Gut microbiota-derived metabolite trimethylamine N-oxide aggravates cognitive dysfunction induced by femoral fracture operation in mice.

An increasing number of elderly individuals are experiencing postoperative cognitive dysfunction (POCD) problems after undergoing hip replacement surgery, with gut microbiota metabolites playing a role in its pathogenesis. Among these, the specific effects of trimethylamine N-oxide (TMAO) on POCD are still unclear. This study aimed to explore the role of TMAO on cognitive dysfunction and underlying mechanisms in mice. The POCD model was created through femoral fracture surgery in elderly mice, followed by cognitive function assessments using the Morris Water Maze and Novel Object Recognition tests. The gut microbiota depletion and fecal microbiota transplantation were performed to examine the relationship between TMAO levels and cognitive outcomes. The effects of TMAO treatment on cognitive dysfunction, microglial activation, and inflammatory cytokine levels in the brain were also evaluated, with additional assessment of the role of microglial ablation in reducing TMAO-induced cognitive impairment. Elevated TMAO levels were found to be associated with cognitive decline in mice following femoral fracture surgery, with gut microbiota depletion mitigating both TMAO elevation and cognitive dysfunction. In contrast, fecal microbiota transplantation from postoperative mice resulted in accelerated cognitive dysfunction and TMAO accumulation in germ-free mice. Furthermore, TMAO treatment worsened cognitive deficits, neuroinflammation, and promoted microglial activation, which were reversed through the ablation of microglia. TMAO exacerbates cognitive dysfunction and neuroinflammation in POCD mice, with microglial activation playing a crucial role in this process. Our findings may provide new therapeutic strategies for managing TMAO-related POCD and improving the quality of life for elderly patients.

Berberine ameliorates vascular dysfunction by downregulating TMAO-endoplasmic reticulum stress pathway via gut microbiota in hypertension.

The gut microbial metabolite trimethylamine N-oxide (TMAO) is regarded as a novel risk factor for hypertension. Berberine (BBR) exerts cardiovascular protective effects by regulating the gut microbiota-metabolite production pathway. However, whether and how BBR alleviates TMAO-induced vascular dysfunction in hypertension remains unclear. In the present study, we observed that plasma TMAO and related bacterial abundance were significantly elevated and negatively correlated with vascular function in 86 hypertensive patients compared with 46 normotensive controls. TMAO activated endoplasmic reticulum stress (ERS) signaling pathway to promote endothelial cell dysfunction and apoptosis in vitro. BBR (100, 200 mg ·â€¯kg-1 ·d-1) for 4 weeks ameliorates TMAO-induced vascular dysfunction and ERS activation in a choline-angiotensin II hypertensive mouse model. We found that plasma TMAO levels in 15 hypertensive patients treated with BBR (0.4 g, tid) were reduced by 8.8 % and 16.7 % at months 1 and 3, respectively, compared with pretreatment baseline. The oral BBR treatment also improved vascular function and lowered blood pressure. Faecal 16 S rDNA showed that BBR altered the gut bacterial composition and reduced the abundance of CutC/D bacteria in hypertensive mice and patients. In vitro bacterial cultures and enzyme reaction systems indicated that BBR inhibited the biosynthesis of TMAO precursor in the gut microbiota by binding to and inhibiting the activity of CutC/D enzyme. Our results indicate that BBR improve vascular dysfunction at least partially by decreasing TMAO via regulation of the gut microbiota in hypertension.

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Ischemic stroke (IS) is a severe cerebrovascular disease that seriously endangers human health. Gut microbiota plays a key role as an intermediate mediator in bidirectional regulation between the brain and the intestine. In recent years, trimethylamine N-oxide (TMAO) as a gut microbiota metabolite has received widespread attention in cardiovascular diseases. Elevated levels of TMAO may increase the risk of IS by affecting IS risk factors such as atherosclerosis, atrial fibrillation, hypertension, and type 2 diabetes. TMAO exacerbates neurological damage in IS patients, increases the risk of IS recurrence, and is an independent predictor of post-stroke cognitive impairment (PSCI) in patients. Current research suggests that the mechanisms of TMAO action include endothelial dysfunction, promoting of foam cell formation, influence on cholesterol metabolism, and enhancement of platelet reactivity. Lowering plasma TMAO levels through the rational use of traditional Chinese medicine, dietary management, vitamins, and probiotics can prevent and treat IS.

Developing and evaluating the construct validity of a dietary pattern predictive of plasma TMAO and choline.

BACKGROUND AND AIMS: The metabolism of choline (highly present in animal products) can produce trimethylamine N-oxide (TMAO), a metabolite with atherosclerotic effects; however, dietary fiber may suppress this metabolic pathway. This study aimed to develop a dietary pattern predictive of plasma TMAO and choline concentrations using reduced rank regression (RRR) and to evaluate its construct validity. METHODS AND RESULTS: Diet and plasma concentrations of choline (µmol/L) and TMAO (µmol/L) were assessed in 1724 post-menopausal women who participated in an ancillary study within the Women's Health Initiative Observational Study (1993-1998). The TMAO dietary pattern was developed using RRR in half of the sample (Training Sample) and applied to the other half of the sample (Validation Sample) to evaluate its construct validity. Energy-adjusted food groups were the predictor variables and plasma choline and TMAO, the response variables. ANCOVA and linear regression models were used to assess associations between each biomarker and the dietary pattern score. Discretionary fat, potatoes, red meat, and eggs were positively associated with the dietary pattern, while yogurt, fruits, added sugar, and starchy vegetables were inversely associated. Mean TMAO and choline concentrations significantly increased across increasing quartiles of the dietary pattern in the Training and Validation samples. Positive associations between the biomarkers and the TMAO dietary pattern were also observed in linear regression models (Validation Sample: TMAO, adjusted beta-coefficient = 0.037 (p-value = 0.0088); Choline, adjusted beta-coefficient = 0.011 (p-value = 0.0224). CONCLUSION: We established the TMAO dietary pattern, a dietary pattern reflecting the potential of the diet to contribute to plasma concentrations of TMAO and choline.

Obesity-associated inflammation countered by a Mediterranean diet: the role of gut-derived metabolites.

The prevalence of obesity has increased dramatically worldwide and has become a critical public health priority. Obesity is associated with many co-morbid conditions, including hypertension, diabetes, and cardiovascular disease. Although the physiology of obesity is complex, a healthy diet and sufficient exercise are two elements known to be critical to combating this condition. Years of research on the Mediterranean diet, which is high in fresh fruits and vegetables, nuts, fish, and olive oil, have demonstrated a reduction in numerous non-communicable chronic diseases associated with this diet. There is strong evidence to support an anti-inflammatory effect of the diet, and inflammation is a key driver of obesity. Changes in diet alter the gut microbiota which are intricately intertwined with human physiology, as gut microbiota-derived metabolites play a key role in biological pathways throughout the body. This review will summarize recent published studies that examine the potential role of gut metabolites, including short-chain fatty acids, bile acids, trimethylamine-N-oxide, and lipopolysaccharide, in modulating inflammation after consumption of a Mediterranean-like diet. These metabolites modulate pathways of inflammation through the NOD-like receptor family pyrin domain-containing 3 (NLRP3) inflammasome, toll-like receptor 4 signaling, and macrophage driven effects in adipocytes, among other mechanisms.

The gut-pancreas axis: investigating the relationship between microbiota metabolites and pancreatic steatosis.

The prevalence of pancreatic steatosis has increased and it has been linked to the rising prevalence of metabolic syndrome. Metabolic syndrome is known to have a strong connection with changes in intestinal microbiota. The aim of this study was to explore the relationship between pancreatic steatosis and the levels of trimethylamine N-oxide (TMAO) and butyrate. In this study, 136 individuals were randomly selected from outpatient clinics at Firat University Hospital. The study evaluated their demographic characteristics, anthropometric measurements, and biochemical parameters. The presence of pancreatic steatosis was assessed using abdominal ultrasonography. Additionally, the levels of TMAO and butyrate were measured. The mean age of individuals in the study was 44.5 ± 14.6. 84 of the subjects were females. Using the waist circumference, 61 were considered obese and 34 overweight. The detection rate of pancreatic steatosis was found to be 70.6%. The study found that individuals with steatosis had higher average age, presence of hepatic steatosis, BMI, waist circumference measurements, and presence of metabolic syndrome than those without steatosis. A significantly higher butyrate level was detected in those without steatosis (p = 0.001). TMAO levels were slightly higher in patients without steatosis than in those with steatosis; however, this was insignificant. Pancreatic steatosis is highly associated with alterations in levels of microbiota metabolites, indicating a potential role of these metabolites in the pathogenesis of the disease and subsequent therapeutic targets. Several other factors, such as age, hepatic steatosis, diabetes, and waist circumference, have also been identified as potential predictors of pancreatic steatosis.

Circulating TMAO, the gut microbiome and cardiometabolic disease risk: an exploration in key precursor disorders.

BACKGROUND: Elevations in the gut metabolite trimethylamine-N-oxide (TMAO) have been linked to cardiovascular and metabolic diseases. Whether elevated TMAO levels reflect early mechanistic involvement or a sequela of evolving disease awaits elucidation. The purpose of this study was to further explore these potential associations. METHODS: We investigated relationships between circulating levels of TMAO and its pre-cursor substrates, dietary factors, gut microbiome profiles and disease risk in individuals with a Healthy BMI (18.5 < BMI < 25, n = 41) or key precursor states for cardiometabolic disease: Overweight (25 < BMI < 30 kg/m2, n = 33), Obese (BMI > 30, n = 27) and Metabolic Syndrome (MetS; ≥ 3 ATPIII report criteria, n = 39). RESULTS: Unexpectedly, plasma [TMAO] did not vary substantially between groups (means of 3-4 µM; p > 0.05), although carnitine was elevated in participants with MetS. Gut microbial diversity and Firmicutes were also significantly reduced in the MetS group (p < 0.05). Exploratory analysis across diverse parameters reveals significant correlations between circulating [TMAO] and seafood intake (p = 0.007), gut microbial diversity (p = 0.017-0.048), and plasma [trimethylamine] (TMA; p = 0.001). No associations were evident with anthropometric parameters or cardiometabolic disease risk. Most variance in [TMAO] within and between groups remained unexplained. CONCLUSIONS: Data indicate that circulating [TMAO] may be significantly linked to seafood intake, levels of TMA substrate and gut microbial diversity across healthy and early disease phenotypes. However, mean concentrations remain < 5 µM, with little evidence of links between TMAO and cardiometabolic disease risk. These observations suggest circulating TMAO may not participate mechanistically in cardiometabolic disease development, with later elevations likely a detrimental sequela of extant disease.

Effect of dapagliflozin on ferroptosis through the gut microbiota metabolite TMAO during myocardial ischemia-reperfusion injury in diabetes mellitus rats.

Dapagliflozin (DAPA) demonstrates promise in the management of diabetic mellitus (DM) and cardiomyopathy. Trimethylamine N-oxide (TMAO) is synthesized by the gut microbiota through the metabolic conversion of choline and phosphatidylcholine. Ferroptosis may offer novel therapeutic avenues for the management of diabetes and myocardial ischemia-reperfusion injury (IRI). However, the precise mechanism underlying ferroptosis in cardiomyocytes and the specific role of TMAO generated by gut microbiota in the therapeutic approach for DM and myocardial IRI utilizing DAPA need to be further explored. Nine male SD rats with specific pathogen-free (SPF) status were randomly divided equally into the normal group, the DM + IRI (DIR) group, and the DAPA group. The diversity of the gut microbiota was analyzed using 16S rRNA gene sequencing. Additionally, the Wekell technique was employed to measure the levels of TMAO in the three groups. Application of network pharmacology to search for intersection targets of DAPA, DIR, and ferroptosis, and RT-PCR experimental verification. Ultimately, the overlapping targets that were acquired were subjected to molecular docking analysis with TMAO. The changes of Bacteroidetes and Firmicutes in the gut microbiota of DIR rats were most significantly affected by DAPA. Escherichia-Shigella and Prevotella_9 within the phylum Bacteroidetes could be identified as the primary effects of DAPA on DIR. Compared with the normal group, the TMAO content in the DIR group was significantly increased, while the TMAO content in the DAPA group was decreased compared to the DIR group. For the network pharmacology analysis, DAPA and DIR generated 43 intersecting target genes, and then further intersected with ferroptosis-related genes, resulting in 11 overlapping target genes. The mRNA expression of ALB, HMOX1, PPARG, CBS, LCN2, and PPARA decreased in the DIR group through reverse transcription polymerase chain reaction (RT-PCR) validation, while the opposite trend was observed in the DAPA group. The docking score between TMAO and DPP4 was - 5.44, and the MM-GBSA result of - 22.02 kcal/mol. It epitomizes the finest docking performance among all the target genes with the lowest score. DAPA could reduce the levels of metabolite TMAO produced by gut microbiota, thereby regulating related target genes to decrease ferroptosis in DIR cardiomyocytes.

Dynamic Changes in Gut Microbiota-Derived Metabolite Trimethylamine-N-Oxide and Risk of Type 2 Diabetes Mellitus: Potential for Dietary Changes in Diabetes Prevention.

BACKGROUND: A gut-microbial metabolite, trimethylamine N-oxide (TMAO), has been associated with type 2 diabetes mellitus (T2DM). Few previous prospective studies have addressed associations between the changes in TMAO and T2DM incidence. METHODS: Data were derived from a longitudinal cohort conducted from 2019 to 2021 in rural areas of Fuxin County, Liaoning Province, China, and 1515 diabetes-free participants aged above 35 years were included. The concentrations of serum TMAO and its precursors were measured at two time points, namely in 2019 and 2021. TMAO and TMAO changes (ΔTMAO) were separately tested in a logistic regression model. For further examination, the odds ratios (ORs) for T2DM were calculated according to a combination of TMAO levels and ΔTMAO levels. RESULTS: During a median follow-up of 1.85 years, 81 incident cases of T2DM (5.35%) were identified. Baseline TMAO levels exhibited a nonlinear relationship, first decreasing and then increasing, and only at the highest quartile was it associated with the risk of T2DM. The OR for T2DM in the highest quartile of serum TMAO was 3.35 (95%CI: 1.55-7.26, p = 0.002), compared with the lowest quartile. As for its precursors, only choline level was associated with T2DM risk and the OR for T2DM in the Q3 and Q4 of serum choline was 3.37 (95%CI: 1.41-8.05, p = 0.006) and 4.72 (95%CI: 1.47-15.13, p = 0.009), respectively. When considering both baseline TMAO levels and ΔTMAO over time, participants with sustained high TMAO levels demonstrated a significantly increased risk of T2DM, with a multivariable-adjusted OR of 8.68 (95%CI: 1.97, 38.34). CONCLUSION: Both initial serum TMAO levels and long-term serum TMAO changes were collectively and significantly associated with the occurrence of subsequent T2DM events. Interventions aimed at normalizing TMAO levels, such as adopting a healthy dietary pattern, may be particularly beneficial in T2DM prevention.

Circulating Trimethylamine-N-Oxide Is Elevated in Liver Transplant Recipients.

Liver transplant recipients (LTRs) have lower long-term survival rates compared with the general population. This underscores the necessity for developing biomarkers to assess post-transplantation mortality. Here we compared plasma trimethylamine-N-oxide (TMAO) levels with those in the general population, investigated its determinants, and interrogated its association with all-cause mortality in stable LTRs. Plasma TMAO was measured in 367 stable LTRs from the TransplantLines cohort (NCT03272841) and in 4837 participants from the population-based PREVEND cohort. TMAO levels were 35% higher in LTRs compared with PREVEND participants (4.3 vs. 3.2 µmol/L, p < 0.001). Specifically, TMAO was elevated in LTRs with metabolic dysfunction-associated steatotic liver disease, alcohol-associated liver disease, and polycystic liver disease as underlying etiology (p < 0.001 for each). Among LTRs, TMAO levels were independently associated with eGFR (std. ß = -0.43, p < 0.001) and iron supplementation (std. ß = 0.13, p = 0.008), and were associated with mortality (29 deaths during 8.6 years follow-up; log-rank test p = 0.017; hazard ratio of highest vs. lowest tertile 4.14, p = 0.007). In conclusion, plasma TMAO is likely elevated in stable LTRs, with impaired eGFR and iron supplementation as potential contributory factors. Our preliminary findings raise the possibility that plasma TMAO could contribute to increased mortality risk in such patients, but this need to be validated through a series of rigorous and methodical studies.

Simultaneous determination of choline, L-carnitine, betaine, trimethylamine, trimethylamine N-oxide, and creatinine in plasma, liver, and feces of hyperlipidemic rats by UHPLC-MS/MS.

BACKGROUND: Due to the close correlation between choline, L-carnitine, betaine and their intestinal microbial metabolites, including trimethylamine (TMA) and trimethylamine N-oxide (TMAO), and creatinine, there has been an increasing interest in the study of these compounds in vivo. METHODS: In this study, a rapid stable isotope dilution (SID)-UHPLC-MS/MS method was developed for the simultaneous determination of choline, L-carnitine, betaine, TMA, TMAO and creatinine in plasma, liver and feces of rats. The method was validated using quality control (QC) samples spiked at low, medium and high levels. Second, we applied the method to quantify the effects of Rosa Roxburghii Tratt juice (RRTJ) on plasma, liver, and fecal levels of choline, L-carnitine, betaine, TMA, TMAO, and creatinine in high-fat diet-induced hyperlipidemic rats, demonstrating the utility of the method. RESULTS: The limits of detection (LOD) were 0.04-0.027 µM and the limits of quantification (LOQ) were 0.009-0.094 µM. The linear ranges for each metabolite in plasma were choline1.50-96 µM; L-carnitine: 2-128 µM; betaine: 3-192 µM; TMA: 0.01-40.96 µM; TMAO: 0.06-61.44 µM and creatinine: 1-64 µM (R2 ≥ 0.9954). The linear ranges for each metabolite in liver were Choline: 12-768 µM; L-carnitine: 1.5-96 µM; betaine: 10-640 µM; TMA: 0.5-32 µM; TMAO: 0.02-81.92 µM and creatinine: 0.2-204.8 µM (R2 ≥ 0.9938). The linear ranges for each metabolite in feces were choline: 1.5-96 µM; L-carnitine: 0.01-40.96 µM; Betaine: 1.5-96 µM; TMA: 1-64 µM; TMAO: 0.02-81.92 µM and Creatinine: 0.02-81.92 µM (R2 ≥ 0.998). The intra-day and inter-day coefficients of variation were < 8 % for all analytes. The samples were stabilized after multiple freeze-thaw cycles (3 freeze-thaw cycles), 24 h at room temperature, 24 h at 4 °C and 20 days at -80 °C. The samples were stable. The average recovery was 89 %-99 %. This method was used to quantify TMAO and its related metabolites and creatinine levels in hyperlipidemic rats. The results showed that high-fat diet led to the disorder of TMAO and its related metabolites and creatinine in rats, which was effectively improved after the intervention of Rosa Roxburghii Tratt juice（RRTJ）. CONCLUSIONS: A method for the determination of choline, L-carnitine, betaine, TMA, TMAO and creatinine in plasma, liver and feces samples was established, which is simple, time-saving, high precision, accuracy and recovery.

Effects of resveratrol on changes in trimethylamine-N-oxide and circulating cardiovascular factors following exercise training among older adults.

PURPOSE: Trimethylamine-N-oxide (TMAO) is a gut-derived metabolite associated with cardiovascular disease (CVD). In preclinical and observational studies, resveratrol and exercise training have been suggested as potential strategies to reduce the systemic levels of TMAO. However, evidence from experimental studies in humans remains unknown. This project examined the dose-dependent effects of a combined resveratrol intervention with exercise training on circulating TMAO and other related metabolite signatures in older adults with high CVD risk. METHODS: Forty-one older adults [mean (±SD) age of 72.1 (6.8) years] participated in a 12-week supervised center-based, multi-component exercise training intervention [2×/week; 80 min/session] and were randomized to one of two resveratrol dosages [Low: 500 vs. High:1000 mg/day] or a cellulose-based placebo. Serum/plasma were collected at baseline and post-intervention and evaluated for TMAO and associated analytes. RESULTS: After the 12-week intervention, TMAO concentration increased over time, regardless of treatment [mean (±SD) Placebo: 11262 (±3970); Low:13252 (±1193); High: 12661(±3359) AUC; p = 0.04]. Each resveratrol dose produced different changes in metabolite signatures. Low dose resveratrol upregulated metabolites associated with bile acids biosynthesis (i.e., glycochenodeoxycholic acid, glycoursodeoxycholic acid, and glycocholic acid). High dose resveratrol modulated metabolites enriched for glycolysis, and pyruvate, propanoate, ß-alanine, and tryptophan metabolism. Different communities tightly correlated to TMAO and resveratrol metabolites were associated with the lipid and vascular inflammatory clinical markers [|r| > 0.4, p < 0.05]. CONCLUSION: These findings suggest a distinct dose-dependent adaptation response to resveratrol supplementation on circulating metabolite signatures but not on TMAO among high-risk CVD older adults when combined with an exercise training intervention.

Betaine supplementation modulates betaine concentration by methylenetetrahydrofolate reductase genotype, but has no effect on amino acid profile in healthy active males: A randomized placebo-controlled cross-over study.

Betaine supplementation is used by athletes, but its mechanism of action is still not fully understood. We hypothesized that betaine supplementation would increase betaine concentration and alter amino acid profiles in relation to MTHFR genotype and dose in physically active males. The study followed a randomized placebo-controlled cross-over design. Blood samples were collected before and after each supplementation period. Serum was analyzed for amino acid profile, homocysteine, betaine, choline, and trimethylamine N-oxide (TMAO) concentrations. For the washout analysis, only participants starting with betaine were included (n = 20). Statistical analysis revealed no differences in the amino acid profile after betaine supplementation. However, betaine concentration significantly increased after betaine supplementation (from 4.89 ± 1.59 µg/mL to 17.31 ± 9.21 µg/mL, P < .001), with a greater increase observed in MTHFR (C677T, rs180113) T-allele carriers compared to CC (P = .027). Betaine supplementation caused a decrease in homocysteine concentration (from 17.04 ± 4.13 µmol/L to 15.44 ± 3.48 µmol/L, P = .00005) and a non-significant increase in TMAO concentrations (from 0.27 ± 0.20 µg/ml to 0.44 ± 0.70 µg/ml, P = .053), but had no effect on choline concentrations. Serum betaine concentrations were not significantly different after the 21-day washout from the baseline values (baseline: 4.93 ± 1.87 µg/mL and after washout: 4.70 ± 1.70 µg/mL, P = 1.000). In conclusion, betaine supplementation increased betaine and decreased homocysteine concentrations, but did not affect the amino acid profile or choline concentrations in healthy active males. Betaine concentrations may be dependent on MTHFR genotype.

The gut microbiota derived metabolite trimethylamine N-oxide: Its important role in cancer and other diseases.

An expanding body of research indicates a correlation between the gut microbiota and various diseases. Metabolites produced by the gut microbiota act as mediators between the gut microbiota and the host, interacting with multiple systems in the human body to regulate physiological or pathological functions. However, further investigation is still required to elucidate the underlying mechanisms. One such metabolite involved in choline metabolism by gut microbes is trimethylamine (TMA), which can traverse the intestinal epithelial barrier and enter the bloodstream, ultimately reaching the liver where it undergoes oxidation catalyzed by flavin-containing monooxygenase 3 (FMO3) to form trimethylamine N-oxide (TMAO). While some TMAO is eliminated through renal excretion, remaining amounts circulate in the bloodstream, leading to systemic inflammation, endoplasmic reticulum (ER) stress, mitochondrial stress, and disruption of normal physiological functions in humans. As a representative microbial metabolite originating from the gut, TMAO has significant potential both as a biomarker for monitoring disease occurrence and progression and for tailoring personalized treatment strategies for patients. This review provides an extensive overview of TMAO sources and its metabolism in human blood, as well as its impact on several major human diseases. Additionally, we explore the latest research areas related to TMAO along with future directions.

Effect of three oral pathogens on the TMA-TMAO metabolic pathway.

Background: Trimethylamine-N-oxide (TMAO) is produced by hepatic flavin-containing monooxygenase 3 (FMO3) from trimethylamine (TMA). High TMAO level is a biomarker of cardiovascular diseases and metabolic disorders, and it also affects periodontitis through interactions with the gastrointestinal microbiome. While recent findings indicate that periodontitis may alter systemic TMAO levels, the specific mechanisms linking these changes and particular oral pathogens require further clarification. Methods: In this study, we established a C57BL/6J male mouse model by orally administering Porphyromonas gingivalis (P. gingivalis, Pg), Fusobacterium nucleatum (F. nucleatum, Fn), Streptococcus mutans (S. mutans, Sm) and PBS was used as a control. We conducted LC-MS/MS analysis to quantify the concentrations of TMAO and its precursors in the plasma and cecal contents of mice. The diversity and composition of the gut microbiome were analyzed using 16S rRNA sequencing. TMAO-related lipid metabolism and enzymes in the intestines and liver were assessed by qPCR and ELISA methods. We further explored the effect of Pg on FMO3 expression and lipid molecules in HepG2 cells by stimulating the cells with Pg-LPS in vitro. Results: The three oral pathogenic bacteria were orally administered to the mice for 5 weeks. The Pg group showed a marked increase in plasma TMAO, betaine, and creatinine levels, whereas no significant differences were observed in the gut TMAO level among the four groups. Further analysis showed similar diversity and composition in the gut microbiomes of both the Pg and Fn groups, which were different from the Sm and control groups. The profiles of TMA-TMAO pathway-related genera and gut enzymes were not significantly different among all groups. The Pg group showed significantly higher liver FMO3 levels and elevated lipid factors (IL-6, TG, TC, and NEFA) in contrast to the other groups. In vitro experiments confirmed that stimulation of HepG2 cells with Pg-LPS upregulated the expression of FMO3 and increased the lipid factors TC, TG, and IL-6. Conclusion: This study conclusively demonstrates that Pg, compared to Fn and Sm, plays a critical role in elevating plasma TMAO levels and significantly influences the TMA-TMAO pathway, primarily by modulating the expression of hepatic FMO3 and directly impacting hepatic lipid metabolism.

Unravelling the Gut Microbiome Role in Cardiovascular Disease: A Systematic Review and a Meta-Analysis.

A notable shift in understanding the human microbiome's influence on cardiovascular disease (CVD) is underway, although the causal association remains elusive. A systematic review and meta-analysis were conducted to synthesise current knowledge on microbial taxonomy and metabolite variations between healthy controls (HCs) and those with CVD. An extensive search encompassing three databases identified 67 relevant studies (2012-2023) covering CVD pathologies from 4707 reports. Metagenomic and metabolomic data, both qualitative and quantitative, were obtained. Analysis revealed substantial variability in microbial alpha and beta diversities. Moreover, specific changes in bacterial populations were shown, including increased Streptococcus and Proteobacteria and decreased Faecalibacterium in patients with CVD compared with HC. Additionally, elevated trimethylamine N-oxide levels were reported in CVD cases. Biochemical parameter analysis indicated increased fasting glucose and triglycerides and decreased total cholesterol and low- and high-density lipoprotein cholesterol levels in diseased individuals. This study revealed a significant relationship between certain bacterial species and CVD. Additionally, it has become clear that there are substantial inconsistencies in the methodologies employed and the reporting standards adhered to in various studies. Undoubtedly, standardising research methodologies and developing extensive guidelines for microbiome studies are crucial for advancing the field.

Trimethylamine N-oxide ameliorates hepatic damage including reduction of hepatic bile acids and cholesterol in Fxr-null mice.

There are conflicting animal experiments on the effect of trimethylamine N-oxide (TMAO), the dietary metabolite, on non-alcoholic fatty liver disease (NAFLD). This study aims to determine the effect of TMAO on NAFLD. A diet containing 0.3% TMAO was fed to farnesoid X receptor (Fxr)-null mice, a model of NAFLD, for 13 weeks. Fxr-null mice fed TMAO showed significant reductions in liver damage markers but not wild-type mice. Hepatic bile acid and cholesterol levels were significantly decreased, and triacylglycerol levels tended to decrease in TMAO-fed Fxr-null mice. Changes in mRNA levels of hepatic bile acid and cholesterol transporters and synthetic enzymes were observed, which could explain the decreased hepatic bile acid and cholesterol levels in Fxr-null mice given the TMAO diet but not in the wild-type mice. These results suggest that TMAO intake ameliorates liver damage in Fxr-null mice, further altering bile acid/cholesterol metabolism in an FXR-independent manner.

Trimethylamine-N-oxide promotes osteoclast differentiation and oxidative stress by activating NF-κB pathway.

BACKGROUND: Senile osteoporosis may be caused by an imbalance in intestinal flora and oxidative stress. Trimethylamine-N-oxide (TMAO), a metabolite of dietary choline dependent on gut microbes, has been found to be significantly increased in osteoporosis. However, the role of TMAO in bone loss during osteoporosis remains poorly understood. In this study, we examined the impact of TMAO on osteoclast differentiation and bone resorption in an in vitro setting. METHODS: Osteoclast differentiation was induced by incubating RAW 264.7 cells in the presence of Receptor Activator for Nuclear Factor-κB Ligand (RANKL) and macrophage-stimulating factor (M-CSF). Flow cytometry, TRAP staining assay, CCK-8, and ELISA were employed to investigate the impact of TMAO on osteoclast differentiation and bone resorption activity in vitro. For mechanistic exploration, RT-PCR and Western blotting were utilized to assess the activation of the NF-κB pathway. Additionally, protein levels of secreted cytokines and growth factors were determined using suspension array technology. RESULTS: Our findings demonstrate that TMAO enhances RANKL and M-CSF-induced osteoclast formation and bone resorption in a dose-dependent manner. Mechanistically, TMAO triggers the upregulation of the NF-κB pathway and osteoclast-related genes (NFATc1, c-Fos, NF-κB p65, Traf6, and Cathepsin K). Furthermore, TMAO markedly elevated the levels of oxidative stress and inflammatory factors. CONCLUSIONS: In conclusion, TMAO enhances RANKL and M-CSF-induced osteoclast differentiation and inflammation in RAW 264.7 cells by activating the NF-κB signaling pathway. These findings offer a new rationale for further academic and clinical research on osteoporosis treatment.

Deciphering the causal relationship between plasma and cerebrospinal fluid metabolites and glioblastoma multiforme: a Mendelian Randomization study.

BACKGROUND: Glioblastoma Multiforme (GBM) is one of the most aggressive and fatal brain cancers. The study of metabolites could be crucial for understanding GBM's biology and reveal new treatment strategies. METHODS: The GWAS data for GBM were sourced from the FinnGen database. A total of 1400 plasma metabolites were collected from the GWAS Catalog dataset. The cerebrospinal fluid (CSF) metabolites data were collected from subsets of participants in the WADRC and WRAP studies. We utilized the inverse variance weighting (IVW) method as the primary tool to explore the causal relationship between metabolites in plasma and CSF and glioblastoma, ensuring the exclusion of instances with horizontal pleiotropy. Additionally, four supplementary analytical methods were applied to reinforce our findings. Aberrant results were identified and omitted based on the outcomes of the leave-one-out sensitivity analysis. Conclusively, a reverse Mendelian Randomization analysis was also conducted to further substantiate our results. RESULTS: The study identified 69 plasma metabolites associated with GBM. Of these, 40 metabolites demonstrated a significant positive causal relationship with GBM, while 29 exhibited a significant negative causal association. Notably, Trimethylamine N-oxide (TMAO) levels in plasma, not CSF, were found to be a significant exposure factor for GBM (OR = 3.1627, 95% CI = (1.6347, 6.1189), P = 0.0006). The study did not find a reverse causal relationship between GBM and plasma TMAO levels. CONCLUSIONS: This research has identified 69 plasma metabolites potentially associated with the incidence of GBM, among which TMAO stands out as a promising candidate for an early detectable biomarker for GBM.

Efficient Reductive N-11C-Methylation Using Arylamines or Alkylamines and In Situ-Generated [11C]Formaldehyde From [11C]Methyl Iodide.

Reductive N-11C-methylation using [11C]formaldehyde and amines has been used to prepare N-11C-methylated compounds. However, the yields of the N-11C-methylated compounds are often insufficient. In this study, we developed an efficient method for base-free reductive N-11C-methylation that is applicable to a wide variety of substrates, including arylamines bearing electron-withdrawing and electron-donating substituents. A 2-picoline borane complex, which is a stable and mild reductant, was used. Dimethyl sulfoxide was used as the primary reaction solvent, and glacial acetic acid or aqueous acetic acid was used as a cosolvent. While reductive N-11C-methylation efficiently proceeded under anhydrous conditions in most cases, the addition of water to the reductive N-11C-methylation generally increased the yield of the N-11C-methylated compounds. Substrates with hydroxy, carboxyl, nitrile, nitro, ester, amide, and phenone moieties and amine salts were applicable to the reaction. This proposed method for reductive N-11C-methylation should be applicable to a wide variety of substrates, including thermo-labile and base-sensitive compounds because the reaction was performed under relatively mild conditions (70°C) without the need for a base.