PubMed

Lactate and energy metabolism during exercise in patients with blocked glycogenolysis (McArdle disease). J Clin Endocrinol Metab. 2015 Jun 1;:jc20151339 Authors: Ørngreen MC, Jeppesen TD, Taivassalo T, Hauerslev S, Preisler N, Heinicke K, Haller RG, Vissing J, van Hall G Abstract CONTEXT: Patients with blocked muscle glycogen breakdown (McArdle disease) have severely reduced exercise capacity compared to healthy individuals and suggested not to produce lactate during exercise. OBJECTIVE: The objectives were to: 1) quantifying systemic and muscle lactate kinetics and oxidation rates and muscle energy utilization during exercise in patients with McArdle disease; 2) elucidate the role of lactate formation in muscle energy production. DESIGN: Single trial Setting: Hospital Participants: Four patients with McArdle disease and seven healthy subjects. INTERVENTION: Patients and healthy controls were studied at rest, followed by 40 min of cycle-ergometer exercise at 60% of the patients maximal oxygen uptake (∼35 W). MAIN OUTCOME MEASURES: Systemic and leg skeletal muscle lactate, alanine, fatty acids and glucose kinetics. RESULTS: McArdle patients had a marked decrease in plasma lactate concentration at the onset of exercise that remained suppressed during exercise. A substantial leg net lactate uptake and subsequent oxidation occurred over the entire exercise period in contrast to a net lactate release or no exchange in the healthy controls. Despite a net lactate uptake by the active leg, a simultaneous unidirectional lactate release was observed in McArdle patients at rates, which were similar to the healthy controls. CONCLUSION: Lactate is an important energy source for contracting skeletal muscle in patients with myophosphorylase deficiency. Although McArdle patients had net leg lactate consumption, a simultaneous release of lactate was observed at rates similar to that found in healthy individuals exercising at the same very low workload, suggesting that lactate formation is mandatory for muscle energy generation during exercise. PMID: 26030324 [PubMed - as supplied by publisher]

Gut Microbiome and Obesity: A Plausible Explanation for Obesity. Curr Obes Rep. 2015 Jun;4(2):250-261 Authors: Sanmiguel C, Gupta A, Mayer EA Abstract Obesity is a multifactorial disorder that results in excessive accumulation of adipose tissue. Although obesity is caused by alterations in the energy consumption/expenditure balance, the factors promoting this disequilibrium are incompletely understood. The rapid development of new technologies and analysis strategies to decode the gut microbiota composition and metabolic pathways has opened a door into the complexity of the guest-host interactions between the gut microbiota and its human host in health and in disease. Pivotal studies have demonstrated that manipulation of the gut microbiota and its metabolic pathways can affect host's adiposity and metabolism. These observations have paved the way for further assessment of the mechanisms underlying these changes. In this review we summarize the current evidence for possible mechanisms underlying gut microbiota induced obesity. The review addresses some well-known effects of the gut microbiota on energy harvesting and changes in metabolic machinery, on metabolic and immune interactions and on possible changes in brain function and behavior. Although there is limited understanding on the symbiotic relationship between us and our gut microbiome, and how disturbances of this relationship affects our health, there is compelling evidence for an important role of the gut microbiota in the development and perpetuation of obesity. PMID: 26029487 [PubMed - as supplied by publisher]

Understanding the foundations of the structural similarities between marketed drugs and endogenous human metabolites. Front Pharmacol. 2015;6:105 Authors: O'Hagan S, Kell DB Abstract BACKGROUND: A recent comparison showed the extensive similarities between the structural properties of metabolites in the reconstructed human metabolic network ("endogenites") and those of successful, marketed drugs ("drugs"). RESULTS: Clustering indicated the related but differential population of chemical space by endogenites and drugs. Differences between the drug-endogenite similarities resulting from various encodings and judged by Tanimoto similarity could be related simply to the fraction of the bitstrings set to 1. By extracting drug/endogenite substructures, we develop a novel family of fingerprints, the Drug Endogenite Substructure (DES) encodings, based on the ranked frequency of the various substructures. These provide a natural assessment of drug-endogenite likeness, and may be used as descriptors with which to derive quantitative structure-activity relationships (QSARs). CONCLUSIONS: "Drug-endogenite likeness" seems to have utility, and leads to a simple, novel and interpretable substructure-based molecular encoding for cheminformatics. PMID: 26029108 [PubMed]

Type I interferons in anticancer immunity. Nat Rev Immunol. 2015 Jun 1; Authors: Zitvogel L, Galluzzi L, Kepp O, Smyth MJ, Kroemer G Abstract Type I interferons (IFNs) are known for their key role in antiviral immune responses. In this Review, we discuss accumulating evidence indicating that type I IFNs produced by malignant cells or tumour-infiltrating dendritic cells also control the autocrine or paracrine circuits that underlie cancer immunosurveillance. Many conventional chemotherapeutics, targeted anticancer agents, immunological adjuvants and oncolytic viruses are only fully efficient in the presence of intact type I IFN signalling. Moreover, the intratumoural expression levels of type I IFNs or of IFN-stimulated genes correlate with favourable disease outcome in several cohorts of patients with cancer. Finally, new anticancer immunotherapies are being developed that are based on recombinant type I IFNs, type I IFN-encoding vectors and type I IFN-expressing cells. PMID: 26027717 [PubMed - as supplied by publisher]

Related Articles Metabolomic analysis of extreme freezing tolerance in Siberian spruce (Picea obovata). New Phytol. 2014 Nov;204(3):545-55 Authors: Angelcheva L, Mishra Y, Antti H, Kjellsen TD, Funk C, Strimbeck RG, Schröder WP Abstract Siberian spruce (Picea obovata) is one of several boreal conifer species that can survive at extremely low temperatures (ELTs). When fully acclimated, its tissues can survive immersion in liquid nitrogen. Relatively little is known about the biochemical and biophysical strategies of ELT survival. We profiled needle metabolites using gas chromatography coupled with mass spectrometry (GC-MS) to explore the metabolic changes that occur during cold acclimation caused by natural temperature fluctuations. In total, 223 metabolites accumulated and 52 were depleted in fully acclimated needles compared with pre-acclimation needles. The metabolite profiles were found to develop in four distinct phases, which are referred to as pre-acclimation, early acclimation, late acclimation and fully acclimated. Metabolite changes associated with carbohydrate and lipid metabolism were observed, including changes associated with increased raffinose family oligosaccharide synthesis and accumulation, accumulation of sugar acids and sugar alcohols, desaturation of fatty acids, and accumulation of digalactosylglycerol. We also observed the accumulation of protein and nonprotein amino acids and polyamines that may act as compatible solutes or cryoprotectants. These results provide new insight into the mechanisms of freezing tolerance development at the metabolite level and highlight their importance in rapid acclimation to ELT in P. obovata. PMID: 25139797 [PubMed - indexed for MEDLINE]

Related Articles The origin of novel avian influenza A (H7N9) and mutation dynamics for its human-to-human transmissible capacity. PLoS One. 2014;9(3):e93094 Authors: Peng J, Yang H, Jiang H, Lin YX, Lu CD, Xu YW, Zeng J Abstract In February 2013, H7N9 (A/H7N9/2013_China), a novel avian influenza virus, broke out in eastern China and caused human death. It is a global priority to discover its origin and the point in time at which it will become transmittable between humans. We present here an interdisciplinary method to track the origin of H7N9 virus in China and to establish an evolutionary dynamics model for its human-to-human transmission via mutations. After comparing influenza viruses from China since 1983, we established an A/H7N9/2013_China virus evolutionary phylogenetic tree and found that the human instances of virus infection were of avian origin and clustered into an independent line. Comparing hemagglutinin (HA) and neuraminidase (NA) gene sequences of A/H7N9/2013_China viruses with all human-to-human, avian, and swine influenza viruses in China in the past 30 years, we found that A/H7N9/2013_China viruses originated from Baer's Pochard H7N1 virus of Hu Nan Province 2010 (HA gene, EPI: 370846, similarity with H7N9 is 95.5%) and duck influenza viruses of Nanchang city 2000 (NA gene, EPI: 387555, similarity with H7N9 is 97%) through genetic re-assortment. HA and NA gene sequence comparison indicated that A/H7N9/2013_China virus was not similar to human-to-human transmittable influenza viruses. To simulate the evolution dynamics required for human-to-human transmission mutations of H7N9 virus, we employed the Markov model. The result of this calculation indicated that the virus would acquire properties for human-to-human transmission in 11.3 years (95% confidence interval (CI): 11.2-11.3, HA gene). PMID: 24671138 [PubMed - indexed for MEDLINE]

Related Articles Evaluation of drug-induced neurotoxicity based on metabolomics, proteomics and electrical activity measurements in complementary CNS in vitro models. Toxicol In Vitro. 2015 May 27; Authors: Schultz L, Zurich MG, Culot M, Costa AF, Landry C, Bellwon P, Kristl T, Hörmann K, Ruzek S, Aiche S, Reinert K, Bielow C, Gosselet F, Cecchelli R, Huber CG, Schroeder OH, Gramowski-Voss A, Weiss DG, Bal-Price A Abstract The present study was performed in an attempt to develop an in vitro integrated testing strategy (ITS) to evaluate drug-induced neurotoxicity. A number of endpoints were analyzed using two complementary brain cell culture models and an in vitro blood-brain barrier (BBB) model after single and repeated exposure treatments with selected drugs that covered the major biological, pharmacological and neuro-toxicological responses. Furthermore, four drugs (diazepam, cyclosporine A, chlorpromazine and amiodarone) were tested more in depth as representatives of different classes of neurotoxicants, inducing toxicity through different pathways of toxicity. The developed in vitro BBB model allowed detection of toxic effects at the level of BBB and evaluation of drug transport through the barrier for predicting free brain concentrations of the studied drugs. The measurement of neuronal electrical activity was found to be a sensitive tool to predict the neuroactivity and neurotoxicity of drugs after acute exposure. The histotypic 3D re-aggregating brain cell cultures, containing all brain cell types, were found to be well suited for OMICs analyses after both acute and long term treatment. The obtained data suggest that an in vitro ITS based on the information obtained from BBB studies and combined with metabolomics, proteomics and neuronal electrical activity measurements performed in stable in vitro neuronal cell culture systems, has high potential to improve current in vitro drug-induced neurotoxicity evaluation. PMID: 26026931 [PubMed - as supplied by publisher]

Related Articles Molecular signature of amniotic fluid derived stem cells in the fetal sheep model of myelomeningocele. J Pediatr Surg. 2015 Apr 28; Authors: Ceccarelli G, Pozzo E, Scorletti F, Benedetti L, Cusella G, Ronzoni FL, Sahakyan V, Zambaiti E, Mimmi MC, Calcaterra V, Deprest J, Sampaolesi M, Pelizzo G Abstract Abnormal cord development results in spinal cord damage responsible for myelomeningocele (MMC). Amniotic fluid-derived stem cells (AFSCs) have emerged as a potential candidate for applications in regenerative medicine. However, their differentiation potential is largely unknown as well as the molecular signaling orchestrating the accurate spinal cord development. Fetal lambs underwent surgical creation of neural tube defect and its subsequent repair. AFSCs were isolated, cultured and characterized at the 12th (induction of MMC), 16th (repair of malformation), and 20th week of gestation (delivery). After performing open hysterectomy, AF collections on fetuses with sham procedures at the same time points as the MMC creation group have been used as controls. Cytological analyses with the colony forming unit assay, XTT and alkaline-phosphatase staining, qRT-PCR gene expression analyses (normalized with aged match controls) and NMR metabolomics profiling were performed. Here we show for the first time the metabolomics and molecular signature variation in AFSCs isolated in the sheep model of MMC, which may be used as diagnostic tools for the in utero identification of the neural tube damage. Intriguingly, PAX3 gene involved in the murine model for spina bifida is modulated in AFSCs reaching the peak of expression at 16weeks of gestation, 4weeks after the intervention. Our data strongly suggest that AFSCs reorganize their differentiation commitment in order to generate PAX3-expressing progenitors to counteract the MMC induced in the sheep model. The gene expression signature of AFSCs highlights the plasticity of these cells reflecting possible alterations of embryonic development. PMID: 26026346 [PubMed - as supplied by publisher]

Related Articles Targeted arginine metabolomics: A rapid, simple UPLC-QToF-MS(E) based approach for assessing the involvement of arginine metabolism in human disease. Clin Chim Acta. 2015 May 27; Authors: Van Dyk M, Mangoni AA, McEvoy M, Attia JR, Sorich MJ, Rowland A Abstract BACKGROUND: Nitric oxide synthase (NOS) mediated conversion of arginine (ARG) to citrulline (CIT) is a key pathway for nitric oxide synthesis. ARG is also metabolised by alternate pathways to ornithine (ORN), homoarginine (HMA), N(G)-monomethyl-L-arginine (MMA), N(G),N(G)-dimethyl-L-arginine (ADMA) and N(G),N(G)'-dimethyl-L-arginine (SDMA), all of which have the capacity to alter NOS activity. Simultaneous assessment of these analytes, when assessing the impact of arginine metabolism in human disease states, is desirable. METHODS: Analytes (ARG, ADMA, SDMA, MMA, HMA, CIT and ORN) were isolated from human plasma by solvent extraction, evaporated and reconstituted. Ultra-performance liquid chromatography (UPLC) was performed on a 150mm x 2.1mm T3 HSS column using a gradient mobile phase comprising ammonium formate (10mM, pH 3.8) in methanol (1% to 63%). Analytes were detected by time-of-flight mass spectrometry (Q-ToF-MS) in positive ion mode with electrospray ionization (ESI+). Data were collected using MS(E). RESULTS: Solvent extraction provided high recovery (>95%). UPLC-QToF-MS(E) facilitated the separation and quantification of the 7 analytes in an analysis time of 6min. The approach has high sensitivity; LOQ range from 0.005μM (NMMA) to 0.25μM (ARG and ORN), and good precision; intra- and inter-day %RSD <6% for all analytes. CONCLUSIONS: This approach provides the capacity to quantify 7 key compounds involved in ARG metabolism in a small sample volume, with a short total analysis time. These characteristics make this approach ideal for undertaking a comprehensive characterisation of this pathway in large data sets (e.g. population studies). PMID: 26026257 [PubMed - as supplied by publisher]

Related Articles Profiling a gut microbiota-generated catechin metabolite's fate in human blood cells using a metabolomic approach. J Pharm Biomed Anal. 2015 May 8;114:71-81 Authors: Mülek M, Fekete A, Wiest J, Holzgrabe U, Mueller MJ, Högger P Abstract The microbial catechin metabolite δ-(3,4-dihydroxy-phenyl)-γ-valerolactone (M1) has been found in human plasma samples after intake of maritime pine bark extract (Pycnogenol(®)). M1 has been previously shown to accumulate in endothelial and blood cells in vitro after facilitated uptake and to exhibit anti-inflammatory activity. The purpose of the present research approach was to systematically and comprehensively analyze the metabolism of M1 in human blood cells in vitro and in vivo. A metabolomic approach that had been successfully applied for drug metabolite profiling was chosen to detect 19 metabolite peaks of M1 which were subsequently further analyzed and validated. The metabolites were categorized into three levels of identification according to the Metabolomics Standards Initiative with six compounds each confirmed at levels 1 and 2 and seven putative metabolites at level 3. The predominant metabolites were glutathione conjugates which were rapidly formed and revealed prolonged presence within the cells. Although a formation of an intracellular conjugate of M1 and glutathione (M1-GSH) was already known two GSH conjugate isomers, M1-S-GSH and M1-N-GSH were observed in the current study. Additionally detected organosulfur metabolites were conjugates with oxidized glutathione and cysteine. Other biotransformation products constituted the open-chained ester form of M1 and a methylated M1. Six of the metabolites determined in in vitro assays were also detected in blood cells in vivo after ingestion of the pine bark extract by two volunteers. The present study provides the first evidence that multiple and structurally heterogeneous polyphenol metabolites can be generated in human blood cells. The bioactivity of the M1 metabolites and their contribution to the previously determined anti-inflammatory effects of M1 now need to be elucidated. PMID: 26025814 [PubMed - as supplied by publisher]

Related Articles Use of the Microbiome in the Practice of Epidemiology: A Primer on -Omic Technologies. Am J Epidemiol. 2015 May 29; Authors: Foxman B, Martin ET Abstract The term microbiome refers to the collective genome of the microbes living in and on our bodies, but it has colloquially come to mean the bacteria, viruses, archaea, and fungi that make up the microbiota (previously known as microflora). We can identify the microbes present in the human body (membership) and their relative abundance using genomics, characterize their genetic potential (or gene pool) using metagenomics, and describe their ongoing functions using transcriptomics, proteomics, and metabolomics. Epidemiologists can make a major contribution to this emerging field by performing well-designed, well-conducted, and appropriately powered studies and by including measures of microbiota in current and future cohort studies to characterize natural variation in microbiota composition and function, identify important confounders and effect modifiers, and generate and test hypotheses about the role of microbiota in health and disease. In this review, we provide an overview of the rapidly growing literature on the microbiome, describe which aspects of the microbiome can be measured and how, and discuss the challenges of including the microbiome as either an exposure or an outcome in epidemiologic studies. PMID: 26025238 [PubMed - as supplied by publisher]

Related Articles Untargeted metabolomics analysis revealed changes in the composition of glycerolipids and phospholipids in Bacillus subtilis under 1-butanol stress. Appl Microbiol Biotechnol. 2015 May 30; Authors: Vinayavekhin N, Mahipant G, Vangnai AS, Sangvanich P Abstract 1-Butanol has been utilized widely in industry and can be produced or transformed by microbes. However, current knowledge about the mechanisms of 1-butanol tolerance in bacteria remains quite limited. Here, we applied untargeted metabolomics to study Bacillus subtilis cells under 1-butanol stress and identified 55 and 37 ions with significantly increased and decreased levels, respectively. Using accurate mass determination, tandem mass spectra, and synthetic standards, 86 % of these ions were characterized. The levels of phosphatidylethanolamine, diglucosyldiacylglycerol, and phosphatidylserine were found to be upregulated upon 1-butanol treatment, whereas those of diacylglycerol and lysyl phosphatidylglycerol were downregulated. Most lipids contained 15:0/15:0, 16:0/15:0, and 17:0/15:0 acyl chains, and all were mapped to membrane lipid biosynthetic pathways. Subsequent two-stage quantitative real-time reverse transcriptase PCR analyses of genes in the two principal membrane lipid biosynthesis pathways revealed elevated levels of ywiE transcripts in the presence of 1-butanol and reduced expression levels of cdsA, pgsA, mprF, clsA, and yfnI transcripts. Thus, the gene transcript levels showed agreement with the metabolomics data. Lastly, the cell morphology was investigated by scanning electron microscopy, which indicated that cells became almost twofold longer after 1.4 % (v/v) 1-butanol stress for 12 h. Overall, the studies uncovered changes in the composition of glycerolipids and phospholipids in B. subtilis under 1-butanol stress, emphasizing the power of untargeted metabolomics in the discovery of new biological insights. PMID: 26025016 [PubMed - as supplied by publisher]

Related Articles Analysis of Eisenia fetida earthworm responses to sub-lethal C60 nanoparticle exposure using (1)H-NMR based metabolomics. Ecotoxicol Environ Saf. 2015 May 26;120:48-58 Authors: Lankadurai BP, Nagato EG, Simpson AJ, Simpson MJ Abstract The enhanced production and environmental release of Buckminsterfullerene (C60) nanoparticles will likely increase the exposure and risk to soil dwelling organisms. We used (1)H NMR-based metabolomics to investigate the response of Eisenia fetida earthworms to sub-lethal C60 nanoparticle exposure in both contact and soil tests. Principal component analysis of (1)H NMR data showed clear separation between controls and exposed earthworms after just 2 days of exposure, however as exposure time increased the separation decreased in soil but increased in contact tests suggesting potential adaptation during soil exposure. The amino acids leucine, valine, isoleucine and phenylalanine, the nucleoside inosine, and the sugars glucose and maltose emerged as potential bioindicators of exposure to C60 nanoparticles. The significant responses observed in earthworms using NMR-based metabolomics after exposure to very low concentrations of C60 nanoparticles suggests the need for further investigations to better understand and predict their sub-lethal toxicity. PMID: 26024814 [PubMed - as supplied by publisher]

Diverse Serum Manganese Species affect Brain Metabolites depending on Exposure Conditions. Chem Res Toxicol. 2015 May 29; Authors: Neth K, Lucio M, Walker A, Kanawati B, Zorn J, Schmitt-Kopplin P, Michalke B Abstract Occupational and environmental exposure to increased concentrations of Manganese (Mn) can lead to an accumulation of this element in the brain. The consequence is an irreversible damage of dopaminergic neurons leading to a disease called manganism with a clinical presentation similar to the one observed in Parkinson´s Disease. Human as well as animal studies indicate that Mn is mainly bound to low molecular mass (LMM) compounds such as Mn-citrate when crossing neural barriers. The shift towards LMM compounds might already take place in serum due to elevated Mn concentrations in the body.In this study we investigated Mn-species pattern in serum in two different animal models by size exclusion chromatography-inductively coupled plasma mass spectrometry (SEC-ICP-MS). A subchronic feeding of rats with elevated levels of Mn led to an increase in LMM compounds, mainly Mn-citrate and Mn bound to amino acids. In addition, a single i.v. injection of Mn showed an increase in Mn-transferrin and Mn bound to amino acids one hour after injection, while species values were rebalanced four days after the injection. Results from Mn-speciation were correlated to the brain metabolome determined by means of electrospray ionization ion cyclotron resonance Fourier transform mass spectrometry (ESI-ICR/FT-MS). The powerful combination of Mn-speciation in serum with metabolomics of the brain underlined the need for Mn-speciation in exposure scenarios instead of determination of whole Mn concentrations in blood. The progress of Mn-induced neuronal inflammation might therefore be assessed on basis of known serum Mn-species. PMID: 26024413 [PubMed - as supplied by publisher]

The Dilemma of Heterogeneity Tests in Meta-Analysis: A Challenge from a Simulation Study. PLoS One. 2015;10(5):e0127538 Authors: Li SJ, Jiang H, Yang H, Chen W, Peng J, Sun MW, Lu CD, Peng X, Zeng J Abstract INTRODUCTION: After several decades' development, meta-analysis has become the pillar of evidence-based medicine. However, heterogeneity is still the threat to the validity and quality of such studies. Currently, Q and its descendant I2 (I square) tests are widely used as the tools for heterogeneity evaluation. The core mission of this kind of test is to identify data sets from similar populations and exclude those are from different populations. Although Q and I2 are used as the default tool for heterogeneity testing, the work we present here demonstrates that the robustness of these two tools is questionable. METHODS AND FINDINGS: We simulated a strictly normalized population S. The simulation successfully represents randomized control trial data sets, which fits perfectly with the theoretical distribution (experimental group: p = 0.37, control group: p = 0.88). And we randomly generate research samples Si that fits the population with tiny distributions. In short, these data sets are perfect and can be seen as completely homogeneous data from the exactly same population. If Q and I2 are truly robust tools, the Q and I2 testing results on our simulated data sets should not be positive. We then synthesized these trials by using fixed model. Pooled results indicated that the mean difference (MD) corresponds highly with the true values, and the 95% confidence interval (CI) is narrow. But, when the number of trials and sample size of trials enrolled in the meta-analysis are substantially increased; the Q and I2 values also increase steadily. This result indicates that I2 and Q are only suitable for testing heterogeneity amongst small sample size trials, and are not adoptable when the sample sizes and the number of trials increase substantially. CONCLUSIONS: Every day, meta-analysis studies which contain flawed data analysis are emerging and passed on to clinical practitioners as "updated evidence". Using this kind of evidence that contain heterogeneous data sets leads to wrong conclusion, makes chaos in clinical practice and weakens the foundation of evidence-based medicine. We suggest more strict applications of meta-analysis: it should only be applied to those synthesized trials with small sample sizes. We call upon that the tools of evidence-based medicine should keep up-to-dated with the cutting-edge technologies in data science. Clinical research data should be made available publicly when there is any relevant article published so the research community could conduct in-depth data mining, which is a better alternative for meta-analysis in many instances. PMID: 26023932 [PubMed - as supplied by publisher]

Metabolomics Reveals that Aryl Hydrocarbon Receptor Activation by Environmental Chemicals Induces Systemic Metabolic Dysfunction in Mice. Environ Sci Technol. 2015 May 29; Authors: Zhang L, Hatzakis E, Nichols RG, Hao R, Correll JB, Smith PB, Chiaro CR, Perdew GH, Patterson AD Abstract Environmental exposure to dioxins and dioxin-like compounds poses a significant health risk for human health. Developing a better understanding of the mechanisms of toxicity through activation of the aryl hydrocarbon receptor (AHR) is likely to improve the reliability of risk assessment. In this study, the AHR-dependent metabolic responses of mice exposed to 2,3,7,8-tetrachlorodibenzofuran (TCDF) were assessed using global 1H nuclear magnetic resonance (NMR)-based metabolomics and targeted metabolic profiling of extracts obtained from serum and liver. 1H NMR analyses revealed that TCDF exposure suppressed gluconeogenesis and glycogenolysis, stimulated lipogenesis, and triggered inflammatory gene expression in an Ahr-dependent manner. Targeted analyses using gas chromatography mass spectrometry showed TCDF treatment altered the ratio of unsaturated/saturated fatty acids. Consistent with this observation, an increase in hepatic expression of stearoyl coenzyme A desaturase 1 was also observed. In addition, TCDF exposure resulted in inhibition of de novo fatty acid biosynthesis manifested by down-regulation of acetyl-CoA, malonyl-CoA and palmitoyl-CoA metabolites and related mRNA levels. In contrast, no significant changes in the levels of glucose and lipid were observed in serum and liver obtained from Ahr-null mice following TCDF treatment, thus strongly supporting the important role of the AHR in mediating the metabolic effects seen following TCDF exposure. PMID: 26023891 [PubMed - as supplied by publisher]

Quantification of folate metabolism using transient metabolic flux analysis. Cancer Metab. 2015;3:6 Authors: Tedeschi PM, Johnson-Farley N, Lin H, Shelton LM, Ooga T, Mackay G, Van Den Broek N, Bertino JR, Vazquez A Abstract BACKGROUND: Systematic quantitative methodologies are needed to understand the heterogeneity of cell metabolism across cell types in normal physiology, disease, and treatment. Metabolic flux analysis (MFA) can be used to infer steady state fluxes, but it does not apply for transient dynamics. Kinetic flux profiling (KFP) can be used in the context of transient dynamics, and it is the current gold standard. However, KFP requires measurements at several time points, limiting its use in high-throughput applications. RESULTS: Here we propose transient MFA (tMFA) as a cost-effective methodology to quantify metabolic fluxes using metabolomics and isotope tracing. tMFA exploits the time scale separation between the dynamics of different metabolites to obtain mathematical equations relating metabolic fluxes to metabolite concentrations and isotope fractions. We show that the isotope fractions of serine and glycine are at steady state 8 h after addition of a tracer, while those of purines and glutathione are following a transient dynamics with an approximately constant turnover rate per unit of metabolite, supporting the application of tMFA to the analysis of folate metabolism. Using tMFA, we investigate the heterogeneity of folate metabolism and the response to the antifolate methotrexate in breast cancer cells. Our analysis indicates that methotrexate not only inhibits purine synthesis but also induces an increase in the AMP/ATP ratio, activation of AMP kinase (AMPK), and the inhibition of protein and glutathione synthesis. We also find that in some cancer cells, the generation of one-carbon units from serine exceeds the biosynthetic demand. CONCLUSIONS: This work validates tMFA as a cost-effective methodology to investigate cell metabolism. Using tMFA, we have shown that the effects of treatment with the antifolate methotrexate extend beyond inhibition of purine synthesis and propagate to other pathways in central metabolism. PMID: 26023330 [PubMed]

Signaling through the Phosphatidylinositol 3-Kinase (PI3K)/ Mammalian Target of Rapamycin (mTOR) Axis is Responsible for Aerobic Glycolysis mediated by Glucose Transporter in Epidermal Growth Factor Receptor (EGFR)-mutated Lung Adenocarcinoma. J Biol Chem. 2015 May 28; Authors: Makinoshima H, Takita M, Saruwatari K, Umemura S, Obata Y, Ishii G, Matsumoto S, Sugiyama E, Ochiai A, Abe R, Goto K, Esumi H, Tsuchihara K Abstract Oncogenic epidermal growth factor receptor (EGFR) signaling plays an important role in regulating global metabolic pathways including aerobic glycolysis, the pentose phosphate pathway (PPP) and pyrimidine biosynthesis.　However, the molecular mechanism by which EGFR signaling regulates cancer cell metabolism is still unclear. To elucidate how EGFR signaling is linked to metabolic activity, we investigated the involvement of the RAS/MEK/ERK and PI3K/AKT/mTOR pathways on metabolic alteration in lung adenocarcinoma (LAD) cell lines with activating EGFR mutations. Although MEK inhibition did not alter lactate production and the extracellular acidification rate (ECAR), PI3K/mTOR inhibitors significantly suppressed glycolysis in EGFR-mutant LAD cells. Moreover, comprehensive metabolomics analysis revealed that the levels of glucose 6-phosphate (G6P) and 6-phosphogluconate (6PG) as early metabolites in glycolysis and PPP were decreased after inhibition of the PI3K/AKT/mTOR pathway, suggesting a linkage between PI3K signaling and the proper function of glucose transporters or hexokinases in glycolysis. Indeed, PI3K/mTOR inhibition effectively suppressed membrane localization of facilitative glucose transporter 1 (GLUT1), which instead accumulated in the cytoplasm. Finally, aerobic glycolysis and cell proliferation were down-regulated when GLUT1 gene expression　was suppressed　by RNA interference (RNAi). Taken together, these results suggest that PI3K/AKT/mTOR signaling is indispensable for the regulation of aerobic glycolysis in EGFR-mutated LAD cells. PMID: 26023239 [PubMed - as supplied by publisher]

Identification of regulatory network hubs that control lipid metabolism in Chlamydomonas reinhardtii. J Exp Bot. 2015 May 28; Authors: Gargouri M, Park JJ, Holguin FO, Kim MJ, Wang H, Deshpande RR, Shachar-Hill Y, Hicks LM, Gang DR Abstract Microalgae-based biofuels are promising sources of alternative energy, but improvements throughout the production process are required to establish them as economically feasible. One of the most influential improvements would be a significant increase in lipid yields, which could be achieved by altering the regulation of lipid biosynthesis and accumulation. Chlamydomonas reinhardtii accumulates oil (triacylglycerols, TAG) in response to nitrogen (N) deprivation. Although a few important regulatory genes have been identified that are involved in controlling this process, a global understanding of the larger regulatory network has not been developed. In order to uncover this network in this species, a combined omics (transcriptomic, proteomic and metabolomic) analysis was applied to cells grown in a time course experiment after a shift from N-replete to N-depleted conditions. Changes in transcript and protein levels of 414 predicted transcription factors (TFs) and transcriptional regulators (TRs) were monitored relative to other genes. The TF and TR genes were thus classified by two separate measures: up-regulated versus down-regulated and early response versus late response relative to two phases of polar lipid synthesis (before and after TAG biosynthesis initiation). Lipidomic and primary metabolite profiling generated compound accumulation levels that were integrated with the transcript dataset and TF profiling to produce a transcriptional regulatory network. Evaluation of this proposed regulatory network led to the identification of several regulatory hubs that control many aspects of cellular metabolism, from N assimilation and metabolism, to central metabolism, photosynthesis and lipid metabolism. PMID: 26022256 [PubMed - as supplied by publisher]

Fungal sterol C22-desaturase is not an antimycotic target as shown by selective inhibitors and testing on clinical isolates. Steroids. 2015 May 25; Authors: Müller C, Binder U, Maurer E, Grimm C, Giera M, Bracher F Abstract Inhibition of concise enzymes in ergosterol biosynthesis is one of the most prominent strategies for antifungal chemotherapy. Nevertheless, the enzymes sterol C5-desaturase and sterol C22-desaturase, which introduce double bonds into the sterol core and side chain, have not been fully investigated yet for their potential as antifungal drug targets. Lathosterol side chain amides bearing N-alkyl groups of proper length are known as potent inhibitors of the enzymes sterol C5-desaturase and sterol Δ(24)-reductase in mammalian cholesterol biosynthesis. Here we present the results of our evaluation of these amides for their ability to inhibit enzymes in fungal ergosterol biosynthesis. In the presence of inhibitor(s) an accumulation of sterols lacking a double bond at C22/23 (mainly ergosta-5,7-dien-3β-ol) was observed in Candida glabrata, Saccharomyces cerevisiae, and Yarrowia lipolytica. Hence, the lathosterol side chain amides were identified as selective inhibitors of the fungal sterol C22-desaturase, which was discussed as a specific target for novel antifungals. One representative inhibitor, (3S,20S)-20-N-butylcarbamoylpregn-7-en-3β-ol) was subjected to antifungal susceptibility testing on patient isolates according to modified EUCAST guidelines. But, the test organisms showed no significant reduction of cell growth and/or viability up to an inhibitor concentration of 100 μg/mL. This leads to the conclusion that sterol C22-desaturase is not an attractive target for the development of antifungals. PMID: 26022150 [PubMed - as supplied by publisher]