Specific circulating miRNA are associated with plasma lipids in a healthy American cohort.

Low-density lipoprotein cholesterol (LDL-c) is both a therapeutic target and a risk factor for cardiovascular disease (CVD). MicroRNA (miRNA) have been shown to regulate cholesterol homeostasis, and miRNA in blood circulation have been linked to hypercholesterolemia. However, few studies to date have associated miRNA with phenotypes like LDL-c in a healthy population. To this end, we analyzed circulating miRNA in relation to LDL-c in a healthy cohort of 353 participants using two separate bioinformatic approaches. The first approach found that miR-15b-5p and miR-16-5p were upregulated in individuals with at-risk levels of LDL-c. The second approach identified two miRNA clusters, one that positively and a second that negatively, correlated with LDL-c. Included in the cluster that positively correlated with LDL-c were miR-15b-5p and miR-16-5p as well as other miRNA from the miR-15/107, miR-30, and let-7 families. Cross-species analyses suggested that several miRNA that associated with LDL-c are conserved between mice and humans. Finally, we examined the influence of diet on circulating miRNA. Our results robustly linked circulating miRNA with LDL-c suggesting that miRNA could be used as biomarkers for hypercholesterolemia or targets for developing cholesterol-lowering drugs.

Analysis of strain, sex, and diet-dependent modulation of gut microbiota reveals candidate keystone organisms driving microbial diversity in response to American and ketogenic diets.

BACKGROUND: The gut microbiota is modulated by a combination of diet, host genetics, and sex effects. The magnitude of these effects and interactions among them is important to understanding inter-individual variability in gut microbiota. In a previous study, mouse strain-specific responses to American and ketogenic diets were observed along with several QTLs for metabolic traits. In the current study, we searched for genetic variants underlying differences in the gut microbiota in response to American and ketogenic diets, which are high in fat and vary in carbohydrate composition, between C57BL/6 J (B6) and FVB/NJ (FVB) mouse strains. RESULTS: Genetic mapping of microbial features revealed 18 loci under the QTL model (i.e., marginal effects that are not specific to diet or sex), 12 loci under the QTL by diet model, and 1 locus under the QTL by sex model. Multiple metabolic and microbial features map to the distal part of Chr 1 and Chr 16 along with eigenvectors extracted from principal coordinate analysis of measures of ß-diversity. Bilophila, Ruminiclostridium 9, and Rikenella (Chr 1) were identified as sex- and diet-independent QTL candidate keystone organisms, and Parabacteroides (Chr 16) was identified as a diet-specific, candidate keystone organism in confirmatory factor analyses of traits mapping to these regions. For many microbial features, irrespective of which QTL model was used, diet or the interaction between diet and a genotype were the strongest predictors of the abundance of each microbial trait. Sex, while important to the analyses, was not as strong of a predictor for microbial abundances. CONCLUSIONS: These results demonstrate that sex, diet, and genetic background have different magnitudes of effects on inter-individual differences in gut microbiota. Therefore, Precision Nutrition through the integration of genetic variation, microbiota, and sex affecting microbiota variation will be important to predict response to diets varying in carbohydrate composition. Video Abstract.

Precision pharmacological reversal of strain-specific diet-induced metabolic syndrome in mice informed by epigenetic and transcriptional regulation.

Diet-related metabolic syndrome is the largest contributor to adverse health in the United States. However, the study of gene-environment interactions and their epigenomic and transcriptomic integration is complicated by the lack of environmental and genetic control in humans that is possible in mouse models. Here we exposed three mouse strains, C57BL/6J (BL6), A/J, and NOD/ShiLtJ (NOD), to a high-fat, high-carbohydrate diet, leading to varying degrees of metabolic syndrome. We then performed transcriptomic and genome-wide DNA methylation analyses for each strain and found overlapping but also highly divergent changes in gene expression and methylation upstream of the discordant metabolic phenotypes. Strain-specific pathway analysis of dietary effects revealed a dysregulation of cholesterol biosynthesis common to all three strains but distinct regulatory networks driving this dysregulation. This suggests a strategy for strain-specific targeted pharmacologic intervention of these upstream regulators informed by epigenetic and transcriptional regulation. As a pilot study, we administered the drug GW4064 to target one of these genotype-dependent networks, the farnesoid X receptor pathway, and found that GW4064 exerts strain-specific protection against dietary effects in BL6, as predicted by our transcriptomic analysis. Furthermore, GW4064 treatment induced inflammatory-related gene expression changes in NOD, indicating a strain-specific effect in its associated toxicities as well as its therapeutic efficacy. This pilot study demonstrates the potential efficacy of precision therapeutics for genotype-informed dietary metabolic intervention and a mouse platform for guiding this approach.

Integrative analysis of hepatic transcriptional profiles reveals genetic regulation of atherosclerosis in hyperlipidemic Diversity Outbred-F1 mice.

Atherogenesis is an insipidus but precipitating process leading to serious consequences of many cardiovascular diseases (CVD). Numerous genetic loci contributing to atherosclerosis have been identified in human genome-wide association studies, but these studies have limitations in the ability to control environmental factors and to decipher cause/effect relationships. To assess the power of hyperlipidemic Diversity Outbred (DO) mice in facilitating quantitative trait loci (QTL) analysis of complex traits, we generated a high-resolution genetic panel of atherosclerosis susceptible (DO-F1) mouse cohort by crossing 200 DO females with C57BL/6J males carrying two human genes: encoding apolipoprotein E3-Leiden and cholesterol ester transfer protein. We examined atherosclerotic traits including plasma lipids and glucose in the 235 female and 226 male progeny before and after 16 weeks of a high-fat/cholesterol diet, and aortic plaque size at 24 weeks. We also assessed the liver transcriptome using RNA-sequencing. Our QTL mapping for atherosclerotic traits identified one previously reported female-specific QTL on Chr10 with a narrower interval of 22.73 to 30.80 Mb, and one novel male-specific QTL at 31.89 to 40.25 Mb on Chr19. Liver transcription levels of several genes within each QTL were highly correlated with the atherogenic traits. A majority of these candidates have already known atherogenic potential in humans and/or mice, but integrative QTL, eQTL, and correlation analyses further pointed Ptprk as a major candidate of the Chr10 QTL, while Pten and Cyp2c67 of the Chr19 QTL in our DO-F1 cohort. Finally, through additional analyses of RNA-seq data we identified genetic regulation of hepatic transcription factors, including Nr1h3, contributes to atherogenesis in this cohort. Thus, an integrative approach using DO-F1 mice effectively validates the influence of genetic factors on atherosclerosis in DO mice and suggests an opportunity to discover therapeutics in the setting of hyperlipidemia.

Randomized crossover clinical trial of coenzyme Q10 and nicotinamide riboside in chronic kidney disease.

BackgroundCurrent studies suggest mitochondrial dysfunction is a major contributor to impaired physical performance and exercise intolerance in chronic kidney disease (CKD). We conducted a clinical trial of coenzyme Q10 (CoQ10) and nicotinamide riboside (NR) to determine their impact on exercise tolerance and metabolic profile in patients with CKD.MethodsWe conducted a randomized, placebo-controlled, double-blind, crossover trial comparing CoQ10, NR, and placebo in 25 patients with an estimated glomerular filtration rate (eGFR) of less than 60mL/min/1.73 m2. Participants received NR (1,000 mg/day), CoQ10 (1,200 mg/day), or placebo for 6 weeks each. The primary outcomes were aerobic capacity measured by peak rate of oxygen consumption (VO2 peak) and work efficiency measured using graded cycle ergometry testing. We performed semitargeted plasma metabolomics and lipidomics.ResultsParticipant mean age was 61.0 ± 11.6 years and mean eGFR was 36.9 ± 9.2 mL/min/1.73 m2. Compared with placebo, we found no differences in VO2 peak (P = 0.30, 0.17), total work (P = 0.47, 0.77), and total work efficiency (P = 0.46, 0.55) after NR or CoQ10 supplementation. NR decreased submaximal VO2 at 30 W (P = 0.03) and VO2 at 60 W (P = 0.07) compared with placebo. No changes in eGFR were observed after NR or CoQ10 treatment (P = 0.14, 0.88). CoQ10 increased free fatty acids and decreased complex medium- and long-chain triglycerides. NR supplementation significantly altered TCA cycle intermediates and glutamate that were involved in reactions that exclusively use NAD+ and NADP+ as cofactors. NR decreased a broad range of lipid groups including triglycerides and ceramides.ConclusionsSix weeks of treatment with NR or CoQ10 improved markers of systemic mitochondrial metabolism and lipid profiles but did not improve VO2 peak or total work efficiency.Trial registrationClinicalTrials.gov NCT03579693.FundingNational Institutes of Diabetes and Digestive and Kidney Diseases (grants R01 DK101509, R03 DK114502, R01 DK125794, and R01 DK101509).

Precision pharmacological reversal of genotype-specific diet-induced metabolic syndrome in mice informed by transcriptional regulation.

Diet-related metabolic syndrome is the largest contributor to adverse health in the United States. However, the study of gene-environment interactions and their epigenomic and transcriptomic integration is complicated by the lack of environmental and genetic control in humans that is possible in mouse models. Here we exposed three mouse strains, C57BL/6J (BL6), A/J, and NOD/ShiLtJ (NOD), to a high-fat high-carbohydrate diet, leading to varying degrees of metabolic syndrome. We then performed transcriptomic and genomic DNA methylation analyses and found overlapping but also highly divergent changes in gene expression and methylation upstream of the discordant metabolic phenotypes. Strain-specific pathway analysis of dietary effects reveals a dysregulation of cholesterol biosynthesis common to all three strains but distinct regulatory networks driving this dysregulation. This suggests a strategy for strain-specific targeted pharmacologic intervention of these upstream regulators informed by transcriptional regulation. As a pilot study, we administered the drug GW4064 to target one of these genotype-dependent networks, the Farnesoid X receptor pathway, and found that GW4064 exerts genotype-specific protection against dietary effects in BL6, as predicted by our transcriptomic analysis, as well as increased inflammatory-related gene expression changes in NOD. This pilot study demonstrates the potential efficacy of precision therapeutics for genotype-informed dietary metabolic intervention, and a mouse platform for guiding this approach.

Analysis of strain, sex, and diet-dependent modulation of gut microbiota reveals candidate keystone organisms driving microbial diversity in response to American and ketogenic diets.

Background The gut microbiota is modulated by a combination of diet, host genetics, and sex effects. The magnitude of these effects and interactions among them is important to understanding inter-individual variability in gut microbiota. In a previous study, mouse strain-specific responses to American and ketogenic diets were observed along with several QTL for metabolic traits. In the current study, we searched for genetic variants underlying differences in the gut microbiota in response to American and ketogenic diets, which are high in fat and vary in carbohydrate composition, between C57BL/6J (B6) and FVB/NJ (FVB) mouse strains. Results Genetic mapping of microbial features revealed 18 loci under the QTL model (i.e., marginal effects that are not specific to diet or sex), 12 loci under the QTL by diet model, and 1 locus under the QTL by sex model. Multiple metabolic and microbial features map to the distal part of Chr 1 and Chr 16 along with eigenvectors extracted from principal coordinate analysis of measures of ß-diversity. Bilophila , Ruminiclostridium 9 , and Rikenella (Chr 1) were identified as sex and diet independent QTL candidate keystone organisms and Rikenelleceae RC9 Gut Group (Chr 16) was identified as a diet-specific, candidate keystone organism in confirmatory factor analyses of traits mapping to these regions. For many microbial features, irrespective of which QTL model was used, diet or the interaction between diet and a genotype were the strongest predictors of the abundance of each microbial trait. Sex, while important to the analyses, was not as strong of a predictor for microbial abundances. Conclusions These results demonstrate that sex, diet, and genetic background have different magnitudes of effects on inter-individual differences in gut microbiota. Therefore, Precision Nutrition through the integration of genetic variation, microbiota, and sex affecting microbiota variation will be important to predict response to diets varying in carbohydrate composition.

Trimethylamine N-Oxide Response to a Mixed Macronutrient Tolerance Test in a Cohort of Healthy United States Adults.

Plasma trimethylamine n-oxide (TMAO) concentration increases in responses to feeding TMAO, choline, phosphatidylcholine, L-carnitine, and betaine but it is unknown whether concentrations change following a mixed macronutrient tolerance test (MMTT) with limited amounts of TMAO precursors. In this proof-of-concept study, we provided healthy female and male adults (n = 97) ranging in age (18-65 years) and BMI (18-44 kg/m2) a MMTT (60% fat, 25% sucrose; 42% of a standard 2000 kilo calorie diet) and recorded their metabolic response at fasting and at 30 min, 3 h, and 6 h postprandially. We quantified total exposure to TMAO (AUC-TMAO) and classified individuals by the blood draw at which they experienced their maximal TMAO concentration (TMAO-response groups). We related AUC-TMAO to the 16S rRNA microbiome, to two SNPs in the exons of the FMO3 gene (rs2266782, G>A, p.Glu158Lys; and rs2266780, A>G, p.Glu308Gly), and to a priori plasma metabolites. We observed varying TMAO responses (timing and magnitude) and identified a sex by age interaction such that AUC-TMAO increased with age in females but not in males (p-value = 0.0112). Few relationships between AUC-TMAO and the fecal microbiome and FMO3 genotype were identified. We observed a strong correlation between AUC-TMAO and TNF-α that depended on TMAO-response group. These findings promote precision nutrition and have important ramifications for the eating behavior of adults who could benefit from reducing TMAO exposure, and for understanding factors that generate plasma TMAO.

The association between plasma choline, growth and neurodevelopment among Malawian children aged 6-15 months enroled in an egg intervention trial.

Choline is an essential micronutrient that may influence growth and development; however, few studies have examined postnatal choline status and children's growth and development in low- and middle-income countries. The aim of this observational analysis was to examine associations of plasma choline with growth and development among Malawian children aged 6-15 months enrolled in an egg intervention trial. Plasma choline and related metabolites (betaine, dimethylglycine and trimethylamine N-oxide) were measured at baseline and 6-month follow-up, along with anthropometric (length, weight, head circumference) and developmental assessments (the Malawi Developmental Assessment Tool [MDAT], the Infant Orienting with Attention task [IOWA], a visual paired comparison [VPC] task and an elicited imitation [EI] task). In cross-sectional covariate-adjusted models, each 1 SD higher plasma choline was associated with lower length-for-age z-score (-0.09 SD [95% confidence interval, CI -0.17 to -0.01]), slower IOWA response time (8.84 ms [1.66-16.03]) and faster processing speed on the VPC task (-203.5 ms [-366.2 to -40.7]). In predictive models, baseline plasma choline was negatively associated with MDAT fine motor z-score at 6-month follow-up (-0.13 SD [-0.22 to -0.04]). There were no other significant associations of plasma choline with child measures. Similarly, associations of choline metabolites with growth and development were null except higher trimethylamine N-oxide was associated with slower information processing on the VPC task and higher memory scores on the EI task. In this cohort of children with low dietary choline intake, we conclude that there were no strong or consistent associations between plasma choline and growth and development.

Chronic Kidney Disease is Associated With Attenuated Plasma Metabolome Response to Oral Glucose Tolerance Testing.

OBJECTIVE: Chronic kidney disease (CKD) is associated with decreased anabolic response to insulin contributing to protein-energy wasting. Targeted metabolic profiling of oral glucose tolerance testing (OGTT) may help identify metabolic pathways contributing to disruptions to insulin response in CKD. METHODS: Using targeted metabolic profiling, we studied the plasma metabolome response in 41 moderate-to-severe nondiabetic CKD patients and 20 healthy controls at fasting and 2 hours after an oral glucose load. We used linear mixed modeling with random intercepts, adjusting for age, gender, race/ethnicity, body weight, and batch to assess heterogeneity in response to OGTT by CKD status. RESULTS: Mean estimated glomerular filtration rate among CKD participants was 38.9 ± 12.7 mL/min per 1.73 m2 compared to 87.2 ± 17.7 mL/min per 1.73 m2 among controls. Glucose ingestion induced an anabolic response resulting in increased glycolysis products and a reduction in a wide range of metabolites including amino acids, tricarboxylic acid cycle intermediates, and purine nucleotides compared to fasting. Participants with CKD demonstrated a blunted anabolic response to OGTT evidenced by significant changes in 13 metabolites compared to controls. The attenuated metabolome response predominant involved mitochondrial energy metabolism, vitamin B family, and purine nucleotides. Compared to controls, CKD participants had elevated lactate:pyruvate (L:P) ratio and decreased guanosine diphosphate:guanosine triphosphate ratio during OGTT. CONCLUSION: Metabolic profiling of OGTT response suggests a broad disruption of mitochondrial energy metabolism in CKD patients. These findings motivate further investigation into the impact of insulin sensitizers and mitochondrial targeted therapeutics on energy metabolism in patients with nondiabetic CKD.

Impaired incretin homeostasis in non-diabetic moderate-severe CKD.

Background: Incretins are regulators of insulin secretion and glucose homeostasis that are metabolized by dipeptidyl peptidase-4 (DPP-4). Moderate-severe CKD may modify incretin release, metabolism, or response. Methods: We performed 2-hour oral glucose tolerance testing (OGTT) in 59 people with non-diabetic CKD (eGFR<60 ml/min per 1.73 m2) and 39 matched controls. We measured total (tAUC) and incremental (iAUC) area under the curve of plasma total glucagon-like peptide-1 (GLP-1) and total glucose-dependent insulinotropic polypeptide (GIP). Fasting DPP-4 levels and activity were measured. Linear regression was used to adjust for demographic, body composition, and lifestyle factors. Results: Mean eGFR was 38 ±13 and 89 ±17ml/min per 1.73 m2 in CKD and controls. GLP-1 iAUC and GIP iAUC were higher in CKD than controls with a mean of 1531 ±1452 versus 1364 ±1484 pMxmin, and 62370 ±33453 versus 42365 ±25061 pgxmin/ml, respectively. After adjustment, CKD was associated with 15271 pMxmin/ml greater GIP iAUC (95% CI 387, 30154) compared to controls. Adjustment for covariates attenuated associations of CKD with higher GLP-1 iAUC (adjusted difference, 122, 95% CI -619, 864). Plasma glucagon levels were higher at 30 minutes (mean difference, 1.6, 95% CI 0.3, 2.8 mg/dl) and 120 minutes (mean difference, 0.84, 95% CI 0.2, 1.5 mg/dl) in CKD compared to controls. There were no differences in insulin levels or plasma DPP-4 activity or levels between groups. Conclusion: Incretin response to oral glucose is preserved or augmented in moderate-severe CKD, without apparent differences in circulating DPP-4 concentration or activity. However, neither insulin secretion nor glucagon suppression are enhanced.

SNPs in apolipoproteins contribute to sex-dependent differences in blood lipids before and after a high-fat dietary challenge in healthy U.S. adults.

BACKGROUND: The effect of genetic polymorphisms on fasting blood lipid levels have been widely studied but the effects of these within the context of a high-fat meal challenge remain less characterized. The current study aimed to investigate the association of SNPs in lipoprotein-related genes with blood lipid profiles in healthy adults in the U.S. METHODS: Subjects (n = 393) between 18-66 years of age with BMIs ranging from 18.5-45 kg/m2 were enrolled the cross-sectional Nutritional Phenotyping Study. Among them, 349 subjects (men: 48%; women: 52%) gave consent for genotyping. SNPs in APOA5, APOB, APOC3, APOE, and LDLR were assessed. The association between lipid markers and genotypes was tested separately for each SNP with analysis of variance (ANOVA), adjusted for sex, age, and BMI. We also examined two-factor interactions between SNPs and sex, age, or BMI. RESULTS: Women carrying the C allele of rs3135506 in APOA5 or men carrying the C allele of rs429358 in APOE had reduced HDL-cholesterol levels during fasting and postprandially. The C allele in APOE was also correlated to increased LDL-C levels. The TT genotype of rs2854116 in APOC3 was associated with elevated total cholesterol. Additive effect of the risk alleles of APOA5 and APOE or APOC3 and APOE was detected. Nevertheless, the tested SNPs had little impact on the postprandial triglyceride responses to the high-fat challenge meal. We found no significant effects of SNPs in APOB (rs1042034) or LDLR (rs2228671) on triglycerides, cholesterol, or free fatty acid levels. CONCLUSIONS: In healthy adults, fasting and postprandial cholesterol levels are strongly correlated with the tested APOA5, APOE, and APOC3 genotypes. Sex contributes to the genetic impact of the tested SNPs on lipid profiles. TRIAL REGISTRATION: ClinicalTrials.gov, NCT02367287. Registered February 20, 2015, https://clinicaltrials.gov/ct2/show/NCT02367287 .

Gut microbiota and host genetics modulate the effect of diverse diet patterns on metabolic health.

Metabolic diseases are major public health issues worldwide and are responsible for disproportionately higher healthcare costs and increased complications of many diseases including SARS-CoV-2 infection. The Western Diet (WD) specifically is believed to be a major contributor to the global metabolic disease epidemic. In contrast, the Mediterranean diet (MeD), Ketogenic diet (KD), and Japanese diet (JD) are often considered beneficial for metabolic health. Yet, there is a growing appreciation that the effect of diet on metabolic health varies depending on several factors including host genetics. Additionally, poor metabolic health has also been attributed to altered gut microbial composition and/or function. To understand the complex relationship between host genetics, gut microbiota, and dietary patterns, we treated four widely used metabolically diverse inbred mouse strains (A/J, C57BL/6J, FVB/NJ, and NOD/ShiLtJ) with four human-relevant diets (MeD, JD, KD, WD), and a control mouse chow from 6 weeks to 30 weeks of age. We found that diet-induced alteration of gut microbiota (α-diversity, ß-diversity, and abundance of several bacteria including Bifidobacterium, Ruminococcus, Turicibacter, Faecalibaculum, and Akkermansia) is significantly modified by host genetics. In addition, depending on the gut microbiota, the same diet could have different metabolic health effects. Our study also revealed that C57BL/6J mice are more susceptible to altered gut microbiota compared to other strains in this study indicating that host genetics is an important modulator of the diet-microbiota-metabolic health axis. Overall, our study demonstrated complex interactions between host genetics, gut microbiota, and diet on metabolic health; indicating the need to consider both host genetics and the gut microbiota in the development of new and more effective precision nutrition strategies to improve metabolic health.

Altered macronutrient composition and genetics influence the complex transcriptional network associated with adiposity in the Collaborative Cross.

BACKGROUND: Obesity is a serious disease with a complex etiology characterized by overaccumulation of adiposity resulting in detrimental health outcomes. Given the liver's critical role in the biological processes that attenuate adiposity accumulation, elucidating the influence of genetics and dietary patterns on hepatic gene expression is fundamental for improving methods of obesity prevention and treatment. To determine how genetics and diet impact obesity development, mice from 22 strains of the genetically diverse recombinant inbred Collaborative Cross (CC) mouse panel were challenged to either a high-protein or high-fat high-sucrose diet, followed by extensive phenotyping and analysis of hepatic gene expression. RESULTS: Over 1000 genes differentially expressed by perturbed dietary macronutrient composition were enriched for biological processes related to metabolic pathways. Additionally, over 9000 genes were differentially expressed by strain and enriched for biological process involved in cell adhesion and signaling. Weighted gene co-expression network analysis identified multiple gene clusters (modules) associated with body fat % whose average expression levels were influenced by both dietary macronutrient composition and genetics. Each module was enriched for distinct types of biological functions. CONCLUSIONS: Genetic background affected hepatic gene expression in the CC overall, but diet macronutrient differences also altered expression of a specific subset of genes. Changes in macronutrient composition altered gene expression related to metabolic processes, while genetic background heavily influenced a broad range of cellular functions and processes irrespective of adiposity. Understanding the individual role of macronutrient composition, genetics, and their interaction is critical to developing therapeutic strategies and policy recommendations for precision nutrition.

Diet, Fecal Microbiome, and Trimethylamine N-Oxide in a Cohort of Metabolically Healthy United States Adults.

TMAO is elevated in individuals with cardiometabolic diseases, but it is unknown whether the metabolite is a biomarker of concern in healthy individuals. We conducted a cross-sectional study in metabolically healthy adults aged 18-66 years with BMI 18-44 kg/m2 and assessed the relationship between TMAO and diet, the fecal microbiome, and cardiometabolic risk factors. TMAO was measured in fasted plasma samples by liquid chromatography mass spectrometry. The fecal microbiome was assessed by 16S ribosomal RNA sequencing and recent food intake was captured by multiple ASA24 dietary recalls. Endothelial function was assessed via EndoPAT. Descriptive statistics were computed by fasting plasma TMAO tertiles and evaluated by ANOVA and Tukey's post-hoc test. Multiple linear regression was used to assess the relationship between plasma TMAO and dietary food intake and metabolic health parameters. TMAO concentrations were not associated with average intake of animal protein foods, fruits, vegetables, dairy, or grains. TMAO was related to the fecal microbiome and the genera Butyribrio, Roseburia, Coprobaciullus, and Catenibacterium were enriched in individuals in the lowest versus the highest TMAO tertile. TMAO was positively associated with α-diversity and compositional differences were identified between groups. TMAO was not associated with classic cardiovascular risk factors in the healthy cohort. Similarly, endothelial function was not related to fasting TMAO, whereas the inflammatory marker TNF-α was significantly associated. Fasting plasma TMAO may not be a metabolite of concern in generally healthy adults unmedicated for chronic disease. Prospective studies in healthy individuals are necessary.

Obesogenic and diabetic effects of CD44 in mice are sexually dimorphic and dependent on genetic background.

INTRODUCTION: CD44 is a candidate gene for obesity and diabetes development and may be a critical mediator of a systemic inflammation associated with obesity and diabetes. METHODS: We investigated the relationship of CD44 with obesity in CD44-deficient mice challenged with a high-fat diet. RESULTS: In mice fed a diet high in fat, cholesterol, and sucrose for 12 weeks fat mass accumulation was reduced in CD44-deficient mice bred onto both a C57BL/6J and the naturally TLR deficient C3H/HeJ background. Reduced fat mass could not be attributed to lower food intake or an increase in energy expenditure as measured by indirect calorimetry. However, we observed a 40-60% lower mRNA expression of the inflammation markers, F4/80, CD11b, TNF-α, and CD14, in adipose tissue of CD44-deficient mice on the C57BL/6J background but not the C3H/HeJ background, perhaps indicating that alternative factors may be affecting adiposity in this model. Measures of hepatic steatosis and insulin sensitivity were improved in CD44-deficient mice on a C57BL/6J but not in the C3H/HeJ mice. These results were highly sexually dimorphic as there were no detectable effects of CD44 inactivation in female mice on a C57BL/6 J or C3H/HeJ background. CONCLUSION: CD44 was associated with adiposity, liver fat, and glucose in male mice. However, the effects of CD44 on obesity may be independent of TLR4 signaling.

Plasma Choline Concentration Was Not Increased After a 6-Month Egg Intervention in 6-9-Month-Old Malawian Children: Results from a Randomized Controlled Trial.

BACKGROUND: Eggs are a rich source of choline, an essential nutrient important for child growth and development. In a randomized trial of 1 egg/d in young children in Ecuador, an egg intervention led to significant improvements in growth, which were partially mediated by increased plasma choline concentration. A similar trial in Malawi (clinicaltrials.gov: NCT03385252) found little improvement in child growth or development. OBJECTIVES: We aimed to evaluate the effect of 1 egg/d for 6 mo on plasma choline concentrations in Malawian children enrolled in a randomized trial. METHODS: Infants aged 6-9 mo in rural Malawi were randomly assigned to receive 1 egg/d (n = 331) or serve as a nonintervention control (n = 329) for 6 mo. Anthropometric, developmental, and dietary data were collected at baseline and 6-mo follow-up, along with a blood draw. Plasma choline, betaine, dimethylglycine, trimethylamine N-oxide (TMAO), and DHA were measured at both time points using ultrahigh performance liquid chromatography-tandem MS (n = 200 per group). Linear regression analysis was used to determine the difference in plasma choline and related metabolites between groups after 6 mo of intervention. RESULTS: Plasma choline, betaine, dimethylglycine, and DHA concentrations did not differ between groups at 6-mo follow-up. Plasma TMAO was significantly (26%; 95% CI: 7%, 48%) higher in the egg intervention group in a fully adjusted model. CONCLUSIONS: Provision of 1 egg/d for 6 mo did not result in increases in plasma choline or related metabolites, except TMAO. This could partially explain the lack of effect on growth and development. Additional interventions are needed to improve choline status, growth, and development in this population.

Sequence meets function-microbiota and cardiovascular disease.

The discovery that gut-microbiota plays a profound role in human health has opened a new avenue of basic and clinical research. Application of ecological approaches where the bacterial 16S rRNA gene is queried has provided a number of candidate bacteria associated with coronary artery disease and hypertension. We examine the associations between gut microbiota and a variety of cardiovascular disease (CVD) including atherosclerosis, coronary artery disease, and blood pressure. These approaches are associative in nature and there is now increasing interest in identifying the mechanisms underlying these associations. We discuss three potential mechanisms including: gut permeability and endotoxemia, increased immune system activation, and microbial derived metabolites. In addition to discussing these potential mechanisms we highlight current studies manipulating the gut microbiota or microbial metabolites to move beyond sequence-based association studies. The goal of these mechanistic studies is to determine the mode of action by which the gut microbiota may affect disease susceptibility and severity. Importantly, the gut microbiota appears to have a significant effect on host metabolism and CVD by producing metabolites entering the host circulatory system such as short-chain fatty acids and trimethylamine N-Oxide. Therefore, the intersection of metabolomics and microbiota research may yield novel targets to reduce disease susceptibility. Finally, we discuss approaches to demonstrate causality such as specific diet changes, inhibition of microbial pathways, and fecal microbiota transplant.

Effects of a diet based on the Dietary Guidelines on vascular health and TMAO in women with cardiometabolic risk factors.

BACKGROUND AND AIMS: Recent evidence links trimethylamine oxide (TMAO) to endothelial dysfunction, an early indicator of cardiovascular disease. We aimed to determine whether short-term consumption of a diet patterned after the 2010 Dietary Guidelines for Americans (DGA) would affect endothelial function, plasma TMAO concentrations, and cardiovascular disease risk, differently than a typical American Diet (TAD). METHODS AND RESULTS: An 8-wk controlled feeding trial was conducted in overweight/obese women pre-screened for insulin resistance and/or dyslipidemia. Women were randomized to a DGA or TAD group (n = 22/group). At wk0 (pre-intervention) and wk8 (post-intervention) vascular age was calculated; endothelial function (reactive hyperemia index (RHI)) and augmentation index (AI@75) were measured using EndoPAT, and plasma TMAO was measured by LC-MS/MS. Vascular age was reduced in DGA at wk8 compared to wk0 but TAD wk8 was not different from wk0 (DGA wk0: 54.2 ± 4.0 vs. wk8: 50.5 ± 3.1 (p = 0.05), vs. TAD wk8: 47.7 ± 2.3). Plasma TMAO concentrations, RHI, and AI@75 were not different between groups or weeks. CONCLUSION: Consumption of a diet based on the 2010 Dietary Guidelines for Americans for 8 weeks did not improve endothelial function or reduce plasma TMAO. CLINICALTRIALS.GOV: NCT02298725.

Genetic architecture modulates diet-induced hepatic mRNA and miRNA expression profiles in Diversity Outbred mice.

Genetic approaches in model organisms have consistently demonstrated that molecular traits such as gene expression are under genetic regulation, similar to clinical traits. The resulting expression quantitative trait loci (eQTL) have revolutionized our understanding of genetic regulation and identified numerous candidate genes for clinically relevant traits. More recently, these analyses have been extended to other molecular traits such as protein abundance, metabolite levels, and miRNA expression. Here, we performed global hepatic eQTL and microRNA expression quantitative trait loci (mirQTL) analysis in a population of Diversity Outbred mice fed two different diets. We identified several key features of eQTL and mirQTL, namely differences in the mode of genetic regulation (cis or trans) between mRNA and miRNA. Approximately 50% of mirQTL are regulated by a trans-acting factor, compared to ∼25% of eQTL. We note differences in the heritability of mRNA and miRNA expression and variance explained by each eQTL or mirQTL. In general, cis-acting variants affecting mRNA or miRNA expression explain more phenotypic variance than trans-acting variants. Finally, we investigated the effect of diet on the genetic architecture of eQTL and mirQTL, highlighting the critical effects of environment on both eQTL and mirQTL. Overall, these data underscore the complex genetic regulation of two well-characterized RNA classes (mRNA and miRNA) that have critical roles in the regulation of clinical traits and disease susceptibility.