Dietary Choline Research Articles

Effects of Egg Consumption and Choline Supplementation on Plasma Choline and Trimethylamine-N-Oxide in a Young Population

ABSTRACTBackground: Plasma trimethylamine-N-oxide (TMAO) concentrations have been associated with cardiovascular disease risk. Eggs are a rich source of choline, which is a precursor of TMAO.Objective: The effects of egg intake versus daily choline supplementation were evaluated on plasma choline and TMAO in a young, healthy population.Methods: Thirty participants (14 males, 16 females; 25.6 ± 2.3 years; body mass index = 24.3 ± 2.9 kg/m2) were enrolled in this 13-week crossover intervention. After a 2-week washout, participants were randomized to consume either 3 eggs/d or a choline bitartrate supplement (∼ 400 mg choline total in eggs or supplement) for 4 weeks. Following a 3-week washout, participants were switched to the alternate treatment. Dietary records were measured at the end of each period. Plasma TMAO and choline were measured at baseline and at the end of each dietary intervention. Gene expression of scavenger receptors associated with plasma TMAO were quantified at the end of each intervention.Results: Compared to the choline supplement, intake of total fat, cholesterol, selenium, and vitamin E were higher (p < 0.05), whereas carbohydrate intake was lower (p < 0.001) with consumption of 3 eggs/d. Fasting plasma choline increased 20% (p = 0.023) with egg intake, while no changes were observed with choline supplementation. Plasma TMAO levels were not different between dietary treatments or compared to baseline.Conclusions: Dietary choline appears to be more bioavailable via egg consumption when compared to a choline supplement. Plasma TMAO concentrations were not affected in healthy participants after 4 weeks of taking ∼400 mg/d choline either via eggs or choline supplementation.

Abstract 105: Increased Circulating Trimethylamine N-oxide (TMAO) Augments the Incidence of Abdominal Aortic Aneurysm in Low Penetrant C57BL/6J Mice

Background: The gut microbiota is a metabolically active endocrine organ critical to the maintenance of cardiovascular health. Dietary sources of choline are metabolized by microbial enzymes to form trimethylamine (TMA). Metabolism by the host hepatic enzyme flavin-containing monooxygenase 3 (FMO3) converts TMA to the pro-inflammatory molecule trimethylamine N-oxide (TMAO). Human clinical trials have correlated high levels of circulating TMAO to an increased risk of cardiometabolic diseases. However, this meta-organismal pathway has not been evaluated in the context of abdominal aortic aneurysm (AAA). The objective of this study was to determine the effects of a high choline diet on the development of AAA. Methods: C57BL/6J male (n=20) and female (n=20) mice were fed either a standard chow control diet (n = 10 each sex) or a choline-rich diet (1%; n = 10 each sex) for 5 weeks. After 1 week of diet, basal abdominal ultrasounds were performed and angiotensin II (AngII; 1,000 ng/kg/min) was infused for 28 days via implantation of osmotic minipumps. Termination ultrasounds were performed on day 27 and mice were sacrificed on day 28. Aortas were harvested for evaluation of aneurysm progression and plasma was analyzed for the metabolites TMA, TMAO, and choline. To determine whether TMAO was elevated in human patients with AAA, plasma samples from participants with fast growing AAAs (n = 85), slow growing AAAs (n = 84), and normal (non-aneurysmal) aortas (n = 115) were analyzed for plasma TMAO levels via liquid chromatography tandem mass spectrometry (LC-MS/MS). Results: Administration of a choline-rich diet augmented the incidence (P &lt; 0.02) and aortic diameter (P &lt; 0.001) of AAAs in both male and female mice versus placebo-fed mice. Plasma levels of TMA, TMAO, and choline were significantly elevated in choline-fed mice versus normal chow (P &lt; 0.05). Importantly, circulating levels of plasma TMAO were significantly elevated in a step-wise fashion with the rate of aneurysm growth versus non-aneurysmal control patients (fast growing &gt; slow growing &gt; normal patients; P &lt; 0.001). Conclusions: Our results indicate increases in circulating TMAO augments the growth status of aneurysms in human patients and the incidence of AAA in a low penetrant C57BL/6J mouse model.

Dietary choline supplementation in adult rats improves performance on a test of recognition memory

In two experiments adult rats (aged at least 6 months at the start of the procedure) received a diet enriched with added choline for a period of 10 weeks; control subjects were maintained on a standard diet during this time. All rats then underwent the spontaneous object recognition (SOR) procedure in which they were exposed to a pair of objects and then tested, after a retention interval, to a display with one object changed. Exploration of the changed object indicates retention and use of information acquired during the exposure phase. All subjects showed retention with a 24-h interval (Experiments 1 and 2) and when retested after a further 24 h (Experiment 1). But when tested for the first time after a 48-h interval (Experiment 2), control subjects showed no evidence of retention, exploring both objects equally, whereas those given the dietary supplement continued to show a preference for the changed object. This supports the conclusion that dietary choline supplementation can enhance performance on a task regarded as a test of declarative memory, and will do so even when the supplementations is given in adulthood.

Metabolic retroconversion of trimethylamine N-oxide and the gut microbiota

BackgroundThe dietary methylamines choline, carnitine, and phosphatidylcholine are used by the gut microbiota to produce a range of metabolites, including trimethylamine (TMA). However, little is known about the use of trimethylamine N-oxide (TMAO) by this consortium of microbes.ResultsA feeding study using deuterated TMAO in C57BL6/J mice demonstrated microbial conversion of TMAO to TMA, with uptake of TMA into the bloodstream and its conversion to TMAO. Microbial activity necessary to convert TMAO to TMA was suppressed in antibiotic-treated mice, with deuterated TMAO being taken up directly into the bloodstream. In batch-culture fermentation systems inoculated with human faeces, growth of Enterobacteriaceae was stimulated in the presence of TMAO. Human-derived faecal and caecal bacteria (n = 66 isolates) were screened on solid and liquid media for their ability to use TMAO, with metabolites in spent media analysed by 1H-NMR. As with the in vitro fermentation experiments, TMAO stimulated the growth of Enterobacteriaceae; these bacteria produced most TMA from TMAO. Caecal/small intestinal isolates of Escherichia coli produced more TMA from TMAO than their faecal counterparts. Lactic acid bacteria produced increased amounts of lactate when grown in the presence of TMAO but did not produce large amounts of TMA. Clostridia (sensu stricto), bifidobacteria, and coriobacteria were significantly correlated with TMA production in the mixed fermentation system but did not produce notable quantities of TMA from TMAO in pure culture.ConclusionsReduction of TMAO by the gut microbiota (predominantly Enterobacteriaceae) to TMA followed by host uptake of TMA into the bloodstream from the intestine and its conversion back to TMAO by host hepatic enzymes is an example of metabolic retroconversion. TMAO influences microbial metabolism depending on isolation source and taxon of gut bacterium. Correlation of metabolomic and abundance data from mixed microbiota fermentation systems did not give a true picture of which members of the gut microbiota were responsible for converting TMAO to TMA; only by supplementing the study with pure culture work and additional metabolomics was it possible to increase our understanding of TMAO bioconversions by the human gut microbiota.

Methodological considerations for the identification of choline and carnitine-degrading bacteria in the gut.

The bacterial formation of trimethylamine (TMA) has been linked to cardiovascular disease. This review focuses on the methods employed to investigate the identity of the bacteria responsible for the formation of TMA from dietary choline and carnitine in the human gut. Recent studies have revealed the metabolic pathways responsible for bacterial TMA production, primarily the anaerobic glycyl radical-containing, choline-TMA lyase, CutC and the aerobic carnitine monooxygenase, CntA. Identification of these enzymes has enabled bioinformatics approaches to screen both human-associated bacterial isolate genomes and whole gut metagenomes to determine which bacteria are responsible for TMA formation in the human gut. We centre on several key methodological aspects for identifying the TMA-producing bacteria and report how these pathways can be identified in human gut microbiota through bioinformatics analysis of available bacterial genomes and gut metagenomes.

Maternal Choline Supplementation during Normal Murine Pregnancy Alters the Placental Epigenome: Results of an Exploratory Study.

The placental epigenome regulates processes that affect placental and fetal development, and could be mediating some of the reported effects of maternal choline supplementation (MCS) on placental vascular development and nutrient delivery. As an extension of work previously conducted in pregnant mice, the current study sought to explore the effects of MCS on various epigenetic markers in the placenta. RNA and DNA were extracted from placentas collected on embryonic day 15.5 from pregnant mice fed a 1X or 4X choline diet, and were subjected to genome-wide sequencing procedures or mass-spectrometry-based assays to examine placental imprinted gene expression, DNA methylation patterns, and microRNA (miRNA) abundance. MCS yielded a higher (fold change = 1.63–2.25) expression of four imprinted genes (Ampd3, Tfpi2, Gatm and Aqp1) in the female placentas and a lower (fold change = 0.46–0.62) expression of three imprinted genes (Dcn, Qpct and Tnfrsf23) in the male placentas (false discovery rate (FDR) ≤ 0.05 for both sexes). Methylation in the promoter regions of these genes and global placental DNA methylation were also affected (p ≤ 0.05). Additionally, a lower (fold change = 0.3; Punadjusted = 2.05 × 10−4; FDR = 0.13) abundance of miR-2137 and a higher (fold change = 1.25–3.92; p < 0.05) expression of its target genes were detected in the 4X choline placentas. These data demonstrate that the placental epigenome is responsive to maternal choline intake during murine pregnancy and likely mediates some of the previously described choline-induced effects on placental and fetal outcomes.

Untargeted metabolomics identifies trimethyllysine, a TMAO-producing nutrient precursor, as a predictor of incident cardiovascular disease risk.

Using an untargeted metabolomics approach in initial (N = 99 subjects) and replication cohorts (N = 1,162), we discovered and structurally identified a plasma metabolite associated with cardiovascular disease (CVD) risks, N6,N6,N6-trimethyl-L-lysine (trimethyllysine, TML). Stable-isotope-dilution tandem mass spectrometry analyses of an independent validation cohort (N = 2,140) confirmed TML levels are independently associated with incident (3-year) major adverse cardiovascular event risks (hazards ratio [HR], 2.4; 95% CI, 1.7-3.4) and incident (5-year) mortality risk (HR, 2.9; 95% CI, 2.0-4.2). Genome-wide association studies identified several suggestive loci for TML levels, but none reached genome-wide significance; and d9(trimethyl)-TML isotope tracer studies confirmed TML can serve as a nutrient precursor for gut microbiota-dependent generation of trimethylamine (TMA) and the atherogenic metabolite trimethylamine N-oxide (TMAO). Although TML was shown to be abundant in both plant- and animal-derived foods, mouse and human fecal cultures (omnivores and vegans) showed slow conversion of TML to TMA. Furthermore, unlike chronic dietary choline, TML supplementation in mice failed to elevate plasma TMAO or heighten thrombosis potential in vivo. Thus, TML is identified as a strong predictor of incident CVD risks in subjects and to serve as a dietary precursor for gut microbiota-dependent generation of TMAO; however, TML does not appear to be a major microbial source for TMAO generation in vivo.

Betaine, choline and folate content in different cereal genotypes

The importance of dietary methyl donors, e.g. betaine, choline and folate, is increasingly being recognised. This study examined variations in methyl donor concentrations in different cereals grown in Sweden. Fourteen cereal samples, representing different genera and cultivars, were analysed using HPLC-UV/FLD. The content of methyl donors in the cereals varied significantly due to cereal genotype. Betaine content varied most, with 28 mg/100 g DM in oats and 176 mg/100 g DM in rye. Total choline varied less, with 67 mg/100 g DM in rye and 149 mg/100 g DM in naked barley. In wheat, the lowest concentration of folate with 36 μg/100 g DM was found, and the highest of 91 μg/100 g DM in barley. Esterified choline was the major contributor to total choline content (80–95%) in the cereals. Free choline was less abundant, ranging from 3 to 27 mg/100 g DM. 5-CHO-H4folate was the dominant folate form in all cereals, amounting to approx. 35–50% of the sum of folates, as determined after pre-column conversion. Due to the limited number of available cultivars, no interpretation regarding effects from cultivar can be made. In conclusion, the studied cereal genotypes are good sources of methyl donors, but concentrations show considerable variation between different cereals.

Simultaneous determination of trimethylamine N-oxide, choline, betaine by UPLC–MS/MS in human plasma: An application in acute stroke patients

Trimethylamine-N-oxide (TMAO) is derived from the gut microbiome and tissues metabolism of dietary choline and betaine. These molecules are closely related to the development of cardiovascular and cerebrovascular diseases. A rapid, sensitive and accurate method has been developed and validated for the simultaneous determination of trimethylamine N-oxide (TMAO), choline and betaine in human plasma using d9-trimethylamine N-oxide (TMAO), d9-choline, d9-betaine as the internal standard (IS). After methanol precipitation with 10 μL plasma samples, the analytes were extracted and then separated on Amide column (2.1 × 100 mm, 1.7 μm, waters) with an isocratic elution program consisting of acetonitrile-water (containing 10 mM ammonium formate pH = 3.0) at a flow of 400 μL/min. The detection was achieved under the selected reaction monitoring (SRM) scan using positive electrospray ionization (ESI+) in 3 min. The mass transitions monitored were as follows: m/z 76.3 → 58.4 for TMAO, m/z 104.2 → 60.3 for choline, m/z 118.1 → 58.3 for betaine, m/z 85.1 → 66.3 for d9-TMAO, m/z 113.2 → 69.3 for d9-choline, and m/z 127.1 → 67.2 for d9-betaine, respectively. The method has been fully validated for specificity, lower limit of quantification, linearity, stability, intra- and inter-day accuracy and precision. This assay combines simple sample processing with a short run time and small plasma volumes, making it well suited for high-throughput routine clinical or research purposes. The newly developed method was successfully applied to the patients (n = 220) suffered from acute stroke, and the concentration of choline was firstly found to be closely related with the prognosis of these patients.

Compared to an Oatmeal Breakfast, Two Eggs/Day Increased Plasma Carotenoids and Choline without Increasing Trimethyl Amine N-Oxide Concentrations

ABSTRACTBackground: Habitual consumption of eggs has been hypothesized to positively modify biomarkers of cardiovascular disease risk through proposed antioxidant properties.Objectives: To examine this relationship, 50 young, healthy men and women were enrolled into a randomized crossover clinical intervention.Methods: Participants consumed either 2 eggs per day or one packet of oatmeal a day for 4 weeks, followed by a 3-week wash-out and crossed over to the alternate breakfast. Fasting blood samples and peripheral blood mononuclear cells (PBMCs) were collected at the end of each intervention period.Results: Increases in plasma large high-density lipoprotein (HDL) and large low-density lipoprotein (LDL) particle concentrations as measured by nuclear magnetic resonance were found following egg consumption (p < 0.001, p < 0.05), respectively, with increases in apolipoprotein concentration as well (p < 0.05). Though there was no difference in the intake of antioxidants lutein and zeaxanthin, a significant increase in plasma concentrations of these carotenoids was observed (p < 0.001) after egg consumption. There was no change in lecithin–cholesterol acyl transferase, cholesteryl ester transfer protein, or paroxanase-1 arylesterase activities between breakfast interventions. Dietary and plasma choline were both higher following egg consumption compared to oatmeal consumption (p < 0.001); however, there was no change in plasma trimethylamine N-oxide (TMAO) concentrations. Two eggs per day had no impact on PBMC gene expression related to cholesterol metabolism, oxidation, or TMAO production.Conclusions: These results suggest that compared to oatmeal, consumption of 2 eggs for breakfast provided increased plasma carotenoids and improved biomarkers of cardiovascular disease (CVD) risk while not affecting TMAO levels in this population.

Choline Supplementation During Pregnancy Protects Against Gestational Lipopolysaccharide-Induced Inflammatory Responses.

To estimate the effects and mechanisms of choline, an essential nutrient and a selective α7 nicotinic acetylcholine receptor (α7nAChR) agonist, on the prevention of symptoms and the effects on the cholinergic anti-inflammatory pathways (CAP) in a lipopolysaccharide (LPS)-induced inflammatory response in a rat model. Inflammation was induced by LPS treatment (1.0 μg LPS/kg body weight) on gestational day (GD) 14. Nonpregnant and pregnant Sprague Dawley rats were placed on a normal choline diet (1.1 g/kg) or supplemented choline diet (5.0 g/kg) from GDs 1 to 20. Systolic blood pressure (SBP), urinary albumin, and pregnancy outcomes were recorded. On GD 20, serum and placentas were assayed for cytokines. Western blots were used to determine the expression of placenta α7nAChR and components of the α7nAChR-CAP, including nuclear factor-κB (NF-κB) and protein kinase B (AKT). Immunohistochemistry was used to localize placental sites for the p65 subunit of NF-κB. Lipopolysaccharide significantly increased SBP and urinary albumin and decreased pregnancy outcomes, and these effects were partially reversed by higher choline treatment. Choline supplementation also significantly attenuated the LPS-induced increase in serum and placental inflammatory cytokines, decreased the expression of placental α7nAChR, lowered the activation of NF-κB signaling in placenta mononuclear cells, and inhibited placental AKT phosphorylation. This study confirms that LPS induces inflammatory conditions in pregnant rats and shows that choline supplementation protects against the inflammatory symptoms through its action on α7nAChR and CAP. These observations have important implications for the prevention and treatment of inflammatory responses associated with pregnancy.

An in vitro exploratory study of dietary strategies based on polyphenol-rich beverages, fruit juices and oils to control trimethylamine production in the colon.

Trimethylamine-N-oxide (TMAO) has been described as a new biomarker of cardiovascular disease (CVD), derived from gut microbial biotransformation of dietary choline and l-carnitine into trimethylamine (TMA) and subsequent hepatic oxidation. (Poly)phenols are among the dietary factors able to interfere with microbial enzymatic activity, possibly modulating TMA biotransformation at the gut level. The aim of this work was to investigate the in vitro biotransformation of choline and carnitine using faecal starters obtained from omnivorous and vegetarian subjects and the effect of (poly)phenol-rich foods on TMA production. Choline and l-carnitine were fermented with vegetarian or omnivorous faecal slurries, alone or in combination with 10 (poly)phenol-rich food items. TMA production from carnitine, but not from choline, was significantly lower when vegetarian faecal starters were used and, among the tested food items, blonde orange juice significantly reduced TMA formation during faecal biotransformation. Consequently, the main compounds of orange juice, namely phenolic compounds, terpenes, limonoids, organic acids and sugars, were tested individually. Sugars exerted the highest inhibitory effect on TMA production. Despite some limitations deriving from the applied in vitro model, this is the first work describing a possible role of some (poly)phenol-rich dietary products on the modulation of TMA colonic production. Free sugars were the main factor responsible for TMA inhibition, suggesting a potential beneficial role of colonic fermentation of carbohydrates in reducing TMA formation from its precursor molecules. This work opens new research directions to evaluate the effect of dietary fermentable fibre on TMA production and, potentially, on circulating TMAO levels.

Effect of long-term dietary sphingomyelin supplementation on atherosclerosis in mice.

Sphingomyelin (SM) levels in the circulation correlate positively with atherosclerosis burden. SM is a ubiquitous component of human diets, but it is unclear if dietary SM increases circulating SM levels. Dietary choline increases atherosclerosis by raising circulating trimethylamine N-oxide (TMAO) levels in mice and humans. As SM has a choline head group, we ask in this study if dietary SM accelerates atherosclerotic lesion development by increasing circulating SM and TMAO levels. Three studies were performed: (Study 1) C57BL/6 mice were maintained on a high fat diet with or without SM supplementation for 4 weeks prior to quantification of serum TMAO and SM levels; (Study 2) atherosclerosis was studied in apoE-/- mice after 16 weeks of a high fat diet without or with SM supplementation and (Study 3) apoE-/- mice were maintained on a chow diet for 19 weeks without or with SM supplementation and antibiotic treatment prior to quantification of atherosclerotic lesions and serum TMAO and SM levels. SM consumption did not increase circulating SM levels or atherosclerosis in high fat-fed apoE-/- mice. Serum TMAO levels in C57BL/6 mice were low and had no effect atherosclerosis lesion development. Dietary SM supplementation significantly reduced atherosclerotic lesion area in the aortic arch of chow-fed apoE-/- mice. This study establishes that dietary SM does not affect circulating SM levels or increase atherosclerosis in high fat-fed apoE-/- mice, but it is anti-atherogenic in chow-fed apoE-/- mice.

Association between choline intake during pregnancy and choline metabolism in parturients and infants

Objective To investigate the association between choline intake during pregnancy and choline metabolism in parturients and infants, and to analyze the difference in plasma choline level between premature and healthy full-term infants at birth. Methods Neonates who were born in the Department of Obstetrics of Xinhua Hospital Affiliated to Shanghai Jiao Tong University School of Medicine from September 2014 to May 2016 were enrolled. Demographic and clinical information of these neonates and their mothers were recorded. Dietary choline intake during pregnancy was determined on the second day after delivery using food frequency questionnaires. Free choline levels in plasma of all parturients and neonates were measured. Then, data were collected and analyzed by independent samples t-test, trend Chi-square test, Chi-square test and linear regression methods. Results (1) A total of 134 parturients and their neonates were enrolled in this study including 80 full-term infants and 54 premature infants. Compared with the full-term group, the parturients in the premature group had higher pregestational BMI (22.7±3.2 vs 20.8±2.3, t=-3.720) and less education background (trend χ2=4.020) (both P<0.05). (2) The food frequency questionnaires showed that 30.0% (24/80) of pregnant women in the full-term group reached adequate intake (AI) of choline during pregnancy, while only 11.1% (6/54) in the premature group did. (3) Plasma choline level in maternal peripheral blood was positively associated with maternal dietary choline intake during pregnancy (r=0.882, r²=0.778, P<0.001). Moreover, plasma choline level in the umbilical artery was closely related to maternal plasma choline level (r=0.965, r²=0.931, P<0.001). (4) The dietary choline intake during pregnancy, maternal plasma choline level and plasma choline level in umbilical artery in premature infant group were all lower than those of full-term group [(305.24±102.17) vs (386.16±121.36) mg/d, t=4.026; (5.23±0.79) vs (7.18±1.51) μmol/L, t=9.783; (20.68±3.52) vs (27.96±3.94) μmol/L, t=10.930; all P<0.01]. Conclusions Insufficient dietary intake of choline during pregnancy is common. The dietary choline intake during pregnancy in parturients of preterm delivery group is lower than that in full-term group. Dietary choline intake during pregnancy is related to plasma choline level in umbilical artery. Therefore, dietary guidance should be emphasized for pregnant and lactating women to increase the intake of foods rich in choline. It is also important to provide premature infants with choline in time to meets the needs of their development. Key words: Pregnancy; Choline; Metabolism; Dietary supplements; Diet; Infant, premature

Maternal Choline Supplementation Modulates Placental Nutrient Transport and Metabolism in Late Gestation of Mouse Pregnancy

BackgroundFetal growth is dependent on placental nutrient supply, which is influenced by placental perfusion and transporter abundance. Previous research indicates that adequate choline nutrition during pregnancy improves placental vascular development, supporting the hypothesis that choline may affect placental nutrient transport. ObjectiveThe present study sought to determine the impact of maternal choline supplementation (MCS) on placental nutrient transporter abundance and nutrient metabolism during late gestation. MethodsFemale non-Swiss albino mice were randomly assigned to the 1×, 2×, or 4× choline diet (1.4, 2.8, and 5.6 g choline chloride/kg diet, respectively) 5 d before mating (n = 16 dams/group). The placentas and fetuses were harvested on gestational day (E) 15.5 and E18.5. The placental abundance of macronutrient, choline, and acetylcholine transporters and glycogen metabolic enzymes, and the placental concentration of glycogen were quantified. Choline metabolites and docosahexaenoic acid (DHA) concentrations were measured in the placentas and/or fetal brains. Data were stratified by gestational day and fetal sex and were analyzed by using mixed linear models. ResultsAt E15.5, MCS downregulated the placental transcript and protein abundance of glucose transporter 1 (GLUT1) (−40% to −73%, P < 0.05) and the placental transcript abundance of glycogen-synthesizing enzymes (−24% to −50%, P ≤ 0.05). At E18.5, MCS upregulated GLUT3 protein abundance (+55%, P = 0.016) and the transcript abundance of glycogen-synthesizing enzymes only in the female placentas (+36% to +60%, P < 0.05), resulting in a doubling (P = 0.01) of the glycogen concentration. A higher placental transcript abundance of the transporters for DHA, choline, and acetylcholine was also detected in response to MCS, consequently altering their concentrations in the placentas or fetal brains (P ≤ 0.05). ConclusionsThese data suggest that MCS modulates placental nutrient transporter abundance and nutrient metabolism in late gestation of mouse pregnancy, with subsequent effects on nutrient supply for the developing fetus.

The Gut Microbial Metabolite Trimethylamine-N-Oxide Is Present in Human Cerebrospinal Fluid

Trimethylamine-N-oxide (TMAO) is a small organic molecule, derived from the intestinal and hepatic metabolism of dietary choline and carnitine. Although the involvement of TMAO in the framework of many chronic diseases has been recently described, no evidence on its putative role in the central nervous system has been provided. The aim of this study was to evaluate whether TMAO is present at detectable levels in human cerebrospinal fluid (CSF). CSF was collected for diagnostic purposes from 58 subjects by lumbar puncture and TMAO was quantified by using liquid chromatography coupled with multiple-reaction monitoring mass spectrometry. The molecule was detected in all samples, at concentrations ranging between 0.11 and 6.43 µmol/L. Further analysis on CSF revealed that a total of 22 subjects were affected by Alzheimer’s disease (AD), 16 were affected by non-AD related dementia, and 20 were affected by other neurological disorders. However, the stratification of TMAO levels according to the neurological diagnoses revealed no differences among the three groups. In conclusion, we provide the first evidence that TMAO can be assessed in human CSF, but the actual impact of this dietary metabolite in the patho-physiolgy of the central nervous system requires further study.

Four-week dietary supplementation with 10- and/or 15-fold basal choline caused decreased body weight in Sprague Dawley rats.

Choline is an essential nutrient utilized for phosphatidylcholine biosynthesis and lipoprotein packaging and secretion. Recently, choline supplementation has been used by athletes and the public for weight loss. However, the potential toxicological impact of choline dietary supplementation requires further investigation. This study examined the effects of choline dietary supplementation in Sprague Dawley rats for 4 weeks. Rats were fed diets containing basal choline levels (control) or 5-, 10-, or 15-fold (5×, 10×, or 15×) basal diet concentration. In groups fed choline-supplemented diets, there were no toxicologically relevant findings in clinical observations, food intake, clinical chemistry, liver weights, or liver histopathology. However, decreased mean body weights (8.5-10.2%) and body weight gains (24-31%) were noted for the 10× choline-supplemented (females only) and 15× choline-supplemented (both sexes) groups relative to the control groups from day 3 onward. These body weight effects were not related to a persistent reduction in average food intake. Serum cholesterol was increased in the 15× choline-supplemented male rats relative to the controls, an expected effect of choline supplementation; however, there were no changes in the serum cholesterol of female rats. Serum choline concentrations were increased in female rats relative to the male rats across all treatment groups. The maximum tolerated dose for male and female rats were the 15× and 10× choline supplements, respectively, based on decreased mean body weight and body weight gains. This study supported the conclusions of a clinical trial that showed a high choline diet can decrease body weight in humans.

Pressing the trimethylamine N-oxide narrative

We read with interest the correspondence titled “Gut Microbe-Generated Trimethylamine N-Oxide from Dietary Choline is Prothrombotic in Subjects”, published by Zhu et al . in the journal Circulation (1). This work builds on an emerging body of evidence which has suggested a role for the gut-derived metabolite, trimethylamine N -oxide (TMAO), in the development of cardiovascular disease (CVD). This current study is an important clinical test of hypotheses generated from innovative preclinical and observational evidence, and seeks to build upon the nature of TMAO’s relationship with CVD risk in humans. To assess this relationship, Zhu et al . enrolled 18 subjects (10 omnivores and 8 vegetarians) and provided choline bitartrate as a supplement (450 mg choline/day) for 2 months. Following choline supplementation, fasting plasma TMAO concentrations increased and were associated with increased ADP-induced platelet aggregation, an effect that was partially blunted by aspirin.

Perfluorooctane Sulfonate-Induced Hepatic Steatosis in Male Sprague Dawley Rats Is Not Attenuated by Dietary Choline Supplementation.

Perfluorooctane sulfonate (PFOS) is an environmentally persistent chemical. Dietary 100 ppm PFOS fed to male mice and rats for 4 weeks caused hepatic steatosis through an unknown mechanism. Choline deficient diets can cause hepatic steatosis. A hepatic choline:PFOS ion complex was hypothesized to cause this effect in mice. This study tested whether dietary choline supplementation attenuates PFOS-induced hepatic steatosis in rats. Sprague Dawley rats (12/sex/group) were fed control, choline supplemented (CS), 100 ppm PFOS, or 100 ppm PFOS + CS diets for 3 weeks. Male rats fed both PFOS-containing diets had decreased serum cholesterol and triglycerides (TGs) on days 9, 16, and/or 23 and increased hepatic free fatty acids and TG (ie, steatosis). Female rats fed both PFOS diets had decreased serum cholesterol on days 9 and 16 and decreased hepatic free fatty acid and TG at termination (ie, no steatosis). Liver PFOS concentrations were similar for both sexes. Liver choline concentrations were increased in male rats fed PFOS (±CS), but the increase was lower in the PFOS + CS group. Female liver choline concentrations were not altered by any diet. These findings demonstrate a clear sex-related difference in PFOS-induced hepatic steatosis in the rat. Additional evaluated mechanisms (ie, nuclear receptor activation, mRNA upregulation, and choline kinase activity inhibition) did not appear to be involved in the hepatic steatosis. Dietary PFOS (100 ppm) induced hepatic steatosis in male, but not female, rats that was not attenuated by choline supplementation. The mechanism of lipid accumulation and the sex-related differences warrant further investigation.

Dietary choline derived TMAO: new role in thrombosis

Increasing pieces of evidence link the intestinal gut microbiota and its produced metabolites to many chronic diseases, including cardiovascular and atherosclerotic diseases (1).