How close are we to complete annotation of metabolomes?

The metabolome describes the full complement of the tens to hundreds of thousands of low molecular weight metabolites present within a biological system. Identification of the metabolome is critical for discovering the maximum amount of biochemical knowledge from metabolomics datasets. Yet no exhaustive experimental characterisation of any organismal metabolome has been reported to date, dramatically contrasting with the genome sequencing of thousands of plants, animals and microbes. Here we review the status of metabolome annotation and describe advances in the analytical methodologies being applied. In part through new international coordination, we conclude that we are now entering a new era of metabolome annotation

Application of combined omics platforms to accelerate biomedical discovery in diabesity

Diabesity has become a popular term to describe the specific form of diabetes that develops late in life and is associated with obesity. While there is a correlation between diabetes and obesity, the association is not universally predictive. Defining the metabolic characteristics of obesity that lead to diabetes, and how obese individuals who develop diabetes different from those who do not, are important goals. The use of large-scale omics analyses (e.g., metabolomic, proteomic, transcriptomic, and lipidomic) of diabetes and obesity may help to identify new targets to treat these conditions. This report discusses how various types of omics data can be integrated to shed light on the changes in metabolism that occur in obesity and diabetes

Advantages of dynamic “closed loop” stable isotope flux phenotyping over static “open loop” clamps in detecting silent genetic and dietary phenotypes

In vivo insulin sensitivity can be assessed using “open loop” clamp or “closed loop” methods. Open loop clamp methods are static, and fix plasma glucose independently from plasma insulin. Closed loop methods are dynamic, and assess glucose disposal in response to a stable isotope labeled glucose tolerance test. Using PPARα−/− mice, open and closed loop assessments of insulin sensitivity/glucose disposal were compared. Indirect calorimetry done for the assessment of diurnal substrate utilization/metabolic flexibility showed that chow fed PPARα−/− mice had increased glucose utilization during the light (starved) cycle. Euglycemic clamps showed no differences in insulin stimulated glucose disposal, whether for chow or high fat diets, but did show differences in basal glucose clearance for chow fed PPARα−/− versus SV129J-wt mice. In contrast, the dynamic stable isotope labeled glucose tolerance tests reveal enhanced glucose disposal for PPARα−/− versus SV129J-wt, for chow and high fat diets. Area under the curve for plasma labeled and unlabeled glucose for PPARα−/− was ≈1.7-fold lower, P < 0.01 during the stable isotope labeled glucose tolerance test for both diets. Area under the curve for plasma insulin was 5-fold less for the chow fed SV129J-wt (P < 0.01) but showed no difference on a high fat diet (0.30 ± 0.1 for SV129J-wt vs. 0.13 ± 0.10 for PPARα−/−, P = 0.28). This study demonstrates that dynamic stable isotope labeled glucose tolerance test can assess “silent” metabolic phenotypes, not detectable by the static, “open loop”, euglycemic or hyperglycemic clamps. Both open loop and closed loop methods may describe different aspects of metabolic inflexibility and insulin sensitivity

Genetically Increasing Flux Through b-Oxidation in Skeletal Muscle Increases Mitochondrial Reductive Stress and Glucose Intolerance

Elevated mitochondrial hydrogen peroxide (H(2)O(2)) emission and an oxidative shift in cytosolic redox environment have been linked to high-fat-diet-induced insulin resistance in skeletal muscle. To test specifically whether increased flux through mitochondrial fatty acid oxidation, in the absence of elevated energy demand, directly alters mitochondrial function and redox state in muscle, two genetic models characterized by increased muscle β-oxidation flux were studied. In mice overexpressing peroxisome proliferator-activated receptor-α in muscle (MCK-PPARα), lipid-supported mitochondrial respiration, membrane potential (ΔΨ(m)), and H(2)O(2) production rate (JH(2)O(2)) were increased, which coincided with a more oxidized cytosolic redox environment, reduced muscle glucose uptake, and whole body glucose intolerance despite an increased rate of energy expenditure. Similar results were observed in lipin-1-deficient, fatty-liver dystrophic mice, another model characterized by increased β-oxidation flux and glucose intolerance. Crossing MCAT (mitochondria-targeted catalase) with MCK-PPARα mice normalized JH(2)O(2) production, redox environment, and glucose tolerance, but surprisingly, both basal and absolute insulin-stimulated rates of glucose uptake in muscle remained depressed. Also surprising, when placed on a high-fat diet, MCK-PPARα mice were characterized by much lower whole body, fat, and lean mass as well as improved glucose tolerance relative to wild-type mice, providing additional evidence that overexpression of PPARα in muscle imposes more extensive metabolic stress than experienced by wild-type mice on a high-fat diet. Overall, the findings suggest that driving an increase in skeletal muscle fatty acid oxidation in the absence of metabolic demand imposes mitochondrial reductive stress and elicits multiple counterbalance metabolic responses in an attempt to restore bioenergetic homeostasis. NEW & NOTEWORTHY Prior work has suggested that mitochondrial dysfunction is an underlying cause of insulin resistance in muscle because it limits fatty acid oxidation and therefore leads to the accumulation of cytotoxic lipid intermediates. The implication has been that therapeutic strategies to accelerate β-oxidation will be protective. The current study provides evidence that genetically increasing flux through β-oxidation in muscle imposes reductive stress that is not beneficial but rather detrimental to metabolic regulation

Lactate efflux from intervertebral disc cells is required for maintenance of spine health.

Maintenance of glycolytic metabolism is postulated to be required for health of the spinal column. In the hypoxic tissues of the intervertebral disc and glycolytic cells of vertebral bone, glucose is metabolized into pyruvate for ATP generation and reduced to lactate to sustain redox balance. The rise in intracellular H+ /lactate concentrations are balanced by plasma-membrane monocarboxylate transporters (MCTs). Using MCT4 null mice and human tissue samples, complimented with genetic and metabolic approaches, we determine that H+ /lactate efflux is critical for maintenance of disc and vertebral bone health. Mechanistically, MCT4 maintains glycolytic and TCA cycle flux and intracellular pH homeostasis in the nucleus pulposus compartment of the disc, where HIF-1α directly activates an intronic enhancer in SLC16A3. Ultimately, our results provide support for research into lactate as a diagnostic biomarker for chronic, painful disc degeneration

Osteocalcin signaling in myofibers is necessary and sufficient for optimum adaptation to exercise

Circulating levels of undercarboxylated and bioactive osteocalcin double during aerobic exercise at the time levels of insulin decrease. In contrast, circulating levels of osteocalcin plummet early during adulthood in mice, monkeys, and humans of both genders. Exploring these observations revealed that osteocalcin signaling in myofibers is necessary for adaptation to exercise by favoring uptake and catabolism of glucose and fatty acids, the main nutrients of myofibers. Osteocalcin signaling in myofibers also accounts for most of the exercise-induced release of interleukin-6, a myokine that promotes adaptation to exercise in part by driving the generation of bioactive osteocalcin. We further show that exogenous osteocalcin is sufficient to enhance the exercise capacity of young mice and to restore to 15-month-old mice the exercise capacity of 3-month-old mice. This study uncovers a bone-to-muscle feedforward endocrine axis that favors adaptation to exercise and can reverse the age-induced decline in exercise capacity

Sirtuin 6 inhibition protects against glucocorticoid-induced skeletal muscle atrophy by regulating IGF/PI3K/AKT signaling

Chronic activation of stress hormones such as glucocorticoids leads to skeletal muscle wasting in mammals. However, the molecular events that mediate glucocorticoid-induced muscle wasting are not well understood. Here, we show that SIRT6, a chromatin-associated deacetylase indirectly regulates glucocorticoid-induced muscle wasting by modulating IGF/PI3K/AKT signaling. Our results show that SIRT6 levels are increased during glucocorticoid-induced reduction of myotube size and during skeletal muscle atrophy in mice. Notably, overexpression of SIRT6 spontaneously decreases the size of primary myotubes in a cell-autonomous manner. On the other hand, SIRT6 depletion increases the diameter of myotubes and protects them against glucocorticoid-induced reduction in myotube size, which is associated with enhanced protein synthesis and repression of atrogenes. In line with this, we find that muscle-specific SIRT6 deficient mice are resistant to glucocorticoid-induced muscle wasting. Mechanistically, we find that SIRT6 deficiency hyperactivates IGF/PI3K/AKT signaling through c-Jun transcription factor-mediated increase in IGF2 expression. The increased activation, in turn, leads to nuclear exclusion and transcriptional repression of the FoxO transcription factor, a key activator of muscle atrophy. Further, we find that pharmacological inhibition of SIRT6 protects against glucocorticoid-induced muscle wasting in mice by regulating IGF/PI3K/AKT signaling implicating the role of SIRT6 in glucocorticoid-induced muscle atrophy.Fil: Mishra, Sneha. No especifíca;Fil: Cosentino, Claudia. Harvard Medical School; Estados UnidosFil: Tamta, Ankit Kumar. No especifíca;Fil: Khan, Danish. No especifíca;Fil: Srinivasan, Shalini. No especifíca;Fil: Ravi, Venkatraman. No especifíca;Fil: Abbotto, Elena. Università degli Studi di Genova; ItaliaFil: Arathi, Bangalore Prabhashankar. No especifíca;Fil: Kumar, Shweta. No especifíca;Fil: Jain, Aditi. No especifíca;Fil: Ramaian, Anand S.. No especifíca;Fil: Kizkekra, Shruti M.. No especifíca;Fil: Rajagopal, Raksha. No especifíca;Fil: Rao, Swathi. No especifíca;Fil: Krishna, Swati. No especifíca;Fil: Asirvatham Jeyaraj, Ninitha. Indian Institute of Technology; IndiaFil: Haggerty, Elizabeth R.. Harvard Medical School; Estados UnidosFil: Silberman, Dafne Magalí. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; ArgentinaFil: Kurland, Irwin J.. No especifíca;Fil: Veeranna, Ravindra P.. No especifíca;Fil: Jayavelu, Tamilselvan. No especifíca;Fil: Bruzzone, Santina. Università degli Studi di Genova; ItaliaFil: Mostoslavsky, Raul. Harvard Medical School; Estados UnidosFil: Sundaresan, Nagalingam R.. No especifíca

Comparison of Fecal Collection Methods for Microbiome and Metabolomics Studies

Background: Integrated microbiome and metabolomics analyses hold the potential to reveal interactions between host and microbiota in relation to disease risks. However, there are few studies evaluating how field methods influence fecal microbiome characterization and metabolomics profiling.Methods: Five fecal collection methods [immediate freezing at −20°C without preservative, OMNIgene GUT, 95% ethanol, RNAlater, and Flinders Technology Associates (FTA) cards] were used to collect 40 fecal samples from eight healthy volunteers. We performed gut microbiota 16S rRNA sequencing, untargeted metabolomics profiling, and targeted metabolomics focusing on short chained fatty acids (SCFAs). Metrics included α-diversity and β-diversity as well as distributions of predominant phyla. To evaluate the concordance with the “gold standard” immediate freezing, the intraclass correlation coefficients (ICCs) for alternate fecal collection systems were calculated. Correlations between SCFAs and gut microbiota were also examined.Results: The FTA cards had the highest ICCs compared to the immediate freezing method for α-diversity indices (ICCs = 0.96, 0.96, 0.76 for Shannon index, Simpson's Index, Chao-1 Index, respectively), followed by OMNIgene GUT, RNAlater, and 95% ethanol. High ICCs (all &gt;0.88) were observed for all methods for the β-diversity metric. For untargeted metabolomics, in comparison to immediate freezing which detected 621 metabolites at ≥75% detectability level, 95% ethanol showed the largest overlapping set of metabolites (n = 430; 69.2%), followed by FTA cards (n = 330; 53.1%) and OMNIgene GUT (n = 213; 34.3%). Both OMNIgene GUT (ICCs = 0.82, 0.93, 0.64) and FTA cards (ICCs = 0.87, 0.85, 0.54) had acceptable ICCs for the top three predominant SCFAs (butyric acid, propionic acid and acetic acid). Nominally significant correlations between bacterial genera and SCFAs (P &lt; 0.05) were observed in fecal samples collected by different methods. Of note, a high correlation between the genus Blautia (known butyrate producer) and butyric acid was observed for both immediate freezing (r = 0.83) and FTA cards (r = 0.74).Conclusions: Four alternative fecal collection methods are generally comparable with immediate freezing, but there are differences in certain measures of the gut microbiome and fecal metabolome across methods. Choice of method depends on the research interests, simplicity of fecal collection procedures and ease of transportation to the lab, especially for large epidemiological studies