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Background The association of circulating trimethylamine-N-oxide (TMAO) with stroke has received limited attention. To address this gap, we examined the associations of serial measures of plasma TMAO with incident ischemic stroke. Methods and Results We used a prospective cohort design with data pooled from 2 cohorts. The settings were the CHS (Cardiovascular Health Study), a cohort of older adults, and the MESA (Multi-Ethnic Study of Atherosclerosis), both in the United States. We measured plasma concentrations of TMAO at baseline and again during the follow-up using high-performance liquid chromatography and mass spectrometry. We assessed the association of plasma TMAO with incident ischemic stroke using proportional hazards regression adjusted for risk factors. The combined cohorts included 11 785 participants without a history of stroke, on average 73 (CHS) and 62 (MESA) years old at baseline, including 60% (CHS) and 53% (MESA) women. We identified 1031 total incident ischemic strokes during a median 15-year follow-up in the combined cohorts. In multivariable analyses, TMAO was significantly associated with incident ischemic stroke risk (hazard ratios comparing a doubling of TMAO: 1.11 [1.03-1.18], P=0.004). The association was linear over the range of TMAO concentrations and appeared restricted to those without diagnosed coronary heart disease. An association with hemorrhagic stroke was not found. Conclusions Plasma TMAO levels are associated with incident ischemic stroke in a diverse population. Registration URL: https://www.clinicaltrials.gov. Unique identifier: NCT00005133.

Trimethylamine n-oxide (TMAO) is a plasma metabolite derived from the meta-organismal pathway involving diet and the gut microbiome and is associated with adverse cardiometabolic phenotypes. Despite robust evidence demonstrating TMAO's clinical significance in populations with cardiometabolic disease, characterizations of plasma TMAO concentrations in healthy adults are few. Further, despite understanding that TMAO generation is due to diet, the gut microbiome, and enzymatic hepatic oxidation, the interplay of these factors by sex, age, and genetics have not been thoroughly studied. Characterizing these relationships will refine our understanding of the factors that influence TMAO concentration and therefore cardiometabolic diseases. Thus, to address these gaps, we first utilized a cohort of healthy adults equally represented by sex, age, and BMI to clarify the relationships among diet, gut microbiome composition, CVD biomarkers, and fasting plasma TMAO in the absence of disease. Next, we used multiple systems-genetics mouse models to perform genome-wide association studies to identify loci regulating fasting plasma TMAO concentrations in different diet environments. Finally, we returned to the healthy human cohort to perform a proof-of-concept precision nutrition study evaluating the postprandial dynamics of plasma TMAO concentration following the consumption of a high-fat mixed macronutrient tolerance test. In all, these studies illuminate the regulation of plasma TMAO in healthy states, which can be leveraged to design preventative and reactive strategies to reduce the metabolite's pathogenicity.

Cardiovascular diseases (CVDs) are the leading cause of death and disability worldwide. The aging population and the rapidly increasing prevalence of obesity and type 2 diabetes will contribute to a growing epidemic of CVDs globally. Despite the extensive investigations in etiology, the pathogenesis of CVDs still not fully understand, and the treatment and prevention for CVDs are still limited. Significant interest has been raised in gut microbiota-host interaction since increasing evidence revealed that gut microbiomes play an important role in human health and diseases, including CVDs. Among more than two thousand gut microbiota metabolites, a compound named trimethylamine N-oxide (TMAO) was revealed to be closely related to CVDs. However, the impact of TMAO on cardiovascular health is still full of controversy and the direct impact of TMAO on heart tissue and cardiomyocytes has not been fully understood yet. In the first chapter, we reviewed the literature on TMAO-related atherosclerosis and cardiomyopathy to give us a general aspect of current research progress in the role of TMAO on CVDs. In this context, we provide an overview of the potential mechanisms underlying TMAO-induced cardiovascular diseases at the cellular and molecular levels, with a focus on atherosclerosis and cardiomyopathy. We also address the direct effects of TMAO on cardiomyocytes (a new and under-researched area) and finally propose TMAO as a potential biomarker and/or therapeutic target for the diagnosis and treatment of patients with CVDs. In the second chapter, the direct impact of TMAO on cardiac function was tested in vivo using wild-type C57B6L mice model. Four experiment groups were enrolled in the feeding protocol, which included 3w (different time points), 6w, and 13w feeding time to reveal the impact of short and longer periods of TMAO consumption on cardiac function. The plasma TMAO was measured by liquid chromatography-tandem mass spectrometry (LC/MS/MS) method at the end of the

Immune dysfunction resulting from allogeneic haematopoietic stem cell transplantation (aHSCT) predisposes one to an elevated risk of cytomegalovirus (CMV) infection. Changes in metabolism have been associated with adverse outcomes, and in this study, we explored the associations between metabolic profiles and post-transplantation CMV infection using plasma samples collected 7-33 days after aHSCT. We included 68 aHSCT recipients from Rigshospitalet, Denmark, 50% of whom experienced CMV infection between days 34-100 post-transplantation. First, we investigated whether 12 metabolites selected based on the literature were associated with an increased risk of post-transplantation CMV infection. Second, we conducted an exploratory network-based analysis of the complete metabolic and lipidomic profiles in relation to clinical phenotypes and biological pathways. Lower levels of trimethylamine N-oxide were associated with subsequent CMV infection (multivariable logistic regression: OR = 0.63; 95% CI = [0.41; 0.87]; p = 0.01). Explorative analysis revealed 12 clusters of metabolites or lipids, among which one was predictive of CMV infection, and the others were associated with conditioning regimens, age upon aHSCT, CMV serostatus, and/or sex. Our results provide evidence for an association between the metabolome and CMV infection post-aHSCT that is independent of known risk factors.

Immune dysfunction resulting from allogeneic haematopoietic stem cell transplantation (aHSCT) predisposes one to an elevated risk of cytomegalovirus (CMV) infection. Changes in metabolism have been associated with adverse outcomes, and in this study, we explored the asso- ciations between metabolic profiles and post-transplantation CMV infection using plasma samples collected 7–33 days after aHSCT. We included 68 aHSCT recipients from Rigshospitalet, Denmark, 50% of whom experienced CMV infection between days 34–100 post-transplantation. First, we inves- tigated whether 12 metabolites selected based on the literature were associated with an increased risk of post-transplantation CMV infection. Second, we conducted an exploratory network-based analysis of the complete metabolic and lipidomic profiles in relation to clinical phenotypes and biological pathways. Lower levels of trimethylamine N-oxide were associated with subsequent CMV infection (multivariable logistic regression: OR = 0.63; 95% CI = [0.41; 0.87]; p = 0.01). Explorative analysis revealed 12 clusters of metabolites or lipids, among which one was predictive of CMV infection, and the others were associated with conditioning regimens, age upon aHSCT, CMV serostatus, and/or sex. Our results provide evidence for an association between the metabolome and CMV infection post-aHSCT that is independent of known risk factors., Immune dysfunction resulting from allogeneic haematopoietic stem cell transplantation (aHSCT) predisposes one to an elevated risk of cytomegalovirus (CMV) infection. Changes in metabolism have been associated with adverse outcomes, and in this study, we explored the associations between metabolic profiles and post-transplantation CMV infection using plasma samples collected 7–33 days after aHSCT. We included 68 aHSCT recipients from Rigshospitalet, Denmark, 50% of whom experienced CMV infection between days 34–100 post-transplantation. First, we investigated whether 12 metabolites selected based on the literature were associated with an increased risk of post-transplantation CMV infection. Second, we conducted an exploratory network-based analysis of the complete metabolic and lipidomic profiles in relation to clinical phenotypes and biological pathways. Lower levels of trimethylamine N-oxide were associated with subsequent CMV infection (multivariable logistic regression: OR = 0.63; 95% CI = [0.41; 0.87]; p = 0.01). Explorative analysis revealed 12 clusters of metabolites or lipids, among which one was predictive of CMV infection, and the others were associated with conditioning regimens, age upon aHSCT, CMV serostatus, and/or sex. Our results provide evidence for an association between the metabolome and CMV infection post-aHSCT that is independent of known risk factors.

The aim of the present study was to analyze the effects of cocoa flavanols and red berry anthocyanins on cardiovascular biomarkers, such as homocysteine, angiotensin-converting enzyme (ACE), nitric oxide (NO), flow-mediated vasodilation (FMD), blood pressure and lipid profile. Additionally, we aimed to ascertain their possible interactions with microbiota related metabolites, such as secondary bile acids (SBA), short-chain fatty acids (SCFA) and trimethylamine N-oxide (TMAO). A randomized, parallel-group study, single-blind for the research team, was performed on 60 healthy volunteers between the ages of 45 and 85, who consumed 2.5 g/day of cocoa powder (9.59 mg/day of total flavanols), 5 g/day of a red berry mixture (13.9 mg/day of total anthocyanins) or 7.5 g/day of a combination of both for 12 weeks. The group that had consumed cocoa showed a significant reduction in TMAO (p = 0.03) and uric acid (p = 0.01) levels in serum, accompanied by an increase in FMD values (p = 0.03) and total polyphenols. corrected by creatinine (p = 0.03) after the intervention. These latter values negatively correlated with the TMAO concentration (R = −0.57, p = 0.02). Additionally, we observed an increase in carbohydrate fermentation in the groups that had consumed cocoa (p = 0.04) and red berries (p = 0.04) between the beginning and the end of the intervention. This increase in carbohydrate fermentation was correlated with lower levels of TC/HDL ratio (p = 0.01), systolic (p = 0.01) and diastolic blood pressure (p = 0.01). In conclusion, our study showed a positive modulation of microbiota metabolism after a regular intake of cocoa flavanols and red berry anthocyanins that led to an improvement in cardiovascular function, especially in the group that consumed cocoa.

Immune dysfunction resulting from allogeneic haematopoietic stem cell transplantation (aHSCT) predisposes one to an elevated risk of cytomegalovirus (CMV) infection. Changes in metabolism have been associated with adverse outcomes, and in this study, we explored the asso- ciations between metabolic profiles and post-transplantation CMV infection using plasma samples collected 7–33 days after aHSCT. We included 68 aHSCT recipients from Rigshospitalet, Denmark, 50% of whom experienced CMV infection between days 34–100 post-transplantation. First, we inves- tigated whether 12 metabolites selected based on the literature were associated with an increased risk of post-transplantation CMV infection. Second, we conducted an exploratory network-based analysis of the complete metabolic and lipidomic profiles in relation to clinical phenotypes and biological pathways. Lower levels of trimethylamine N-oxide were associated with subsequent CMV infection (multivariable logistic regression: OR = 0.63; 95% CI = [0.41; 0.87]; p = 0.01). Explorative analysis revealed 12 clusters of metabolites or lipids, among which one was predictive of CMV infection, and the others were associated with conditioning regimens, age upon aHSCT, CMV serostatus, and/or sex. Our results provide evidence for an association between the metabolome and CMV infection post-aHSCT that is independent of known risk factors., Immune dysfunction resulting from allogeneic haematopoietic stem cell transplantation (aHSCT) predisposes one to an elevated risk of cytomegalovirus (CMV) infection. Changes in metabolism have been associated with adverse outcomes, and in this study, we explored the associations between metabolic profiles and post-transplantation CMV infection using plasma samples collected 7–33 days after aHSCT. We included 68 aHSCT recipients from Rigshospitalet, Denmark, 50% of whom experienced CMV infection between days 34–100 post-transplantation. First, we investigated whether 12 metabolites selected based on the literature were associated with an increased risk of post-transplantation CMV infection. Second, we conducted an exploratory network-based analysis of the complete metabolic and lipidomic profiles in relation to clinical phenotypes and biological pathways. Lower levels of trimethylamine N-oxide were associated with subsequent CMV infection (multivariable logistic regression: OR = 0.63; 95% CI = [0.41; 0.87]; p = 0.01). Explorative analysis revealed 12 clusters of metabolites or lipids, among which one was predictive of CMV infection, and the others were associated with conditioning regimens, age upon aHSCT, CMV serostatus, and/or sex. Our results provide evidence for an association between the metabolome and CMV infection post-aHSCT that is independent of known risk factors.

Patients with inflammatory bowel disease (IBD) have an increased risk of developing venous thromboembolic events, which have a considerable impact on morbidity and mortality. Chronic inflammation plays a crucial role in the pathogenesis of thrombotic events in patients with IBD. However, many unresolved questions remain, particularly regarding the mechanisms that determine the persistent inflammatory state independent of disease activity. This review explored the role of gut microbiota dysbiosis and intestinal barrier dysfunction, which are considered distinctive features of IBD, in determining pro-thrombotic tendencies. Gut-derived endotoxemia due to the translocation of bacterial lipopolysaccharides (LPS) from the intestine to the bloodstream and the bacterial metabolite trimethylamine-N-oxide (TMAO) are the most important molecules involved in gut dysbiosis-related thrombosis. The pathogenic prothrombotic pathways linked to LPS and TMAO have been discussed. Finally, we present emerging therapeutic approaches that can help reduce LPS-mediated endotoxemia and TMAO, such as restoring intestinal eubiosis, normalizing intestinal barrier function, and counterbalancing the effects of LPS and TMAO.

Background: Trimethylamine-N-oxide (TMAO) is a gut microbiota-derived metabolite produced from dietary nutrients. Many studies have discovered that circulating TMAO concentrations are linked to a wide range of health outcomes. Objectives: This study aimed to summarize health outcomes related to circulating TMAO concentrations. Methods: We searched the Embase. Medline, Web of Science, and Scopus databases from inception to 15 February, 2022 to identify and update meta-analyses examining the associations between 'TAO and multiple health outcomes. For each health outcome, we estimated the summary effect size. 95% prediction CI. between-study heterogeneity. evidence of small-study effects, and evidence of excess-significance bias. These metrics were used to evaluate the evidence credibility of the identified associations. Results: This umbrella review identified 24 meta-analyses that investigated the association between circulating 'TAO concentrations and health outcomes including all-cause mortality, cardiovascular diseases (CVDs), diabetes mellitus (DM), cancer. and renal function. We updated these meta-analyses by including a total of 82 individual studies on 18 unique health outcomes. Among them, 14 associations were nominally significant. After evidence credibility assessment, we found 6 (33%) associations (i.e., all-cause mortality, CVD mortality, major adverse cardiovascular events, hypertension. DM, and glomerular filtration rate) to present highly suggestive evidence. Conclusions: TMAO might be a novel biomarker related to human health conditions including all-cause mortality, hypertension. CVD, DM. cancer, and kidney function. Further studies are needed to investigate whether circulating 'MAO concentrations could be an intervention target for chronic disease.

Background: Trimethylamine-N-oxide (TMAO) is a gut microbiota-derived metabolite produced from dietary nutrients. Many studies have discovered that circulating TMAO concentrations are linked to a wide range of health outcomes. Objectives: This study aimed to summarize health outcomes related to circulating TMAO concentrations. Methods: We searched the Embase. Medline, Web of Science, and Scopus databases from inception to 15 February, 2022 to identify and update meta-analyses examining the associations between 'TAO and multiple health outcomes. For each health outcome, we estimated the summary effect size. 95% prediction CI. between-study heterogeneity. evidence of small-study effects, and evidence of excess-significance bias. These metrics were used to evaluate the evidence credibility of the identified associations. Results: This umbrella review identified 24 meta-analyses that investigated the association between circulating 'TAO concentrations and health outcomes including all-cause mortality, cardiovascular diseases (CVDs), diabetes mellitus (DM), cancer. and renal function. We updated these meta-analyses by including a total of 82 individual studies on 18 unique health outcomes. Among them, 14 associations were nominally significant. After evidence credibility assessment, we found 6 (33%) associations (i.e., all-cause mortality, CVD mortality, major adverse cardiovascular events, hypertension. DM, and glomerular filtration rate) to present highly suggestive evidence. Conclusions: TMAO might be a novel biomarker related to human health conditions including all-cause mortality, hypertension. CVD, DM. cancer, and kidney function. Further studies are needed to investigate whether circulating 'MAO concentrations could be an intervention target for chronic disease.

Background: Trimethylamine-N-oxide (TMAO) is a gut microbiota-derived metabolite produced from dietary nutrients. Many studies have discovered that circulating TMAO concentrations are linked to a wide range of health outcomes. Objectives: This study aimed to summarize health outcomes related to circulating TMAO concentrations. Methods: We searched the Embase. Medline, Web of Science, and Scopus databases from inception to 15 February, 2022 to identify and update meta-analyses examining the associations between 'TAO and multiple health outcomes. For each health outcome, we estimated the summary effect size. 95% prediction CI. between-study heterogeneity. evidence of small-study effects, and evidence of excess-significance bias. These metrics were used to evaluate the evidence credibility of the identified associations. Results: This umbrella review identified 24 meta-analyses that investigated the association between circulating 'TAO concentrations and health outcomes including all-cause mortality, cardiovascular diseases (CVDs), diabetes mellitus (DM), cancer. and renal function. We updated these meta-analyses by including a total of 82 individual studies on 18 unique health outcomes. Among them, 14 associations were nominally significant. After evidence credibility assessment, we found 6 (33%) associations (i.e., all-cause mortality, CVD mortality, major adverse cardiovascular events, hypertension. DM, and glomerular filtration rate) to present highly suggestive evidence. Conclusions: TMAO might be a novel biomarker related to human health conditions including all-cause mortality, hypertension. CVD, DM. cancer, and kidney function. Further studies are needed to investigate whether circulating 'MAO concentrations could be an intervention target for chronic disease.

Heart failure (HF) represents a cardiovascular disease with high mortality and morbidity. The latest evidence shows that changes in the composition of the gut microbiota might play a pivotal role in the prevention and management of HF. This systematic review aims at assessing the potential associations between the diet, gut microbiota, and derived metabolites with the outcomes of HF. A systematic literature search was performed up to July 2022 on the PubMed, Web of Science, and Scopus databases. The PRISMA guidelines were followed when possible. The risk of bias was assessed with the SYRCLE and ARRIVE tools. A total of nine pre-clinical studies on animal models, with considerable heterogeneity in dietary interventions, were included. High-fiber/prebiotic diets (n = 4) and a diet rich in polyphenols (n = 1) modified the gut microbiota composition and increased microbial metabolites' activities, linked with an improvement in HF outcomes, such as a reduction in systolic blood pressure, cardiac hypertrophy, and left ventricular thickness. A high-fat diet (n = 2) or a diet rich in choline (n = 2) induced an increase in TMAO and indole derivative production associated with a decrease in cardiac function, systemic endotoxemia, and inflammation and an increase in cardiac fibrosis and cardiac remodeling. Although results are retrieved from animal studies, this systematic review shows the key role of the diet-especially a high-fiber and prebiotic diet-on gut microbial metabolites in improving HF outcomes. Further studies on human cohorts are needed to identify personalized therapeutic dietary interventions to improve cardiometabolic health.

Trimethylamine-N-oxide (TMAO) is a uremic toxin, which has been associated with chronic kidney disease (CKD). Renal tubular epithelial cells play a central role in the pathophysiology of CKD. Megalin is an albumin-binding surface receptor on tubular epithelial cells, which is indispensable for urine protein reabsorption. To date, no studies have investigated the effect of TMAO on megalin expression and the functional properties of human tubular epithelial cells. The aim of this study was first to identify the functional effect of TMAO on human renal proximal tubular cells and second, to unravel the effects of TMAO on megalin-cubilin receptor expression. We found through global gene expression analysis that TMAO was associated with kidney disease. The microarray analysis also showed that megalin expression was suppressed by TMAO, which was also validated at the gene and protein level. High glucose and TMAO was shown to downregulate megalin expression and albumin uptake similarly. We also found that TMAO suppressed megalin expression via PI3K and ERK signaling. Furthermore, we showed that candesartan, dapagliflozin and enalaprilat counteracted the suppressive effect of TMAO on megalin expression. Our results may further help us unravel the role of TMAO in CKD development and to identify new therapeutic targets to counteract TMAOs effects., Funding agency:Faculty of Medicine and Health at Örebro University

Atherosclerotic cardiovascular disease (ASCVD) is a prime example of a systems disease. In the initial phase, apolipoprotein B-containing cholesterol-rich lipoproteins deposit excess cholesterol in macrophage-like cells that subsequently develop into foam cells. A multitude of systemic as well as environmental factors are involved in further progression of atherosclerotic plaque formation. In recent years, both oral and gut microbiota have been proposed to play an important role in the process at different stages. Particularly bacteria from the oral cavity may easily reach the circulation and cause low-grade inflammation, a recognized risk factor for ASCVD. Gut-derived microbiota on the other hand can influence host metabolism on various levels. Next to translocation across the intestinal wall, these prokaryotes produce a great number of specific metabolites such as trimethylamine and short-chain fatty acids but can also metabolize endogenously formed bile acids and convert these into metabolites that may influence signal transduction pathways. In this overview, we critically discuss the novel developments in this rapidly emerging research field.

Background: Trimethylamine-N-oxide (TMAO) is a gut microbiota-derived metabolite produced from dietary nutrients. Many studies have discovered that circulating TMAO concentrations are linked to a wide range of health outcomes. Objectives: This study aimed to summarize health outcomes related to circulating TMAO concentrations. Methods: We searched the Embase. Medline, Web of Science, and Scopus databases from inception to 15 February, 2022 to identify and update meta-analyses examining the associations between 'TAO and multiple health outcomes. For each health outcome, we estimated the summary effect size. 95% prediction CI. between-study heterogeneity. evidence of small-study effects, and evidence of excess-significance bias. These metrics were used to evaluate the evidence credibility of the identified associations. Results: This umbrella review identified 24 meta-analyses that investigated the association between circulating 'TAO concentrations and health outcomes including all-cause mortality, cardiovascular diseases (CVDs), diabetes mellitus (DM), cancer. and renal function. We updated these meta-analyses by including a total of 82 individual studies on 18 unique health outcomes. Among them, 14 associations were nominally significant. After evidence credibility assessment, we found 6 (33%) associations (i.e., all-cause mortality, CVD mortality, major adverse cardiovascular events, hypertension. DM, and glomerular filtration rate) to present highly suggestive evidence. Conclusions: TMAO might be a novel biomarker related to human health conditions including all-cause mortality, hypertension. CVD, DM. cancer, and kidney function. Further studies are needed to investigate whether circulating 'MAO concentrations could be an intervention target for chronic disease.

Background: Trimethylamine-N-oxide (TMAO) is a gut microbiota-derived metabolite produced from dietary nutrients. Many studies have discovered that circulating TMAO concentrations are linked to a wide range of health outcomes. Objectives: This study aimed to summarize health outcomes related to circulating TMAO concentrations. Methods: We searched the Embase. Medline, Web of Science, and Scopus databases from inception to 15 February, 2022 to identify and update meta-analyses examining the associations between 'TAO and multiple health outcomes. For each health outcome, we estimated the summary effect size. 95% prediction CI. between-study heterogeneity. evidence of small-study effects, and evidence of excess-significance bias. These metrics were used to evaluate the evidence credibility of the identified associations. Results: This umbrella review identified 24 meta-analyses that investigated the association between circulating 'TAO concentrations and health outcomes including all-cause mortality, cardiovascular diseases (CVDs), diabetes mellitus (DM), cancer. and renal function. We updated these meta-analyses by including a total of 82 individual studies on 18 unique health outcomes. Among them, 14 associations were nominally significant. After evidence credibility assessment, we found 6 (33%) associations (i.e., all-cause mortality, CVD mortality, major adverse cardiovascular events, hypertension. DM, and glomerular filtration rate) to present highly suggestive evidence. Conclusions: TMAO might be a novel biomarker related to human health conditions including all-cause mortality, hypertension. CVD, DM. cancer, and kidney function. Further studies are needed to investigate whether circulating 'MAO concentrations could be an intervention target for chronic disease.

A Mediterranean-style eating pattern (MED-EP) may include moderate red meat intake. However, it is unknown if the pro-atherogenic metabolite trimethylamine N-oxide (TMAO) is affected by the amount of red meat consumed with a MED-EP. The results presented are from a secondary, retrospective objective of an investigator-blinded, randomized, crossover, controlled feeding trial (two 5-wk interventions separated by a 4-wk washout) to determine if a MED-EP with 200g unprocessed lean red meat/wk (MED-CONTROL) reduces circulating TMAO concentrations compared to a MED-EP with 500g unprocessed lean red meat/wk (MED-RED). Participants were 27 women and 12 men (n=39 total) who were either overweight or obese (BMI: 30.5 ± 0.3 kg/m2 mean ± SEM). Serum samples were obtained following an overnight fast both before (pre) and after (post) each intervention. Fasting serum TMAO, choline, carnitine, and betaine concentrations were measured using a targeted Liquid chromatography-mass spectrometry. Data were analyzed to assess if (a) TMAO and related metabolites differed by intervention, and (b) if changes in TMAO were associated with changes in Framingham 10-year risk score. Serum TMAO was lower post-intervention following MED-CONTROL compared to MED-RED intervention (post-MED-CONTROL 3.1 ± 0.2 µM vs. post-MED-RED 5.0 ± 0.5 µM, p<0.001), and decreased following MED-CONTROL (pre- vs post-MED-CONTROL, p = 0.025). Exploratory analysis using mixed model analysis of covariance identified a positive association between changes in TMAO and changes in HOMA-IR (p = 0.036). These results suggest that lower amounts of red meat intake leads to lower TMAO concentrations in the context of a MED-EP.

AIMS: Trimethylamine N-oxide (TMAO), choline and betaine serum levels have been associated with metabolic diseases including type 2 diabetes (T2D) and non-alcoholic fatty liver disease (NAFLD). These associations could be mediated by insulin resistance. However, the relationships among these metabolites, insulin resistance and NAFLD have not been thoroughly investigated. Moreover, it has recently been suggested that TMAO could play a role in NAFLD by altering bile acid metabolism. We examined the association between circulating TMAO, choline and betaine levels and NAFLD in obese subjects. METHODS: Serum TMAO, choline, betaine and bile acid levels were measured in 357 Mexican obese patients with different grades of NAFLD as determined by liver histology. Associations of NAFLD with TMAO, choline and betaine levels were tested. Moreover, association of TMAO levels with non-alcoholic steatohepatitis (NASH) was tested separately in patients with and without T2D. RESULTS: TMAO and choline levels were significantly associated with NAFLD histologic features and NASH risk. While increased serum TMAO levels were significantly associated with NASH in patients with T2D, in non-T2D subjects this association lost significance after adjusting for sex, BMI and HOMA2-IR. Moreover, circulating secondary bile acids were associated both with increased TMAO levels and NASH. CONCLUSIONS: In obese patients, circulating TMAO levels were associated with NASH mainly in the presence of T2D. Functional studies are required to evaluate the role of insulin resistance and T2D in this association, both highly prevalent in NASH patients.

Trimethylamine-N-oxide (TMAO), a microbiome-derived metabolite from the metabolism of choline, betaine, and carnitines, is associated to adverse cardiovascular outcomes. A method suitable for routine quantification of TMAO and its precursors (trimethylamine (TMA), choline, betaine, creatinine, and propionyl-, acetyl-, and l-carnitine) in clinical and food samples has been developed based on LC-MS. TMA was successfully derivatized using iodoacetonitrile, and no cross-reactions with TMAO or the other methylamines were detected. Extraction from clinical samples (plasma and urine) was performed after protein precipitation using acetonitrile:methanol. For food samples (meatballs and eggs), water extraction was shown to be sufficient, but acid hydrolysis was required to release bound choline before extraction. Baseline separation of the methylamines was achieved using a neutral HILIC column and a mobile phase consisting of 25 mmol/L ammonium formate in water:ACN (30:70). Quantification was performed by MS using external calibration and isotopic labelled internal standards. The assay proved suitable for both clinical and food samples and was linear from approximate to 0.1 up to 200 mu mol/L for all methylamines except for TMA and TMAO, which were linear up to 100 mu mol/L. Recoveries were 91-107% in clinical samples and 76-98% in food samples. The interday (n=8, four duplicate analysis) CVs were below 9% for all metabolites in clinical and food samples. The method was applied successfully to determine the methylamine concentrations in plasma and urine from the subjects participating in an intervention trial (n=10) to determine the effect of animal food ingestion on methylamine concentrations.

Bacteria can colonize virtually any environment on Earth due to their remarkable capacity to detect and respond quickly and adequately to environmental stressors. Vibrio cholerae is a cosmopolitan bacterium that inhabits a vast range of environments. The V. cholerae life cycle comprises diverse environmental and infective stages. The bacterium is found in aquatic ecosystems both under free-living conditions or associated with a wide range of aquatic organisms, and some strains are also capable of causing epidemics in humans. In order to adapt between environments, V. cholerae possesses a versatile metabolism characterized by the rapid cross-regulation of energy-producing pathways. Low oxygen concentration is a key environmental factor that governs V. cholerae physiology. This article reviews the metabolic plasticity that enables V. cholerae to thrive on low oxygen concentrations and its role in environmental and host adaptation.

Bacteria can colonize virtually any environment on Earth due to their remarkable capacity to detect and respond quickly and adequately to environmental stressors. Vibrio cholerae is a cosmopolitan bacterium that inhabits a vast range of environments. The V. cholerae life cycle comprises diverse environmental and infective stages. The bacterium is found in aquatic ecosystems both under free-living conditions or associated with a wide range of aquatic organisms, and some strains are also capable of causing epidemics in humans. In order to adapt between environments, V. cholerae possesses a versatile metabolism characterized by the rapid cross-regulation of energy-producing pathways. Low oxygen concentration is a key environmental factor that governs V. cholerae physiology. This article reviews the metabolic plasticity that enables V. cholerae to thrive on low oxygen concentrations and its role in environmental and host adaptation.

As a crucial part of the symbiotic system, the gut microbiome is metabolically connected to many diseases and conditions, including cardiovascular diseases (CVD). Trimethylamine (TMA) is produced by gut bacteria from dietary choline, betaine, or L-carnitine, and is then converted in the liver to Trimethylamine N-oxide (TMAO), which in turn affects hepatic and intestinal lipid metabolism. Circulating TMAO is positively associated with CVD risk. Because eggs are rich in choline, it has been speculated that their consumption may increase plasma TMAO. In this study, we hypothesized that 2 eggs per day increases plasma TMAO level by altering gut microbiome composition in mildly hypercholesterolemic postmenopausal women. In this randomized, cross-over study, 20 overweight, postmenopausal women were given 2 whole eggs and the equivalent amount of yolk-free substitute as breakfast for 4 weeks, in randomized order, with a 4-week washout in between. Fasting blood draws and stool were collected at the beginning and end of each treatment period. Plasma TMAO, choline, betaine and other metabolites were analyzed using LC/MS, while gut microbiome composition was analyzed using 16S amplicon sequencing. Plasma choline and betaine were significantly increased after whole egg but not yolk-free substitute, however TMAO level was not significantly affected by treatments. Gut microbiome composition showed large inter-individual variability at baseline and in response to the treatments. The consumption of 2 eggs per day in overweight, postmenopausal mildly hypercholesterolemic women significantly increased plasma choline and betaine, but did not increase plasma TMAO or alter gut microbiome composition.

© 2019, The Author(s). Background: The Paleolithic diet is promoted worldwide for improved gut health. However, there is little evidence available to support these claims, with existing literature examining anthropometric and cardiometabolic outcomes. Objective: To determine the association between dietary intake, markers of colonic health, microbiota, and serum trimethylamine-N-oxide (TMAO), a gut-derived metabolite associated with cardiovascular disease. Design: In a cross-sectional design, long-term (n = 44, > 1 year) self-reported followers of a Paleolithic diet (PD) and controls (n = 47) consuming a diet typical of national recommendations were recruited. Diets were assessed via 3-day weighed diet records; 48-h stool for short chain fatty acids using GC/MS, microbial composition via 16S rRNA sequencing of the V4 region using Illumina MiSeq. TMAO was quantified using LC–MS/MS. Results: Participants were grouped according to PD adherence; namely excluding grains and dairy products. Strict Paleolithic (SP) (n = 22) and Pseudo-Paleolithic (PP) (n = 22) groups were formed. General linear modelling with age, gender, energy intake and body fat percentage as covariates assessed differences between groups. Intake of resistant starch was lower in both Paleolithic groups, compared to controls [2.62, 1.26 vs 4.48 g/day (P < 0.05)]; PERMANOVA analysis showed differences in microbiota composition (P < 0.05), with higher abundance of TMA-producer Hungatella in both Paleolithic groups (P < 0.001). TMAO was higher in SP compared to PP and control (P < 0.01), and inversely associated with whole grain intake (r = − 0.34, P < 0.01). Conclusions: Although the PD is promoted for improved gut health, results indicate long-term adherence is associated with different gut microbiota and increased TMAO. A variety of fiber components, including whole grain sources may be required to maintain gut and cardiovascular health. Clinical trial registrations: Australian and New Zealand Clinical T

© 2019, The Author(s). Background: The Paleolithic diet is promoted worldwide for improved gut health. However, there is little evidence available to support these claims, with existing literature examining anthropometric and cardiometabolic outcomes. Objective: To determine the association between dietary intake, markers of colonic health, microbiota, and serum trimethylamine-N-oxide (TMAO), a gut-derived metabolite associated with cardiovascular disease. Design: In a cross-sectional design, long-term (n = 44, > 1 year) self-reported followers of a Paleolithic diet (PD) and controls (n = 47) consuming a diet typical of national recommendations were recruited. Diets were assessed via 3-day weighed diet records; 48-h stool for short chain fatty acids using GC/MS, microbial composition via 16S rRNA sequencing of the V4 region using Illumina MiSeq. TMAO was quantified using LC–MS/MS. Results: Participants were grouped according to PD adherence; namely excluding grains and dairy products. Strict Paleolithic (SP) (n = 22) and Pseudo-Paleolithic (PP) (n = 22) groups were formed. General linear modelling with age, gender, energy intake and body fat percentage as covariates assessed differences between groups. Intake of resistant starch was lower in both Paleolithic groups, compared to controls [2.62, 1.26 vs 4.48 g/day (P < 0.05)]; PERMANOVA analysis showed differences in microbiota composition (P < 0.05), with higher abundance of TMA-producer Hungatella in both Paleolithic groups (P < 0.001). TMAO was higher in SP compared to PP and control (P < 0.01), and inversely associated with whole grain intake (r = − 0.34, P < 0.01). Conclusions: Although the PD is promoted for improved gut health, results indicate long-term adherence is associated with different gut microbiota and increased TMAO. A variety of fiber components, including whole grain sources may be required to maintain gut and cardiovascular health. Clinical trial registrations: Australian and New Zealand Clinical T

© 2019, The Author(s). Background: The Paleolithic diet is promoted worldwide for improved gut health. However, there is little evidence available to support these claims, with existing literature examining anthropometric and cardiometabolic outcomes. Objective: To determine the association between dietary intake, markers of colonic health, microbiota, and serum trimethylamine-N-oxide (TMAO), a gut-derived metabolite associated with cardiovascular disease. Design: In a cross-sectional design, long-term (n = 44, > 1 year) self-reported followers of a Paleolithic diet (PD) and controls (n = 47) consuming a diet typical of national recommendations were recruited. Diets were assessed via 3-day weighed diet records; 48-h stool for short chain fatty acids using GC/MS, microbial composition via 16S rRNA sequencing of the V4 region using Illumina MiSeq. TMAO was quantified using LC–MS/MS. Results: Participants were grouped according to PD adherence; namely excluding grains and dairy products. Strict Paleolithic (SP) (n = 22) and Pseudo-Paleolithic (PP) (n = 22) groups were formed. General linear modelling with age, gender, energy intake and body fat percentage as covariates assessed differences between groups. Intake of resistant starch was lower in both Paleolithic groups, compared to controls [2.62, 1.26 vs 4.48 g/day (P < 0.05)]; PERMANOVA analysis showed differences in microbiota composition (P < 0.05), with higher abundance of TMA-producer Hungatella in both Paleolithic groups (P < 0.001). TMAO was higher in SP compared to PP and control (P < 0.01), and inversely associated with whole grain intake (r = − 0.34, P < 0.01). Conclusions: Although the PD is promoted for improved gut health, results indicate long-term adherence is associated with different gut microbiota and increased TMAO. A variety of fiber components, including whole grain sources may be required to maintain gut and cardiovascular health. Clinical trial registrations: Australian and New Zealand Clinical T

As a crucial part of the symbiotic system, the gut microbiome is metabolically connected to many diseases and conditions, including cardiovascular diseases (CVD). Trimethylamine (TMA) is produced by gut bacteria from dietary choline, betaine, or L-carnitine, and is then converted in the liver to Trimethylamine N-oxide (TMAO), which in turn affects hepatic and intestinal lipid metabolism. Circulating TMAO is positively associated with CVD risk. Because eggs are rich in choline, it has been speculated that their consumption may increase plasma TMAO. In this study, we hypothesized that 2 eggs per day increases plasma TMAO level by altering gut microbiome composition in mildly hypercholesterolemic postmenopausal women. In this randomized, cross-over study, 20 overweight, postmenopausal women were given 2 whole eggs and the equivalent amount of yolk-free substitute as breakfast for 4 weeks, in randomized order, with a 4-week washout in between. Fasting blood draws and stool were collected at the beginning and end of each treatment period. Plasma TMAO, choline, betaine and other metabolites were analyzed using LC/MS, while gut microbiome composition was analyzed using 16S amplicon sequencing. Plasma choline and betaine were significantly increased after whole egg but not yolk-free substitute, however TMAO level was not significantly affected by treatments. Gut microbiome composition showed large inter-individual variability at baseline and in response to the treatments. The consumption of 2 eggs per day in overweight, postmenopausal mildly hypercholesterolemic women significantly increased plasma choline and betaine, but did not increase plasma TMAO or alter gut microbiome composition.

Nanoscale transportation in engineered environments is critical towards designing efficient and smart hybrid bio-nanodevices. Biomolecular motors, the smallest natural machines, are promising as actuators as well as sensors in hybrid nanodevices and hold enormous potentials in nanoscale transportation. Highly specific regulation of the activity of biomolecular motors is the key to control such integrated nanodevices. We present a simple method to regulate the activity of a biomolecular motor system, microtubule (MT)-kinesin by using a natural osmolyte trimethylamine-N-oxide (TMAO). Motility of kinesin-driven MTs in an in vitro gliding assay is regulated over a broad spectrum by using TMAO in a concentration dependent manner. The regulation of MT motility is rapid, reversible and repeatable over multiple cycles. Interestingly, the motility of MTs can be completely turned off using TMAO of a relatively high concentration. The halted motility of MTs is fully restored upon elimination of TMAO. Repeated cycles of TMAO addition and removal enable cyclical inhibition and restoration of the motility of MTs. These results demonstrate an ability to control nanoscale motion of a biomolecular motor in an artificial environment. This work facilitates further tunability over functions of biomolecular motors, which in turn will foster their nanotechnological applications, such as in nano-transportation.

Nanoscale transportation in engineered environments is critical towards designing efficient and smart hybrid bio-nanodevices. Biomolecular motors, the smallest natural machines, are promising as actuators as well as sensors in hybrid nanodevices and hold enormous potentials in nanoscale transportation. Highly specific regulation of the activity of biomolecular motors is the key to control such integrated nanodevices. We present a simple method to regulate the activity of a biomolecular motor system, microtubule (MT)-kinesin by using a natural osmolyte trimethylamine-N-oxide (TMAO). Motility of kinesin-driven MTs in an in vitro gliding assay is regulated over a broad spectrum by using TMAO in a concentration dependent manner. The regulation of MT motility is rapid, reversible and repeatable over multiple cycles. Interestingly, the motility of MTs can be completely turned off using TMAO of a relatively high concentration. The halted motility of MTs is fully restored upon elimination of TMAO. Repeated cycles of TMAO addition and removal enable cyclical inhibition and restoration of the motility of MTs. These results demonstrate an ability to control nanoscale motion of a biomolecular motor in an artificial environment. This work facilitates further tunability over functions of biomolecular motors, which in turn will foster their nanotechnological applications, such as in nano-transportation.

© 2019, The Author(s). Background: The Paleolithic diet is promoted worldwide for improved gut health. However, there is little evidence available to support these claims, with existing literature examining anthropometric and cardiometabolic outcomes. Objective: To determine the association between dietary intake, markers of colonic health, microbiota, and serum trimethylamine-N-oxide (TMAO), a gut-derived metabolite associated with cardiovascular disease. Design: In a cross-sectional design, long-term (n = 44, > 1 year) self-reported followers of a Paleolithic diet (PD) and controls (n = 47) consuming a diet typical of national recommendations were recruited. Diets were assessed via 3-day weighed diet records; 48-h stool for short chain fatty acids using GC/MS, microbial composition via 16S rRNA sequencing of the V4 region using Illumina MiSeq. TMAO was quantified using LC–MS/MS. Results: Participants were grouped according to PD adherence; namely excluding grains and dairy products. Strict Paleolithic (SP) (n = 22) and Pseudo-Paleolithic (PP) (n = 22) groups were formed. General linear modelling with age, gender, energy intake and body fat percentage as covariates assessed differences between groups. Intake of resistant starch was lower in both Paleolithic groups, compared to controls [2.62, 1.26 vs 4.48 g/day (P < 0.05)]; PERMANOVA analysis showed differences in microbiota composition (P < 0.05), with higher abundance of TMA-producer Hungatella in both Paleolithic groups (P < 0.001). TMAO was higher in SP compared to PP and control (P < 0.01), and inversely associated with whole grain intake (r = − 0.34, P < 0.01). Conclusions: Although the PD is promoted for improved gut health, results indicate long-term adherence is associated with different gut microbiota and increased TMAO. A variety of fiber components, including whole grain sources may be required to maintain gut and cardiovascular health. Clinical trial registrations: Australian and New Zealand Clinical T

Bacteria can colonize virtually any environment on Earth due to their remarkable capacity to detect and respond quickly and adequately to environmental stressors. Vibrio cholerae is a cosmopolitan bacterium that inhabits a vast range of environments. The V. cholerae life cycle comprises diverse environmental and infective stages. The bacterium is found in aquatic ecosystems both under free-living conditions or associated with a wide range of aquatic organisms, and some strains are also capable of causing epidemics in humans. In order to adapt between environments, V. cholerae possesses a versatile metabolism characterized by the rapid cross-regulation of energy-producing pathways. Low oxygen concentration is a key environmental factor that governs V. cholerae physiology. This article reviews the metabolic plasticity that enables V. cholerae to thrive on low oxygen concentrations and its role in environmental and host adaptation.

Bacteria can colonize virtually any environment on Earth due to their remarkable capacity to detect and respond quickly and adequately to environmental stressors. Vibrio cholerae is a cosmopolitan bacterium that inhabits a vast range of environments. The V. cholerae life cycle comprises diverse environmental and infective stages. The bacterium is found in aquatic ecosystems both under free-living conditions or associated with a wide range of aquatic organisms, and some strains are also capable of causing epidemics in humans. In order to adapt between environments, V. cholerae possesses a versatile metabolism characterized by the rapid cross-regulation of energy-producing pathways. Low oxygen concentration is a key environmental factor that governs V. cholerae physiology. This article reviews the metabolic plasticity that enables V. cholerae to thrive on low oxygen concentrations and its role in environmental and host adaptation.

Choline is a water-soluble nutrient essential for human life. Gut microbial metabolism of choline results in the production of trimethylamine (TMA), which, upon absorption by the host is converted into trimethylamine-N-oxide (TMAO) in the liver. A high accumulation of both components is related to cardiovascular disease, inflammatory bowel disease, non-alcoholic fatty liver disease, and chronic kidney disease. However, the relationship between the microbiota production of these components and its impact on these diseases still remains unknown. In this review, we will address which microbes contribute to TMA production in the human gut, the extent to which host factors (e.g., the genotype) and diet affect TMA production, and the colonization of these microbes and the reversal of dysbiosis as a therapy for these diseases.

Bacteria can colonize virtually any environment on Earth due to their remarkable capacity to detect and respond quickly and adequately to environmental stressors. Vibrio cholerae is a cosmopolitan bacterium that inhabits a vast range of environments. The V. cholerae life cycle comprises diverse environmental and infective stages. The bacterium is found in aquatic ecosystems both under free-living conditions or associated with a wide range of aquatic organisms, and some strains are also capable of causing epidemics in humans. In order to adapt between environments, V. cholerae possesses a versatile metabolism characterized by the rapid cross-regulation of energy-producing pathways. Low oxygen concentration is a key environmental factor that governs V. cholerae physiology. This article reviews the metabolic plasticity that enables V. cholerae to thrive on low oxygen concentrations and its role in environmental and host adaptation.

© 2019, The Author(s). Background: The Paleolithic diet is promoted worldwide for improved gut health. However, there is little evidence available to support these claims, with existing literature examining anthropometric and cardiometabolic outcomes. Objective: To determine the association between dietary intake, markers of colonic health, microbiota, and serum trimethylamine-N-oxide (TMAO), a gut-derived metabolite associated with cardiovascular disease. Design: In a cross-sectional design, long-term (n = 44, > 1 year) self-reported followers of a Paleolithic diet (PD) and controls (n = 47) consuming a diet typical of national recommendations were recruited. Diets were assessed via 3-day weighed diet records; 48-h stool for short chain fatty acids using GC/MS, microbial composition via 16S rRNA sequencing of the V4 region using Illumina MiSeq. TMAO was quantified using LC–MS/MS. Results: Participants were grouped according to PD adherence; namely excluding grains and dairy products. Strict Paleolithic (SP) (n = 22) and Pseudo-Paleolithic (PP) (n = 22) groups were formed. General linear modelling with age, gender, energy intake and body fat percentage as covariates assessed differences between groups. Intake of resistant starch was lower in both Paleolithic groups, compared to controls [2.62, 1.26 vs 4.48 g/day (P < 0.05)]; PERMANOVA analysis showed differences in microbiota composition (P < 0.05), with higher abundance of TMA-producer Hungatella in both Paleolithic groups (P < 0.001). TMAO was higher in SP compared to PP and control (P < 0.01), and inversely associated with whole grain intake (r = − 0.34, P < 0.01). Conclusions: Although the PD is promoted for improved gut health, results indicate long-term adherence is associated with different gut microbiota and increased TMAO. A variety of fiber components, including whole grain sources may be required to maintain gut and cardiovascular health. Clinical trial registrations: Australian and New Zealand Clinical T

BackgroundAcute Coronary Syndrome (ACS) is a leading cause of morbidity and mortality. Perturbed gut- microbiota (dysbiosis) and increased intestinal permeability (leaky-gut) with translocation of bacterial antigens, play critical role in obesity and metabolic syndrome, which are also major ACS risk factors. Additionally, Trimethylamine-N-Oxide (TMAO), a metabolite produced by phylum Proteobacteria in gut is implicated in developing ACS. As Proteobacteria is a major source of translocated antigen lipopolysaccharides (LPS), we hypothesized that ACS patients have leaky-gut condition characterized by dysbiosis with increased Proteobacteria, leading to elevated blood levels of TMAO and LPS.MethodsIn a pilot case-control study, we enrolled 19 ACS patients (within 72-h of cardiac events) and 19 healthy-controls. Gut barrier function was determined using lactulose-to-mannitol urinary excretion ratio (L/M ratio). Stool microbiome composition was examined using16S sequencing and predictive functional analysis for LPS biosynthesis pathway by PICRUSt tool. Serum TMAO and LPS levels were measured.ResultsACS patients had increased Gammaproteobacteria compared to controls:1.8 ±3.0 vs. 0.2 ±0.4% (P =0.04). Though Proteobacteria level was increased but not statistically significant: 4.1 ±3.8 vs. 2.1 ±1.7% (P =0.056). L/M-ratio was three times higher in ACS patients; 0.06 ±0.07 vs 0.023 ±0.02, (P =0.014). Surprisingly, there was no difference in the mean serum LPS or TMAO levels. However, PICRUSt analysis indicated increased Proteobacteria population increasingly contributed to LPS biosynthesis in ACS patients only.ConclusionsACS patients likely to have leaky-gut and perturbed gut microbiota. Further studies are required to precisely define the role of dysbiosis in ACS.

Scope: The impact of dietary protein types on the gut microbiome is scarcely studied. The aim of the present study is therefore to examine the effects of lean-seafood and non-seafood proteins on the gut microbiome composition and activity and elucidate potential associations to cardiovascular disease (CVD) risk factors. Methods: A crossover intervention study in which 20 healthy subjects consumed two diets that varied in protein source was conducted. 1H NMR spectroscopy and 16S rDNA sequencing analyses were applied to characterize fecal metabolites and gut microbiota composition, respectively. Results: A twofold increase in fecal trimethylamine excretion was observed after the lean-seafood diet period. Circulating TAG and the total to high-density lipoprotein (HDL) cholesterol ratio as well as circulating TMAO levels were each associated with specific gut bacteria. Following the non-seafood diet period, a decreased relative abundance of Clostridium cluster IV and a tendency toward an increased Firmicutes/Bacteroidetes ratio were found. Conclusions: Lean-seafood and non-seafood diets differentially modulate the gut microbiome composition and activity. Furthermore, the gut microbiota composition seems to affect circulating TMAO levels and CVD risk factors.

Higher dietary protein intake is increasingly recommended for the elderly; however, high protein diets have also been linked to increased cardiovascular disease (CVD) risk. Trimethylamine-N-oxide (TMAO) is a bacterial metabolite derived from choline and carnitine abundant from animal protein-rich foods. TMAO may be a novel biomarker for heightened CVD risk. The purpose of this study was to assess the impact of a high protein diet on TMAO. Healthy men (74.2 ± 3.6 years, n = 29) were randomised to consume the recommended dietary allowance of protein (RDA: 0.8 g protein/kg bodyweight/day) or twice the RDA (2RDA) as part of a supplied diet for 10 weeks. Fasting blood samples were collected pre- and post-intervention for measurement of TMAO, blood lipids, glucose tolerance, insulin sensitivity, and inflammatory biomarkers. An oral glucose tolerance test was also performed. In comparison with RDA, the 2RDA diet increased circulatory TMAO (p = 0.002) but unexpectedly decreased renal excretion of TMAO (p = 0.003). LDL cholesterol was increased in 2RDA compared to RDA (p = 0.049), but no differences in other biomarkers of CVD risk and insulin sensitivity were evident between groups. In conclusion, circulatory TMAO is responsive to changes in dietary protein intake in older healthy males.

Higher dietary protein intake is increasingly recommended for the elderly; however, high protein diets have also been linked to increased cardiovascular disease (CVD) risk. Trimethylamine-N-oxide (TMAO) is a bacterial metabolite derived from choline and carnitine abundant from animal protein-rich foods. TMAO may be a novel biomarker for heightened CVD risk. The purpose of this study was to assess the impact of a high protein diet on TMAO. Healthy men (74.2 ± 3.6 years, n = 29) were randomised to consume the recommended dietary allowance of protein (RDA: 0.8 g protein/kg bodyweight/day) or twice the RDA (2RDA) as part of a supplied diet for 10 weeks. Fasting blood samples were collected pre- and post-intervention for measurement of TMAO, blood lipids, glucose tolerance, insulin sensitivity, and inflammatory biomarkers. An oral glucose tolerance test was also performed. In comparison with RDA, the 2RDA diet increased circulatory TMAO (p = 0.002) but unexpectedly decreased renal excretion of TMAO (p = 0.003). LDL cholesterol was increased in 2RDA compared to RDA (p = 0.049), but no differences in other biomarkers of CVD risk and insulin sensitivity were evident between groups. In conclusion, circulatory TMAO is responsive to changes in dietary protein intake in older healthy males.

Trimethylamine-n-oxide (TMAO) is a metabolite found in plasma/serum in humans. Elevated levels of TMAO have been associated with several types of heart disease. It’s therefore of interest to make a simple analytical method to analyse TMAO and other metabolites that are degraded to TMAO, including betaine. In this study, the goal was to develop a method for the sample preparation and analysis of these compounds in human plasma. Sample preparation was performed with an Ostro 96-well method for sample clean-up. The analysis was performed by ultraperformance liquid chromatography – hydrophilic interaction liquid chromatography – tandem masspectrometry (UPLC-HILIC-MS/MS) in multiple reaction monitoring (MRM)-mode using electrospray ionization in positive mode (ESI+)-mode as the ion source. The analytes eluted under five minutes and were all baseline separated in the chromatogram. TMAO and betaine were quantified in quality control (QC) plasma samples using external calibration. Concentration of TMAO ranged from 132 ng/mL – 253 ng/mL and 1025-2084 ng/mL for betaine. Due to the lack of isotopically labelled standards for TMAO and betaine, valine-d8 was tested as an internal standard for the extraction; however, it was not a suitable option due to the low recovery obtained (5-34%) and the low response in ESI+. The recovery needs to be investigated further using isotopically labelled TMAO or betaine. Overall, the developed UPLC-HILIC-MS/MS method was found to be suitable for analysis of TMAO and betaine in human plasma. Further development and validation is required before application to samples from clinical studies.

Background: Acute Coronary Syndrome (ACS) is a leading cause of morbidity and mortality. Metabolic syndrome and obesity are major risk factors for atherosclerosis and ACS. Dysbiosis plays an important role in metabolic syndrome and obesity. Studies show a markedly increased risk of heart attacks in patients with high levels of the pro-atherogenic metabolite trimethylamine-N-Oxide (TMAO). TMAO is produced by the intestinal microbial flora through metabolism of dietary phospholipids; Gram-negative bacteria (Phylum Proteobacteria) is the major source of TMAO metabolism. Patients with obesity and metabolic syndrome have a defective intestinal tight-junctional (TJ) barrier, which allows paracellular permeation of luminal antigens such as lipopolysaccharides (LPS; endotoxins); leading to endotoxemia. Endotoxins are also associated with increased risk of atherosclerosis and cardiovascular disease. We aim to study the hypothesis that patients with ACS have dysbiosis, including higher proportion of Gram-negative bacterial species capable of vi producing the pro-atherogenic metabolite TMAO, and test if dysbiosis in ACS patients is associated with increased intestinal permeability and endotoxemia. Methods: This is a prospective case-control study conducted at a single university hospital. We enrolled ACS patients (within 72 hours of acute cardiac events) and healthy controls. Relevant clinical and demographic data were collected. Stool microbiome composition was examined using the 16S ribosomal RNA (Illumina) method to identify microbiota taxonomic genera. LPS serum level was measured by an ELISA-based method. The intestinal TJ barrier function was evaluated using lactulose-to-mannitol urinary excretion ratio (L/M ratio). Serum TMAO was measured using a mass spectrometry. Results: We enrolled 19 patients and 19 controls. ACS patients had increased relative abundance of Gammaproteobacteria and Proteobacteria compared to healthy controls, with mean proportions o

Background: Acute Coronary Syndrome (ACS) is a leading cause of morbidity and mortality. Metabolic syndrome and obesity are major risk factors for atherosclerosis and ACS. Dysbiosis plays an important role in metabolic syndrome and obesity. Studies show a markedly increased risk of heart attacks in patients with high levels of the pro-atherogenic metabolite trimethylamine-N-Oxide (TMAO). TMAO is produced by the intestinal microbial flora through metabolism of dietary phospholipids; Gram-negative bacteria (Phylum Proteobacteria) is the major source of TMAO metabolism. Patients with obesity and metabolic syndrome have a defective intestinal tight-junctional (TJ) barrier, which allows paracellular permeation of luminal antigens such as lipopolysaccharides (LPS; endotoxins); leading to endotoxemia. Endotoxins are also associated with increased risk of atherosclerosis and cardiovascular disease. We aim to study the hypothesis that patients with ACS have dysbiosis, including higher proportion of Gram-negative bacterial species capable of vi producing the pro-atherogenic metabolite TMAO, and test if dysbiosis in ACS patients is associated with increased intestinal permeability and endotoxemia. Methods: This is a prospective case-control study conducted at a single university hospital. We enrolled ACS patients (within 72 hours of acute cardiac events) and healthy controls. Relevant clinical and demographic data were collected. Stool microbiome composition was examined using the 16S ribosomal RNA (Illumina) method to identify microbiota taxonomic genera. LPS serum level was measured by an ELISA-based method. The intestinal TJ barrier function was evaluated using lactulose-to-mannitol urinary excretion ratio (L/M ratio). Serum TMAO was measured using a mass spectrometry. Results: We enrolled 19 patients and 19 controls. ACS patients had increased relative abundance of Gammaproteobacteria and Proteobacteria compared to healthy controls, with mean proportions o

Trimethylamine-n-oxide (TMAO) is a metabolite found in plasma/serum in humans. Elevated levels of TMAO have been associated with several types of heart disease. It’s therefore of interest to make a simple analytical method to analyse TMAO and other metabolites that are degraded to TMAO, including betaine. In this study, the goal was to develop a method for the sample preparation and analysis of these compounds in human plasma. Sample preparation was performed with an Ostro 96-well method for sample clean-up. The analysis was performed by ultraperformance liquid chromatography – hydrophilic interaction liquid chromatography – tandem masspectrometry (UPLC-HILIC-MS/MS) in multiple reaction monitoring (MRM)-mode using electrospray ionization in positive mode (ESI+)-mode as the ion source. The analytes eluted under five minutes and were all baseline separated in the chromatogram. TMAO and betaine were quantified in quality control (QC) plasma samples using external calibration. Concentration of TMAO ranged from 132 ng/mL – 253 ng/mL and 1025-2084 ng/mL for betaine. Due to the lack of isotopically labelled standards for TMAO and betaine, valine-d8 was tested as an internal standard for the extraction; however, it was not a suitable option due to the low recovery obtained (5-34%) and the low response in ESI+. The recovery needs to be investigated further using isotopically labelled TMAO or betaine. Overall, the developed UPLC-HILIC-MS/MS method was found to be suitable for analysis of TMAO and betaine in human plasma. Further development and validation is required before application to samples from clinical studies.

This thesis explores lipid self-assembly and aims to give a broad picture of self-assembly structures in simple and complex lipid-protein systems. The systems studied are lung surfactant, stratum corneum and simple model membranes. The lung surfactant mixture lines the alveolus in our lungs and stabilises the air- tissue interface. The lung surfactant lipid phase behaviour was here investigated with respect to the effects of cholesterol concentration and changes in the external conditions of temperature and a water gradient. Taken together the studies of lung surfactant give a comprehensive picture of the phase behaviour of a clinical lung surfactant extract, showing the importance of cholesterol and non-equilibrium conditions. It is a recurrent observation for all studies that the addition of physiologically relevant levels of cholesterol to the clinical lung surfactant forms a single robust liquid ordered phase under both equilibrium and non- equilibrium conditions.Conditions such as draught, high salinity or freezing, exerts the lipid systems to osmotic stress, which can lead to phase changes between different self-assembled structures. The outer layer of the skin, the stratum corneum is most of the time exposed to osmotic stress from dry air in the environment. The stratum corneum contains small polar molecules, to counteract phase changes due to osmotic stress. We study how the self-assembly in stratum corneum and model membranes are affected by the presence of osmolytes under conditions of osmotic stress. It is shown that these compounds under dry conditions act to replace the water in both stratum corneum and model membranes and they may stabilize the fluid lipid phases at lower humidities.In plants, there are several strategies for protection against osmotic stress. One strategy involves the expression of specific proteins. We have studied how one such protein from the family of dehydrins, influences lipid self-assembly, aiming at molecular understanding of

BackgroundThe gut microbiome is a complex and metabolically active community that directly influences host phenotypes. In this study, we profile gut microbiota using 16S rRNA gene sequencing in 531 well-phenotyped Finnish men from the Metabolic Syndrome In Men (METSIM) study.ResultsWe investigate gut microbiota relationships with a variety of factors that have an impact on the development of metabolic and cardiovascular traits. We identify novel associations between gut microbiota and fasting serum levels of a number of metabolites, including fatty acids, amino acids, lipids, and glucose. In particular, we detect associations with fasting plasma trimethylamine N-oxide (TMAO) levels, a gut microbiota-dependent metabolite associated with coronary artery disease and stroke. We further investigate the gut microbiota composition and microbiota-metabolite relationships in subjects with different body mass index and individuals with normal or altered oral glucose tolerance. Finally, we perform microbiota co-occurrence network analysis, which shows that certain metabolites strongly correlate with microbial community structure and that some of these correlations are specific for the pre-diabetic state.ConclusionsOur study identifies novel relationships between the composition of the gut microbiota and circulating metabolites and provides a resource for future studies to understand host-gut microbiota relationships.

BackgroundThe gut microbiome is a complex and metabolically active community that directly influences host phenotypes. In this study, we profile gut microbiota using 16S rRNA gene sequencing in 531 well-phenotyped Finnish men from the Metabolic Syndrome In Men (METSIM) study.ResultsWe investigate gut microbiota relationships with a variety of factors that have an impact on the development of metabolic and cardiovascular traits. We identify novel associations between gut microbiota and fasting serum levels of a number of metabolites, including fatty acids, amino acids, lipids, and glucose. In particular, we detect associations with fasting plasma trimethylamine N-oxide (TMAO) levels, a gut microbiota-dependent metabolite associated with coronary artery disease and stroke. We further investigate the gut microbiota composition and microbiota-metabolite relationships in subjects with different body mass index and individuals with normal or altered oral glucose tolerance. Finally, we perform microbiota co-occurrence network analysis, which shows that certain metabolites strongly correlate with microbial community structure and that some of these correlations are specific for the pre-diabetic state.ConclusionsOur study identifies novel relationships between the composition of the gut microbiota and circulating metabolites and provides a resource for future studies to understand host-gut microbiota relationships.

BACKGROUND: Cardiovascular risk and plasma total homocysteine (tHcy) are high in patients with renal failure. High tHcy may account for a substantial part of the increased risk. We assessed mediation by tHcy of the association of estimated glomerular filtration rate (eGFR CKD/EPI) with carotid total plaque area (TPA) and carotid stenosis. METHODS: TPA and carotid stenosis were measured by ultrasound. Multiple linear regression was used to assess the effects of eGFR and/or tHcy after adjustment for age, sex, systolic blood pressure (SBP), smoking, LDL, HDL and weight. RESULTS: Complete data were available for 1967 patients. eGFR decreased, and TPA and total stenosis increased linearly with age. After adjustment [age, sex, SBP, smoking (in pack years), low-density lipoprotein (LDL), high-density lipoprotein (HDL) and weight], eGFR and tHcy were independently associated with TPA (P < 0.01), but when both were added to the model, their significance was attenuated (P = 0.06 for eGFR, 0.03 for tHcy). Mediation analysis showed that tHcy seems to contribute to a significant mediation of the association of eGFR with TPA but not stenosis; after adjustment for the set of risk factors listed above, tHcy still demonstrated significant mediation on TPA (P = 0.03), but not on stenosis (P = 0.16). CONCLUSIONS: tHcy accounts for a significant part, but not all of the effect of renal impairment on atherosclerosis. Other uremic toxins including metabolic products of the intestinal microbiome may explain residual effects of renal failure on atherosclerosis. Therapeutic approaches arising from that hypothesis are discussed.

Epidermolysis bullosa simplex (EBS) is a dominantly inherited skin disease caused by mutations in the keratin 5 (KRT5) or KRT14 genes (1). Some reports suggested that fever and/or hot weather may exacerbate EBS phenotype (2). Effective EBS therapies are still lacking. Molecular chaperones are proteins whose main function is to promote the correct folding of polypeptides (s1). Molecules such as trimethylamine N-oxide (TMAO) and sodium 4-phenylbutyrate (4-PBA) act as chemical chaperones (s2) with protein folding and stabilization activities (s3, s4, s5).

L'épidermolyse bulleuse simplex (EBS) est une maladie orpheline dermatologique qui a une prévalence mondiale de 1/50 000 à 1/30 000. Elle est caractérisée par des mutations transmises de manière autosomale dominante dans les gènes de la kératine 5 (KRT5) et 14 (KRT14), menant à une agrégation des filaments intermédiaires. Phénotypiquement, il y a présence d'ampoules causées par un décollement de l'épiderme. Jusqu'à présent, aucune thérapie efficace n'est disponible. Dans cette étude, des essais cellulaires évaluant le potentiel thérapeutique de deux chaperonnes chimiques, le triméthylamine N-oxyde (TMAO) et le sodium 4-phénylbutyrate (4-PBA), ont été réalisés. Pour ce faire, trois lignées immortalisées de kératinocytes de patients EBS ayant une mutation dans KRT14 et une lignée contrôle ont été utilisées pour déterminer la capacité des molécules à réduire le nombre de cellules ayant des agrégats. Les résultats ont montré un effet bénéfique significatif du TMAO seulement, et ce dans deux lignées uniquement. Ainsi, le TMAO se qualifie comme une approche thérapeutique potentielle.

[EN] The present invention describes a transgenic plant resistant to cold and to saline stress based on the use of a new gene, gene RCI5, encoding a monooxygenase enzyme which induces the appearance of TMAO. Moreover, the invention provides a sequence of nucleotides, vectors and cells necessary to carry out the procedure for the obtainment of said transgenic plants. Finally, there is added the use of compositions containing TMAO to improve the tolerance to saline stress of affected plants by means of exogenous application of said solutions onto the plant or seed, for example spraying or injection., [ES] La presente invención describe una planta transgénica resistente al frió y al estrés salino basada en el uso de un nuevo gen, el gen RCI5, codificante de una enzima monoxigenasa que induce la aparición de TMAO. Además la invención proporciona secuencia de nucleótidos, vectores y células necesarias para llevar a cabo el procedimiento para obtener dichas plantas transgénicas. Finalmente, se añade el uso de composiciones que contienen TMAO para la mejora de la tolerancia al estrés salino de las plantas afectadas mediante la aplicación exógena de dichas soluciones sobre la planta o semilla, por ejemplo pulverización o inyección.

BACKGROUND: Epidermolysis bullosa simplex (EBS) is an autosomal inherited mechano-bullous disease, characterized by intraepidermal blistering and skin fragility caused by mutations in the keratin (KRT) 5 or 14 genes. Despite a vast knowledge about the intermediate filament pathology in this disease, the progress in therapy has been slow. Animal models and well-characterized continuous cell culture models of EBS are needed prior to clinical testing. OBJECTIVES: Our aim was to generate immortalized cell lines as an in vitro model for the study of EBS and test a chemical chaperone, trimethylamine N-oxide (TMAO), as a putative novel therapy. METHODS: We generated four immortalized cell lines, two each from an EBS patient with a KRT5-mutation (V186L) and a healthy control, using human papillomavirus 16 (HPV16) E6E7 as transducer. Cell lines were established in serum-free and serum-containing medium and assessed for growth characteristics, keratin expression profiles, ability to differentiate in organotypic cultures, and response to heat stress with and without the presence of TMAO. RESULTS: All cell lines have been expanded >160 population doublings and their cellular characteristics are similar. However, the formation of cytoplasmic keratin filament aggregates in response to heat-shock treatment differed between EBS and normal cell lines. Notably, serum-free established EBS-cell line was most vulnerable to heat shock but both cell lines exhibited significant reduction in the number of keratin aggregates containing cells by TMAO. CONCLUSION: The immortalized cell lines represent a suitable model for studying novel therapies for EBS. TMAO is a promising new agent for future development as a novel EBS therapy.