictegravir, weight loss or weight gain under treatment, allowing a machine learning model to predict 15.9% of the weight gain variability regardless of sex, age and body mass index (RMSE: 0.0126). Dysosmobacter sp. and Haemophilus sp., two bacteria previously associated with host metabolism, were among the strongest predictors. Our findings link INSTIs, weight gain, and the gut microbiome. Future research should investigate the causal role of the identified taxa to improve our understanding of microbiome-drug interactions and further support personalized antiretroviral strategies.Trial registration: Eudra-CT 2020-001103-17 (registration date: 2020-12-01).

In recent years, the gut microbiota and derived metabolites have emerged as relevant players in modulating several brain functions, including energy balance control1–3. This form of distant communication mirrors that of metabolic hormones (for example, leptin, ghrelin), which convey information about the organism’s energy status by exerting effects on diverse brain regions, including the master homeostatic centre, the hypothalamus4. However, whether the hypothalamus is also able to influence gut microbiota composition remains enigmatic. Here we present a study designed to unravel this challenging question. To this aim, we used chemogenetics5 (to selectively activate or inhibit hypothalamic pro-opiomelanocortin or agouti-related peptide neurons) or centrally administered leptin or ghrelin to male mice. Subsequently, we conducted microbiota composition analysis throughout the gut using 16S rRNA gene sequencing. Our results showed that these brain interventions significantly changed the gut microbiota in an anatomical and short-term (2–4 h) fashion. Transcriptomic analysis indicated that these changes were associated with the reconfiguration of neuronal and synaptic pathways in the duodenum concomitant with increased sympathetic tone. Interestingly, diet-induced obesity attenuated the brain-mediated changes triggered by leptin in gut microbiota communities and sympathetic activation. Our findings reveal a previously unanticipated brain–gut axis that acutely attunes microbiota composition on fast timescales, with potential implications for meal-to-meal adjustments and systemic energy balance control. Toledo et al. show that metabolic hormones acting in the hypothalamus, along with direct manipulation of hypothalamic neurons, can drive remodelling of microbial communities in the gut within short periods of time.

SUMMARYThe gut microbiota is a major factor contributing to the regulation of energy homeostasis and has been linked to both excessive body weight and accumulation of fat mass (i.e., overweight, obesity) or body weight loss, weakness, muscle atrophy, and fat depletion (i.e., cachexia). These syndromes are characterized by multiple metabolic dysfunctions including abnormal regulation of food reward and intake, energy storage, and low-grade inflammation. Given the increasing worldwide prevalence of obesity, cachexia, and associated metabolic disorders, novel therapeutic strategies are needed. Among the different mechanisms explaining how the gut microbiota is capable of influencing host metabolism and energy balance, numerous studies have investigated the complex interactions existing between nutrition, gut microbes, and their metabolites. In this review, we discuss how gut microbes and different microbiota-derived metabolites regulate host metabolism. We describe the role of the gut barrier function in the onset of inflammation in this context. We explore the importance of the gut-to-brain axis in the regulation of energy homeostasis and glucose metabolism but also the key role played by the liver. Finally, we present specific key examples of how using targeted approaches such as prebiotics and probiotics might affect specific metabolites, their signaling pathways, and their interactions with the host and reflect on the challenges to move from bench to bedside.

IntroductionThe gut barrier, comprising gut microbiota, plays a pivotal role in chronic kidney disease (CKD) progression and nutritional status. This study aimed to explore gut barrier alterations in hemodialyzed (HD) patients, non-HD (NHD) CKD patients, and healthy volunteers.MethodsOur cross-sectional study enrolled 22 HD patients, 11 NHD patients, and 11 healthy volunteers. We evaluated fecal microbiota composition (assessed via bacterial 16S rRNA gene sequencing), fecal IgA levels, surrogate markers of gut permeability, serum cytokines, appetite mediators, nutritional status, physical activity, and quality of life.ResultsHD patients exhibited significant alterations in fecal microbiota composition compared to healthy volunteers, with observed shifts in taxa known to be associated with dietary patterns or producing metabolites acting on human host. In comparison to healthy volunteers, individuals with HD patients exhibited elevated levels of inflammatory markers (CRP, IL-6 and TNF-α), glucagon-like peptide-2, and potential anorexigenic markers (including leptin and peptide YY). NHD patients had increased levels of CRP and peptide YY. Overall fecal microbiota composition was associated with height, soft lean mass, resting energy expenditure, handgrip strength, bone mineral content and plasma albumin and TNF-α.DiscussionCompared to healthy volunteers, HD patients have an altered fecal microbiota composition, a higher systemic inflammation, and a modification in plasma levels of appetite mediators. While some differences align with previous findings, heterogeneity exists likely due to various factors including lifestyle and comorbidities. Despite limitations such as sample size, our study underscores the multifaceted interplay between gut microbiota, physiological markers, and kidney function, warranting further investigation in larger cohorts.

Overconsumption of added sugars has been pointed out as a major culprit in the increasing rates of obesity worldwide, contributing to the rising popularity of non-caloric sweeteners. In order to satisfy the growing demand, industrial efforts have been made to purify the sweet-tasting molecules found in the natural sweetener stevia, which are characterized by a sweet taste free of unpleasant aftertaste. Although the use of artificial sweeteners has raised many concerns regarding metabolic health, the impact of purified stevia components on the latter remains poorly studied. The objective of this project was to evaluate the impact of two purified sweet-tasting components of stevia, rebaudioside A and D (RebA and RebD), on the development of obesity, insulin resistance, hepatic health, bile acid profile, and gut microbiota in a mouse model of diet-induced obesity. Male C57BL/6 J mice were fed an obesogenic high-fat/high-sucrose (HFHS) diet and orally treated with 50 mg/kg of RebA, RebD or vehicle (water) for 12 weeks. An additional group of chow-fed mice treated with the vehicle was included as a healthy reference. At weeks 10 and 12, insulin and oral glucose tolerance tests were performed. Liver lipids content was analyzed. Whole-genome shotgun sequencing was performed to profile the gut microbiota. Bile acids were measured in the feces, plasma, and liver. Liver lipid content and gene expression were analyzed. As compared to the HFHS-vehicle treatment group, mice administered RebD showed a reduced weight gain, as evidenced by decreased visceral adipose tissue weight. Liver triglycerides and cholesterol from RebD-treated mice were lower and lipid peroxidation was decreased. Interestingly, administration of RebD was associated with a significant enrichment of Faecalibaculum rodentium in the gut microbiota and an increased secondary bile acid metabolism. Moreover, RebD decreased the level of lipopolysaccharide-binding protein (LBP). Neither RebA nor RebD treatments were found to impact glucose homeostasis. The daily consumption of two stevia components has no detrimental effects on metabolic health. In contrast, RebD treatment was found to reduce adiposity, alleviate hepatic steatosis and lipid peroxidation, and decrease LBP, a marker of metabolic endotoxemia in a mouse model of diet-induced obesity.

The understanding that changes in microbiome composition can influence chronic human diseases and the efficiency of therapies has driven efforts to develop microbiota-centred therapies such as first and next generation probiotics, prebiotics and postbiotics, microbiota editing and faecal microbiota transplantation. Central to microbiome research is understanding how disease impacts microbiome composition and vice versa, yet there is a problematic issue with the term ‘dysbiosis’, which broadly links microbial imbalances to various chronic illnesses without precision or definition. Another significant issue in microbiome discussions is defining ‘healthy individuals’ to ascertain what characterises a healthy microbiome. This involves questioning who represents the healthiest segment of our population—whether it is those free from illnesses, athletes at peak performance, individuals living healthily through regular exercise and good nutrition or even elderly adults or centenarians who have been tested by time and achieved remarkable healthy longevity. This review advocates for delineating ‘what defines a healthy microbiome?’ by considering a broader range of factors related to human health and environmental influences on the microbiota. A healthy microbiome is undoubtedly linked to gut health. Nevertheless, it is very difficult to pinpoint a universally accepted definition of ‘gut health’ due to the complexities of measuring gut functionality besides the microbiota composition. We must take into account individual variabilities, the influence of diet, lifestyle, host and environmental factors. Moreover, the challenge in distinguishing causation from correlation between gut microbiome and overall health is presented. The review also highlights the resource-heavy nature of comprehensive gut health assessments, which hinders their practicality and broad application. Finally, we call for continued research and a nuanced approach to better understand the intricate and evolving concept of gut health, emphasising the need for more precise and inclusive definitions and methodologies in studying the microbiome.

Here, we present a protocol for the pre-clinical evaluation of gut barrier function and immune interaction in an HT-29/PBMC co-culture model. We describe steps for culturing HT-29 cells and peripheral blood mononuclear cells (PBMCs), assembling the co-culture model, and testing cell viability. We then detail procedures for performing efficacy tests through stress challenges and barrier permeability assays.For complete details on the use and execution of this protocol, please refer to Abot et al.1

Bioluminescence imaging has become an essential non-invasive tool in cancer research for monitoring various cellular processes and tumor progression in vivo. In this article, we aimed to propose a transduction and selection protocol for reliable in vivo bioluminescent measurements in immunocompetent mouse models. Using two different heterogenous luciferase-expressing cell models, we underlined factors influencing transduction. The protocol was tested through an in vitro luciferase activity assay as well as using in vivo longitudinal monitoring of metastases formation (In Vivo Imaging System®). The data were cross validated with histological assessment. Our results demonstrated stable and proportional in vitro and in vivo bioluminescent signals correlating with actual metastatic burden. Furthermore, ex vivo analysis confirmed the accuracy of bioluminescent imaging in quantifying metastatic surface area. This protocol should ensure reliable and reproducible measurements in cancer research utilizing luciferase-positive cell lines, confirming the validity and accuracy of preclinical studies in immunocompetent models.

Plasma proteomics is a precious tool in human disease research but requires extensive sample preparation in order to perform in-depth analysis and biomarker discovery using traditional data-dependent acquisition (DDA). Here, we highlight the efficacy of combining moderate plasma prefractionation and data-independent acquisition (DIA) to significantly improve proteome coverage and depth while remaining cost-efficient. Using human plasma collected from a 20-patient COVID-19 cohort, our method utilizes commonly available solutions for depletion, sample preparation, and fractionation, followed by 3 liquid chromatography-mass spectrometry/MS (LC-MS/MS) injections for a 360 min total DIA run time. We detect 1321 proteins on average per patient and 2031 unique proteins across the cohort. Differential analysis further demonstrates the applicability of this method for plasma proteomic research and clinical biomarker identification, identifying hundreds of differentially abundant proteins at biological concentrations as low as 47 ng/L in human plasma. Data are available via ProteomeXchange with the identifier PXD047901. In summary, this study introduces a streamlined, cost-effective approach to deep plasma proteome analysis, expanding its utility beyond classical research environments and enabling larger-scale multiomics investigations in clinical settings. Our comparative analysis revealed that fractionation, whether the samples were pooled or separate postfractionation, significantly improved the number of proteins quantified. This underscores the value of fractionation in enhancing the depth of plasma proteome analysis, thereby offering a more comprehensive landscape for biomarker discovery in diseases such as COVID-19.

The cell-envelope of Gram-negative bacteria contains endotoxic lipopolysaccharides (LPS) that are recognized by the innate immune system via Toll-Like Receptors (TLRs). The intestinal mucosal symbiont Akkermansia muciniphila is known to confer beneficial effects on the host and has a Gram-negative architecture. Here we show that A. muciniphila LPS lacks the O-polysaccharide repeating unit, with the resulting lipooligosaccharide (LOS) having unprecedented structural and signaling properties. The LOS consists of a complex glycan chain bearing two distinct undeca- and hexadecasaccharide units each containing three 2-keto-3-deoxy-D-manno-octulosonic acid (Kdo) residues. The lipid A moiety appears as a mixture of differently phosphorylated and acylated species and carries either linear or branched acyl moieties. Peritoneal injection of the LOS in mice increased higher gene expression of liver TLR2 than TLR4 (100-fold) and induced high IL-10 gene expression. A. muciniphila LOS was found to signal both through TLR4 and TLR2, whereas lipid A only induced TLR2 in a human cell line. We propose that the unique structure of the A. muciniphila LOS allows interaction with TLR2, thus generating an anti-inflammatory response as to compensate for the canonical inflammatory signaling associated with LOS and TLR4, rationalizing its beneficial host interaction.