Myelopoiesis is necessary for the generation of mature myeloid cells during homeostatic turnover and immunological insults; however, the metabolic requirements for this process remain poorly defined. ...Here, we demonstrate that myelopoiesis, including monocyte and macrophage differentiation, requires mechanistic target of rapamycin complex 1 (mTORC1) signaling and anabolic metabolism. Loss of mTORC1 impaired myelopoiesis under steady state and dampened innate immune responses against
infection. Stimulation of hematopoietic progenitors with macrophage colony-stimulating factor (M-CSF) resulted in mTORC1-dependent anabolic metabolism, which in turn promoted expression of M-CSF receptor and transcription factors PU.1 and IRF8, thereby constituting a feed-forward loop for myelopoiesis. Mechanistically, mTORC1 engaged glucose metabolism and initiated a transcriptional program involving Myc activation and sterol biosynthesis after M-CSF stimulation. Perturbation of glucose metabolism or disruption of Myc function or sterol biosynthesis impaired myeloid differentiation. Integrative metabolomic and genomic profiling further identified one-carbon metabolism as a central node in mTORC1-dependent myelopoiesis. Therefore, the interplay between mTORC1 signaling and metabolic reprogramming underlies M-CSF-induced myelopoiesis.
Abstract Background Pulmonary hypertension and associated right ventricular (RV) dysfunction are important determinants of morbidity and mortality, which are optimally characterized by invasive ...hemodynamic measurements. Objectives This study sought to determine whether metabolite profiling could identify plasma signatures of right ventricular-pulmonary vascular (RV-PV) dysfunction. Methods We measured plasma concentrations of 105 metabolites using targeted mass spectrometry in 71 individuals (discovery cohort) who underwent comprehensive physiological assessment with right-sided heart catheterization and radionuclide ventriculography at rest and during exercise. Our findings were validated in a second cohort undergoing invasive hemodynamic evaluations (n = 71), as well as in an independent cohort with or without known pulmonary arterial (PA) hypertension (n = 30). Results In the discovery cohort, 21 metabolites were associated with 2 or more hemodynamic indicators of RV-PV function (i.e., resting right atrial pressure, mean PA pressure, pulmonary vascular resistance PVR, and PVR and PA pressure-flow response ΔPQ during exercise). We identified novel associations of RV-PV dysfunction with circulating indoleamine 2,3-dioxygenase (IDO)–dependent tryptophan metabolites (TMs), tricarboxylic acid intermediates, and purine metabolites and confirmed previously described associations with arginine–nitric oxide metabolic pathway constituents. IDO-TM levels were inversely related to RV ejection fraction and were particularly well correlated with exercise PVR and ΔPQ. Multisite sampling demonstrated transpulmonary release of IDO-TMs. IDO-TMs also identified RV-PV dysfunction in a validation cohort with known risk factors for pulmonary hypertension and in patients with established PA hypertension. Conclusions Metabolic profiling identified reproducible signatures of RV-PV dysfunction, highlighting both new biomarkers and pathways for further functional characterization.
The role of branched-chain amino acids (BCAAs) in cardiovascular disease (CVD) remains poorly understood. We hypothesized that baseline BCAA concentrations predict future risk of CVD and that a ...Mediterranean diet (MedDiet) intervention may counteract this effect.
We developed a case-cohort study within the Prevención con Dieta Mediterránea (PREDIMED), with 226 incident CVD cases and 744 noncases. We used LC-MS/MS to measure plasma BCAAs (leucine, isoleucine, and valine), both at baseline and after 1 year of follow-up. The primary outcome was a composite of incident stroke, myocardial infarction, or cardiovascular death.
After adjustment for potential confounders, baseline leucine and isoleucine concentrations were associated with higher CVD risk: the hazard ratios (HRs) for the highest vs lowest quartile were 1.70 (95% CI, 1.05-2.76) and 2.09 (1.27-3.44), respectively. Stronger associations were found for stroke. For both CVD and stroke, we found higher HRs across successive quartiles of BCAAs in the control group than in the MedDiet groups. With stroke as the outcome, a significant interaction (P = 0.009) between baseline BCAA score and intervention with MedDiet was observed. No significant effect of the intervention on 1-year changes in BCAAs or any association between 1-year changes in BCAAs and CVD were observed.
Higher concentrations of baseline BCAAs were associated with increased risk of CVD, especially stroke, in a high cardiovascular risk population. A Mediterranean-style diet had a negligible effect on 1-year changes in BCAAs, but it may counteract the harmful effects of BCAAs on stroke.
The inflammatory bowel diseases (IBDs), which include Crohn's disease (CD) and ulcerative colitis (UC), are multifactorial chronic conditions of the gastrointestinal tract. While IBD has been ...associated with dramatic changes in the gut microbiota, changes in the gut metabolome-the molecular interface between host and microbiota-are less well understood. To address this gap, we performed untargeted metabolomic and shotgun metagenomic profiling of cross-sectional stool samples from discovery (n = 155) and validation (n = 65) cohorts of CD, UC and non-IBD control patients. Metabolomic and metagenomic profiles were broadly correlated with faecal calprotectin levels (a measure of gut inflammation). Across >8,000 measured metabolite features, we identified chemicals and chemical classes that were differentially abundant in IBD, including enrichments for sphingolipids and bile acids, and depletions for triacylglycerols and tetrapyrroles. While > 50% of differentially abundant metabolite features were uncharacterized, many could be assigned putative roles through metabolomic 'guilt by association' (covariation with known metabolites). Differentially abundant species and functions from the metagenomic profiles reflected adaptation to oxidative stress in the IBD gut, and were individually consistent with previous findings. Integrating these data, however, we identified 122 robust associations between differentially abundant species and well-characterized differentially abundant metabolites, indicating possible mechanistic relationships that are perturbed in IBD. Finally, we found that metabolome- and metagenome-based classifiers of IBD status were highly accurate and, like the vast majority of individual trends, generalized well to the independent validation cohort. Our findings thus provide an improved understanding of perturbations of the microbiome-metabolome interface in IBD, including identification of many potential diagnostic and therapeutic targets.
Rigorous studies are necessary to demonstrate suitability of metabolomics platforms to profile metabolites in archived plasma within epidemiologic studies of human disease, for which attenuation of ...effect estimates due to measurement error is a key concern.
Using a liquid chromatography-tandem mass spectrometry platform, we quantified 257 metabolites from archived plasma to evaluate metabolite interassay reproducibility, reproducibility with delayed processing, and within-person reproducibility over time. Interassay reproducibility was assessed with CVs from 60 duplicate plasma samples donated by participants in the Nurses' Health Study and Health Professionals Follow-up Study, and 20 QC pool plasma replicates. Metabolite reproducibility over a 24- to 48-h processing delay (n = 48 samples) and within-person reproducibility over 1-2 years (n = 80 samples) were assessed using Spearman and intraclass correlation coefficients (ICCs).
CVs were <20% for 92% of metabolites and generally were similar by plasma anticoagulant type (heparin or EDTA) and fasting time. Approximately 75% of metabolites were reproducible over delays in processing of blood samples (Spearman correlation or ICC ≥ 0.75, comparing immediate and 24-h delayed processing). Carbohydrates and purine/pyrimidine derivatives were most adversely affected by the processing delay. Ninety percent of metabolites were reproducible over 1-2 years within individuals (Spearman correlation or ICC ≥ 0.4).
For potential use in epidemiologic studies, the majority of plasma metabolites had low CVs and were reproducible over a 24-h processing delay and within individuals over 1-2 years. Certain metabolites, such as carbohydrates and purine/pyrimidine derivatives, may be challenging to evaluate if samples have delayed processing.
Aberrant activation of the mammalian target of rapamycin complex 1 (mTORC1) is a common molecular event in a variety of pathological settings, including genetic tumor syndromes, cancer, and obesity. ...However, the cell-intrinsic consequences of mTORC1 activation remain poorly defined. Through a combination of unbiased genomic, metabolomic, and bioinformatic approaches, we demonstrate that mTORC1 activation is sufficient to stimulate specific metabolic pathways, including glycolysis, the oxidative arm of the pentose phosphate pathway, and de novo lipid biosynthesis. This is achieved through the activation of a transcriptional program affecting metabolic gene targets of hypoxia-inducible factor (HIF1α) and sterol regulatory element-binding protein (SREBP1 and SREBP2). We find that SREBP1 and 2 promote proliferation downstream of mTORC1, and the activation of these transcription factors is mediated by S6K1. Therefore, in addition to promoting protein synthesis, mTORC1 activates specific bioenergetic and anabolic cellular processes that are likely to contribute to human physiology and disease.
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► Unbiased genomic and metabolomic approaches to reveal downstream functions of mTORC1 ► mTORC1 stimulates glucose uptake and glycolysis through HIF1 ► mTORC1 stimulates the pentose phosphate pathway and lipid biosynthesis through SREBP1 ► S6K1 regulates SREBP1 processing and transcriptional activation of target genes
Inflammatory bowel diseases, which include Crohn's disease and ulcerative colitis, affect several million individuals worldwide. Crohn's disease and ulcerative colitis are complex diseases that are ...heterogeneous at the clinical, immunological, molecular, genetic, and microbial levels. Individual contributing factors have been the focus of extensive research. As part of the Integrative Human Microbiome Project (HMP2 or iHMP), we followed 132 subjects for one year each to generate integrated longitudinal molecular profiles of host and microbial activity during disease (up to 24 time points each; in total 2,965 stool, biopsy, and blood specimens). Here we present the results, which provide a comprehensive view of functional dysbiosis in the gut microbiome during inflammatory bowel disease activity. We demonstrate a characteristic increase in facultative anaerobes at the expense of obligate anaerobes, as well as molecular disruptions in microbial transcription (for example, among clostridia), metabolite pools (acylcarnitines, bile acids, and short-chain fatty acids), and levels of antibodies in host serum. Periods of disease activity were also marked by increases in temporal variability, with characteristic taxonomic, functional, and biochemical shifts. Finally, integrative analysis identified microbial, biochemical, and host factors central to this dysregulation. The study's infrastructure resources, results, and data, which are available through the Inflammatory Bowel Disease Multi'omics Database ( http://ibdmdb.org ), provide the most comprehensive description to date of host and microbial activities in inflammatory bowel diseases.
Alternative splicing of the Pkm gene product generates the PKM1 and PKM2 isoforms of pyruvate kinase (PK), and PKM2 expression is closely linked to embryogenesis, tissue regeneration, and cancer. To ...interrogate the functional requirement for PKM2 during development and tissue homeostasis, we generated germline PKM2-null mice (Pkm2(-/-)). Unexpectedly, despite being the primary isoform expressed in most wild-type adult tissues, we found that Pkm2(-/-) mice are viable and fertile. Thus, PKM2 is not required for embryonic or postnatal development. Loss of PKM2 leads to compensatory expression of PKM1 in the tissues that normally express PKM2. Strikingly, PKM2 loss leads to spontaneous development of hepatocellular carcinoma (HCC) with high penetrance that is accompanied by progressive changes in systemic metabolism characterized by altered systemic glucose homeostasis, inflammation, and hepatic steatosis. Therefore, in addition to its role in cancer metabolism, PKM2 plays a role in controlling systemic metabolic homeostasis and inflammation, thereby preventing HCC by a non-cell-autonomous mechanism.
Whereas regular exercise is associated with lower risk of cardiovascular disease and mortality, mechanisms of exercise-mediated health benefits remain less clear. We used metabolite profiling before ...and after acute exercise to delineate the metabolic architecture of exercise response patterns in humans.
Cardiopulmonary exercise testing and metabolite profiling was performed on Framingham Heart Study participants (age 53±8 years, 63% women) with blood drawn at rest (n=471) and at peak exercise (n=411).
We observed changes in circulating levels for 502 of 588 measured metabolites from rest to peak exercise (exercise duration 11.9±2.1 minutes) at a 5% false discovery rate. Changes included reductions in metabolites implicated in insulin resistance (glutamate, -29%;
=1.5×10
; dimethylguanidino valeric acid DMGV, -18%;
=5.8×10
) and increases in metabolites associated with lipolysis (1-methylnicotinamide, +33%;
=6.1×10
), nitric oxide bioavailability (arginine/ornithine + citrulline, +29%;
=2.8×10
), and adipose browning (12,13-dihydroxy-9Z-octadecenoic acid +26%;
=7.4×10
), among other pathways relevant to cardiometabolic risk. We assayed 177 metabolites in a separate Framingham Heart Study replication sample (n=783, age 54±8 years, 51% women) and observed concordant changes in 164 metabolites (92.6%) at 5% false discovery rate. Exercise-induced metabolite changes were variably related to the amount of exercise performed (peak workload), sex, and body mass index. There was attenuation of favorable excursions in some metabolites in individuals with higher body mass index and greater excursions in select cardioprotective metabolites in women despite less exercise performed. Distinct preexercise metabolite levels were associated with different physiologic dimensions of fitness (eg, ventilatory efficiency, exercise blood pressure, peak Vo
). We identified 4 metabolite signatures of exercise response patterns that were then analyzed in a separate cohort (Framingham Offspring Study; n=2045, age 55±10 years, 51% women), 2 of which were associated with overall mortality over median follow-up of 23.1 years (
≤0.003 for both).
In a large sample of community-dwelling individuals, acute exercise elicits widespread changes in the circulating metabolome. Metabolic changes identify pathways central to cardiometabolic health, cardiovascular disease, and long-term outcome. These findings provide a detailed map of the metabolic response to acute exercise in humans and identify potential mechanisms responsible for the beneficial cardiometabolic effects of exercise for future study.
Gut dysbiosis during inflammatory bowel disease involves alterations in the gut microbiota associated with inflammation of the host gut. We used a combination of shotgun metagenomic sequencing and ...metabolomics to analyze fecal samples from pediatric patients with Crohn's disease and found an association between disease severity, gut dysbiosis, and bacterial production of free amino acids. Nitrogen flux studies using
N in mice showed that activity of bacterial urease, an enzyme that releases ammonia by hydrolysis of host urea, led to the transfer of murine host-derived nitrogen to the gut microbiota where it was used for amino acid synthesis. Inoculation of a conventional murine host (pretreated with antibiotics and polyethylene glycol) with commensal
engineered to express urease led to dysbiosis of the gut microbiota, resulting in a predominance of Proteobacteria species. This was associated with a worsening of immune-mediated colitis in these animals. A potential role for altered urease expression and nitrogen flux in the development of gut dysbiosis suggests that bacterial urease may be a potential therapeutic target for inflammatory bowel diseases.
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