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Background: The rupture of atherosclerotic plaque contributes significantly to cardiovascular disease. Plasma concentrations of bilirubin—a byproduct of heme catabolism—inversely associate with risk of cardiovascular disease, although the link between bilirubin and atherosclerosis remains unclear. Methods: To assess the role of bilirubin in atherosclerotic plaque stability, we crossed Bvra −/− with Apoe −/− mice and used the tandem stenosis model of plaque instability. Human coronary arteries were obtained from heart transplant recipients. Analysis of bile pigments, heme metabolism, and proteomics were performed by liquid chromatography tandem mass spectrometry. MPO (myeloperoxidase) activity was determined by in vivo molecular magnetic resonance imaging, liquid chromatography tandem mass spectrometry analysis, and immunohistochemical determination of chlorotyrosine. Systemic oxidative stress was evaluated by plasma concentrations of lipid hydroperoxides and the redox status of circulating Prx2 (peroxiredoxin 2), whereas arterial function was assessed by wire myography. Atherosclerosis and arterial remodeling were quantified by morphometry and plaque stability by fibrous cap thickness, lipid accumulation, infiltration of inflammatory cells, and the presence of intraplaque hemorrhage. Results: Compared with Bvra +/+ Apoe −/− tandem stenosis littermates, Bvra −/− Apoe −/− tandem stenosis mice were deficient in bilirubin, showed signs of increased systemic oxidative stress, endothelial dysfunction, as well as hyperlipidemia, and had a higher atherosclerotic plaque burden. Heme metabolism was increased in unstable compared with stable plaque of both Bvra +/+ Apoe −/− and Bvra −/− Apoe −/− tandem stenosis mice and in human coronary plaques. In mice, Bvra deletion selectively destabilized unstable plaque, characterized by positive arterial remodeling and increased cap thinning, intraplaque hemorrhage, infiltration of neutrophils, and MPO activity. Proteomic analysis confirmed Bvra deletion enhanced extracellular matrix degradation, recruitment and activation of neutrophils, and associated oxidative stress in unstable plaque. Conclusions: Bilirubin deficiency, resulting from global Bvra deletion, generates a proatherogenic phenotype and selectively enhances neutrophil-mediated inflammation and destabilization of unstable plaque, thereby providing a link between bilirubin and cardiovascular disease risk.

Conversion of the redox probe hydroethidine (HE) to 2-chloroethidium (2-Cl-E

Near-infrared autofluorescence (NIRAF) in unstable atherosclerotic plaque has been suggested as a novel imaging technology for high-risk atherosclerosis. Intraplaque hemorrhage (IPH) and bilirubin, derived from the subsequent degradation of heme, have been proposed as the source of NIRAF, although their roles and the underlying mechanism responsible for NIRAF remain unclear. To test the proposed role of bilirubin as the source of NIRAF in high-risk atherosclerosis, Biliverdin reductase a gene and apolipoprotein E gene double-knockout (Bvra−/−Apoe−/−) mice were subjected to the Western diet and tandem stenosis (TS) surgery, as a model of both bilirubin deficiency and plaque instability. Human coronary arteries containing atherosclerotic plaques were obtained from heart transplant recipients. The NIRAF was determined by in vivo fluorescence emission computed tomography, and ex vivo infrared imaging. The cholesterol content was quantified by HPLC with UV detection. In Bvra+/+Apoe−/− TS mice, the NIRAF intensity was significantly higher in unstable plaque than in stable plaque, yet the NIRAF in unstable plaque was undistinguishable in Bvra+/+Apoe−/− and littermate Bvra−/−Apoe−/− TS mice. Moreover, the unstable plaque in TS mice exhibited a lower NIRAF compared with highly cellular plaque that lacked most of the features of unstable plaque. In human coronary arteries, the NIRAF associated with cholesterol-rich, calcified lesions, rather than just cholesterol-rich lesions. The NIRAF in atherosclerotic plaque can be dissociated from IPH and bilirubin, such that the compositional meaning of an elevated NIRAF remains obscure.

The completeness of the system of eigenfunctions of the complex Schrodinger operator L c = − d 2 / d x 2 + c x 2 / 3 on the semi-axis in L 2 ( R + ) with Dirichlet boundary conditions is proved for all c: | arg ⁡ c | π / 2 + θ 0 , where π / 10 θ 0 π / 2 is determined as the only solution of a certain transcendental equation.

Supplementary Methods

Figure S1-S5 with legends

Table S1

Figure S1 shows the effect of statins on cholesterol and coenzyme Q levels and synthesis. Figure S2 shows statins reduce oxidative phosphorylation and hence increase glucose uptake and lactate secretion. Figure S3 shows the metabolic rewiring that occurs as a consequence of statin-mediated ROS production. Figure S4 shows simvastatin treatment in vivo does not alter cholesterol or dolichol synthesis or systemic coenzyme Q synthesis. Figure S5 shows simvastatin and AZD6244 are synergistic.

Statins are widely prescribed inhibitors of the mevalonate pathway, acting to lower systemic cholesterol levels. The mevalonate pathway is critical for tumorigenesis and is frequently upregulated in cancer. Nonetheless, reported effects of statins on tumor progression are ambiguous, making it unclear whether statins, alone or in combination, can be used for chemotherapy. Here, using advanced mass spectrometry and isotope tracing, we showed that statins only modestly affected cancer cholesterol homeostasis. Instead, they significantly reduced synthesis and levels of another downstream product, the mitochondrial electron carrier coenzyme Q, both in cultured cancer cells and tumors. This compromised oxidative phosphorylation, causing severe oxidative stress. To compensate, cancer cells upregulated antioxidant metabolic pathways, including reductive carboxylation, proline synthesis, and cystine import. Targeting cystine import with an xCT transporter–lowering MEK inhibitor, in combination with statins, caused profound tumor cell death. Thus, statin-induced ROS production in cancer cells can be exploited in a combinatorial regimen.Significance:Cancer cells induce specific metabolic pathways to alleviate the increased oxidative stress caused by statin treatment, and targeting one of these pathways synergizes with statins to produce a robust antitumor response.See related commentary by Cordes and Metallo, p. 151

AGPATs (1-acylglycerol-3-phosphate O-acyltransferases) catalyze the acylation of lysophosphatidic acid to form phosphatidic acid (PA), a key step in the glycerol-3-phosphate pathway for the synthesis of phospholipids and triacylglycerols. AGPAT2 is the only AGPAT isoform whose loss-of-function mutations cause a severe form of human congenital generalized lipodystrophy. Paradoxically, AGPAT2 deficiency is known to dramatically increase the level of its product, PA. Here, we find that AGPAT2 deficiency impairs the biogenesis and growth of lipid droplets. We show that AGPAT2 deficiency compromises the stability of CDP-diacylglycerol (DAG) synthases (CDSs) and decreases CDS activity in both cell lines and mouse liver. Moreover, AGPAT2 and CDS1/2 can directly interact and form functional complexes, which promote the metabolism of PA along the CDP-DAG pathway of phospholipid synthesis. Our results provide key insights into the regulation of metabolic flux during lipid synthesis and suggest substrate channelling at a major branch point of the glycerol-3-phosphate pathway., AGPATs (1-acylglycerol-3-phosphate O-acyltransferases) catalyze the acylation of lysophosphatidic acid to form phosphatidic acid (PA), a key step in the synthesis of all glycerolipids. Here, the authors show that AGPAT2 and CDP-DAG synthases (CDS1 and CDS2) form functional complexes that promote further conversion of PA along the CDP-DAG pathway of phospholipid synthesis.

The impact of the Covid-19 pandemic on the national development processes is under study. The article considers methods for assessing changes that affect the territory development prepared by Russian and Polish researchers. A comparative characteristic of methodological approaches is given. The discussion made it possible to establish that the result of the development of any territory, of course, should be considered in terms of changes in the quality of life of the population. In this regard, the methodology developed by Russian authors, which provides for the gradation of influencing factors by groups of influence, was taken as the basis. In total, four special interest groups on changes in the quality of life of the population were identified, such as factors associated with the population itself; business structure development factors; infrastructural factors, which are the responsibility of the municipalities; and institutional factors providing for the activities of public institutions. The study presented in the article is based on the sociological survey data among young population conducted on the basis of several universities in the two countries of Russia and Poland in the pre-pandemic and pandemic periods, since the youth student environment determines the potential for future development of territories. The main changes (positive and negative) in the assessments of the impact of various factors on the quality of life of the population caused by the Covid-19 pandemic have been established.

The article raises the issues of the ultimate effectiveness of regulatory measures of a social nature carried out by the state in rural areas of Russia and Belarus. For this purpose, the results of individual government directives are compared with the dynamics of demographic indicators, employment, real incomes, housing provision in rural areas, the structure of household expenditures. The problem areas of social policy in rural areas of Russia and Belarus are considered. As creative views on the solution of social problems in rural areas, options for the introduction and development of innovative agro-industrial ecosystems, which are defined by the authors as a set of economic, social, energy, information, marketing, financial and other entrepreneurial business structures that cover all the problem areas of development, are proposed.

Mitochondrial energy production and function rely on optimal concentrations of the essential redox-active lipid, coenzyme Q (CoQ). CoQ deficiency results in mitochondrial dysfunction associated with increased mitochondrial oxidative stress and a range of pathologies. What drives CoQ deficiency in many of these pathologies is unknown, just as there currently is no effective therapeutic strategy to overcome CoQ deficiency in humans. To date, large-scale studies aimed at systematically interrogating endogenous systems that control CoQ biosynthesis and their potential utility to treat disease have not been carried out. Therefore, we developed a quantitative high-throughput method to determine CoQ concentrations in yeast cells. Applying this method to the Yeast Deletion Collection as a genome-wide screen, 30 genes not known previously to regulate cellular concentrations of CoQ were discovered. In combination with untargeted lipidomics and metabolomics, phosphatidylethanolamine N-methyltransferase (PEMT) deficiency was confirmed as a positive regulator of CoQ synthesis, the first identified to date. Mechanistically, PEMT deficiency alters mitochondrial concentrations of one-carbon metabolites, characterized by an increase in the S-adenosylmethionine to S-adenosylhomocysteine (SAM-to-SAH) ratio that reflects mitochondrial methylation capacity, drives CoQ synthesis, and is associated with a decrease in mitochondrial oxidative stress. The newly described regulatory pathway appears evolutionary conserved, as ablation of PEMT using antisense oligonucleotides increases mitochondrial CoQ in mouse-derived adipocytes that translates to improved glucose utilization by these cells, and protection of mice from high-fat diet-induced insulin resistance. Our studies reveal a previously unrecognized relationship between two spatially distinct lipid pathways with potential implications for the treatment of CoQ deficiencies, mitochondrial oxidative stress/dysfunction, and associated diseases., Graphical abstract Image 1, Highlights • Mitochondrial CoQ deficiency results in oxidative stress and a range of pathologies • The drivers of mitochondrial CoQ deficiency remain largely unknown • PEMT deficiency is the first identified positive regulator of mitochondrial CoQ • PEMT deficiency increases CoQ by increasing the mitochondrial SAM-to-SAH ratio • PEMT deficiency prevents insulin resistance by increasing mitochondrial CoQ

Atherosclerosis is a complex, multifactorial disease characterized by the buildup of plaque in the arterial wall. Apolipoprotein E gene deficient (Apoe-/-) mice serve as a commonly used tool to elucidate the pathophysiology of atherosclerosis because of their propensity to spontaneously develop arterial lesions. To date, however, an integrated omics assessment of atherosclerotic lesions in individual Apoe-/- mice has been challenging because of the small amount of diseased and nondiseased tissue available. To address this current limitation, we developed a multiomics method (Multi-ABLE) based on the proteomic method called accelerated Barocycler lysis and extraction (ABLE) to assess the depth of information that can be obtained from arterial tissue derived from a single mouse by splitting ABLE to allow for a combined proteomics-metabolomics-lipidomics analysis (Multi-ABLE). The new method includes tissue lysis via pressure cycling technology (PCT) in a Barocycler, followed by proteomic analysis of half the sample by nanoLC-MS and sequential extraction of lipids (organic extract) and metabolites (aqueous extract) combined with HILIC and reversed phase chromatography and time-of-flight mass spectrometry on the other half. Proteomic analysis identified 845 proteins, 93 of which were significantly altered in lesion-containing arteries. Lipidomic and metabolomic analyses detected 851 lipid and 362 metabolite features, which included 215 and 65 identified lipids and metabolites, respectively. The Multi-ABLE method is the first to apply a concurrent multiomics pipeline to cardiovascular disease using small (

Pancreatic ductal adenocarcinoma (PDAC) develops a pronounced stromal response reflecting an aberrant wound-healing process. This stromal reaction features transdifferentiation of tissue-resident pancreatic stellate cells (PSC) into activated cancer-associated fibroblasts, a process induced by PDAC cells but of unclear significance for PDAC progression. Here, we show that PSCs undergo a dramatic lipid metabolic shift during differentiation in the context of pancreatic tumorigenesis, including remodeling of the intracellular lipidome and secretion of abundant lipids in the activated, fibroblastic state. Specifically, stroma-derived lysophosphatidylcholines support PDAC cell synthesis of phosphatidylcholines, key components of cell membranes, and also facilitate production of the potent wound-healing mediator lysophosphatidic acid (LPA) by the extracellular enzyme autotaxin, which is overexpressed in PDAC. The autotaxin–LPA axis promotes PDAC cell proliferation, migration, and AKT activation, and genetic or pharmacologic autotaxin inhibition suppresses PDAC growth in vivo. Our work demonstrates how PDAC cells exploit the local production of wound-healing mediators to stimulate their own growth and migration. Significance: Our work highlights an unanticipated role for PSCs in producing the oncogenic LPA signaling lipid and demonstrates how PDAC tumor cells co-opt the release of wound-healing mediators by neighboring PSCs to promote their own proliferation and migration. See related commentary by Biffi and Tuveson, p. 578. This article is highlighted in the In This Issue feature, p. 565

Objective: Hmox1 (heme oxygenase-1) is a stress-induced enzyme that catalyzes the degradation of heme to carbon monoxide, iron, and biliverdin. Induction of Hmox1 and its products protect against cardiovascular disease, including ischemic injury. Hmox1 is also a downstream target of the transcription factor HIF-1α (hypoxia-inducible factor-1α), a key regulator of the body’s response to hypoxia. However, the mechanisms by which Hmox1 confers protection against ischemia-mediated injury remain to be fully understood. Approach and Results: Hmox1 deficient ( Hmox1 –/– ) mice had impaired blood flow recovery with severe tissue necrosis and autoamputation following unilateral hindlimb ischemia. Autoamputation preceded the return of blood flow, and bone marrow transfer from littermate wild-type mice failed to prevent tissue injury and autoamputation. In wild-type mice, ischemia-induced expression of Hmox1 in skeletal muscle occurred before stabilization of HIF-1α. Moreover, HIF-1α stabilization and glucose utilization were impaired in Hmox1 –/– mice compared with wild-type mice. Experiments exposing dermal fibroblasts to hypoxia (1% O 2 ) recapitulated these key findings. Metabolomics analyses indicated a failure of Hmox1 –/– mice to adapt cellular energy reprogramming in response to ischemia. Prolyl-4-hydroxylase inhibition stabilized HIF-1α in Hmox1 –/– fibroblasts and ischemic skeletal muscle, decreased tissue necrosis and autoamputation, and restored cellular metabolism to that of wild-type mice. Mechanistic studies showed that carbon monoxide stabilized HIF-1α in Hmox1 –/– fibroblasts in response to hypoxia. Conclusions: Our findings suggest that Hmox1 acts both downstream and upstream of HIF-1α, and that stabilization of HIF-1α contributes to Hmox1’s protection against ischemic injury independent of neovascularization.

Background and Aims The volatile thiols, such as 3‐mercaptohexanol, 3‐mercaptohexyl acetate and 4‐mercapto‐4‐methylpentan‐2‐one, are major aroma compounds responsible for the distinctive tropical notes (passionfruit and grapefruit) in New Zealand Sauvignon Blanc wines. Previously, it was observed that traces of linoleic acid in grape juice can significantly affect the development of these volatile thiols and other aroma compounds in wine. Here, we examined the effect of the fatty acids commonly found in Sauvignon Blanc grape juice as well as juice lipid lipolysis on the development of aroma compounds in the resulting wines. Methods and Results Experiments were performed to increase the total free fatty acid concentration in grape juice through juice treatment with acidic lipase from Aspergillus niger and through co‐cultured fermentations with non‐Saccharomyces lipase‐secreting yeasts. We observed a significant reduction in the production of 3‐mercaptohexyl acetate in fermented wines upon supplementation with linoleic acid and lipase. Supplementation with lipase increased the concentration of 3‐mercaptohexanol and all ethyl esters detected, while reducing the development of acetate esters. Co‐cultured fermentations also affected the development of many classes of aroma compounds. Conclusions This study demonstrated that the aroma profile of wines could be effectively manipulated by altering the amount of free unsaturated fatty acids in grape juice through lipolytic enzyme supplementation or co‐fermentation with non‐Saccharomyces lipolytic yeast species. Significance of the Study The key findings of this study may lead to new winemaking strategies through the manipulation of the lipid component of grape juice.

Metabolic reprogramming is a key hallmark of cancer, but less is known about metabolic plasticity of the same tumor at different sites. Here, we investigated the metabolic adaptation of leukemia in two different microenvironments, the bone marrow and the central nervous system (CNS). We identified a metabolic signature of fatty-acid synthesis in CNS leukemia, highlighting Stearoyl-CoA desaturase (SCD1) as a key player. In vivo SCD1 overexpression increases CNS disease, whilst genetic or pharmacological inhibition of SCD1 decreases CNS load. Overall, we demonstrated that leukemic cells dynamically rewire metabolic pathways to suit local conditions and that targeting these adaptations can be exploited therapeutically.

Statins are widely prescribed inhibitors of the mevalonate pathway, acting to lower systemic cholesterol levels. The mevalonate pathway is critical for tumorigenesis and is frequently upregulated in cancer. Nonetheless, reported effects of statins on tumor progression are ambiguous, making it unclear whether statins, alone or in combination, can be used for chemotherapy. Here, using advanced mass spectrometry and isotope tracing, we showed that statins only modestly affected cancer cholesterol homeostasis. Instead, they significantly reduced synthesis and levels of another downstream product, the mitochondrial electron carrier coenzyme Q, both in cultured cancer cells and tumors. This compromised oxidative phosphorylation, causing severe oxidative stress. To compensate, cancer cells upregulated antioxidant metabolic pathways, including reductive carboxylation, proline synthesis, and cystine import. Targeting cystine import with an xCT transporter–lowering MEK inhibitor, in combination with statins, caused profound tumor cell death. Thus, statin-induced ROS production in cancer cells can be exploited in a combinatorial regimen. Significance: Cancer cells induce specific metabolic pathways to alleviate the increased oxidative stress caused by statin treatment, and targeting one of these pathways synergizes with statins to produce a robust antitumor response. See related commentary by Cordes and Metallo, p. 151

This study presents a comprehensive lipidome analysis of Sauvignon blanc grape juice by combining GC-MS based fatty acid profiling with shotgun lipidomics strategy. We observed that despite grape juice being a water based matrix it contains a diverse range of lipid species, including common saturated and unsaturated free and intact fatty acids as well as odd-numbered and hydroxy fatty acids. Based on GC-MS quantitative data, we found that the total lipid content of grape juice could be as high as 2.80 g/L. The majority of lipids were present in the form of complex lipids with relatively small amount of free fatty acids (15%). Therefore we concluded that the lipidome should be considered an important component of grape juice with the potential to impact on fermentation processes as well as on the sensorial properties of fermented products. This work serves as a hypothesis generating tool, the results of which justify follow-up studies to explore the influence of the grape juice lipidome and lipid metabolism in yeast on the aroma profile of wine.

Succinate dehydrogenase (SDH) is a hetero-tetrameric nuclear-encoded complex responsible for the oxidation of succinate to fumarate in the tricarboxylic acid (TCA) cycle. Loss-of-function mutations in any of the SDH genes are associated with cancer formation. However, the impact of SDH loss on cell metabolism and the mechanisms enabling growth of SDH-defective cells are largely unknown. Here, we generated Sdhb-ablated kidney mouse cells and employed comparative metabolomics and stable isotope-labelling approaches to identify nutritional requirements and metabolic adaptations to SDH loss. We found that lack of SDH activity commits cells to consume extracellular pyruvate, which sustains Warburg-like bioenergetic features. We further demonstrated that pyruvate carboxylation diverts glucose-derived carbons into aspartate biosynthesis, thus sustaining cell growth. By identifying pyruvate carboxylase as an essential gene for the proliferation and tumorigenic capacity of SDH-deficient cells, this study revealed a metabolic vulnerability for potential future treatment of SDH-associated malignancies.

Cultures of dissociated hippocampal neurons are often used to study neuronal cell biology. We report that the development of these neurons is strongly affected by chemicals leaching from commonly used disposable medical-grade syringes and syringe filters. Contamination of culture medium by bioactive substance(s) from syringes and filters occurred with multiple manufacturing lots and filter types under normal use conditions and resulted in changes to neurite growth, axon formation and the neuronal microtubule cytoskeleton. The effects on neuronal morphology were concentration dependent and significant effects were detected even after substantial dilution of the contaminated medium. Gas chromatography-mass spectrometry analyses revealed many chemicals eluting from the syringes and filters. Three of these chemicals (stearic acid, palmitic acid and 1,2-ethanediol monoacetate) were tested but showed no effects on neurite growth. Similar changes in neuronal morphology were seen with high concentrations of bisphenol A and dibutyl phthalate, two hormonally active plasticisers. Although no such compounds were detected by gas chromatography–mass spectrometry, unknown plasticisers in leachates may affect neurites. This is the first study to show that leachates from laboratory consumables can alter the growth of cultured hippocampal neurons. We highlight important considerations to ensure leachate contamination does not compromise cell biology experiments.

Melanoma cells steer out of tumours using self-generated lysophosphatidic acid (LPA) gradients. The cells break down LPA, which is present at high levels around the tumours, creating a dynamic gradient that is low in the tumour and high outside. They then migrate up this gradient, creating a complex and evolving outward chemotactic stimulus. Here, we introduce a new assay for self-generated chemotaxis, and show that raising LPA levels causes a delay in migration rather than loss of chemotactic efficiency. Knockdown of the lipid phosphatase LPP3 – but not of its homologues LPP1 or LPP2 – diminishes the cell's ability to break down LPA. This is specific for chemotactically active LPAs, such as the 18:1 and 20:4 species. Inhibition of autotaxin-mediated LPA production does not diminish outward chemotaxis, but loss of LPP3-mediated LPA breakdown blocks it. Similarly, in both 2D and 3D invasion assays, knockdown of LPP3 diminishes the ability of melanoma cells to invade. Our results demonstrate that LPP3 is the key enzyme in the breakdown of LPA by melanoma cells, and confirm the importance of attractant breakdown in LPA-mediated cell steering. This article has an associated First Person interview with the first author of the paper., Highlighted Article: Melanoma cells can create and follow their own gradients of attractant, via a new mechanism by which tumour cells may undergo metastasis.

We evaluated the quality and content of fish oil supplements in New Zealand. All encapsulated fish oil supplements marketed in New Zealand were eligible for inclusion. Fatty acid content was measured by gas chromatography. Peroxide values (PV) and anisidine values (AV) were measured, and total oxidation values (Totox) calculated. Only 3 of 32 fish oil supplements contained quantities of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) that were equal or higher than labelled content, with most products tested (69%) containing

Serine catabolism results in formate efflux that exceeds anabolic demands for purine synthesis., Serine catabolism to glycine and a one-carbon unit has been linked to the anabolic requirements of proliferating mammalian cells. However, genome-scale modeling predicts a catabolic role with one-carbon release as formate. We experimentally prove that in cultured cancer cells and nontransformed fibroblasts, most of the serine-derived one-carbon units are released from cells as formate, and that formate release is dependent on mitochondrial reverse 10-CHO-THF synthetase activity. We also show that in cancer cells, formate release is coupled to mitochondrial complex I activity, whereas in nontransformed fibroblasts, it is partially insensitive to inhibition of complex I activity. We demonstrate that in mice, about 50% of plasma formate is derived from serine and that serine starvation or complex I inhibition reduces formate synthesis in vivo. These observations transform our understanding of one-carbon metabolism and have implications for the treatment of diabetes and cancer with complex I inhibitors.

Technological advances in the area of analytical chemistry and the development of state-of-the-art analytical instrumentation have allowed for a shift in the focus from a previously strict targeted approach towards the approach adopted in metabolomics, the essence of which is non-targeted providing an unbiased analysis of metabolites in biological samples. Metabolite profiling methods have served their purpose in providing descriptive information about biological systems through qualitative and relative semi-quantitative data. However, quantitative characterization of a system cannot be fully accomplished without using absolute metabolite concentrations, an area which is lacking in most current metabolomics platforms. The objective of this work was to develop a calibration-curve free method for quantitation of amino and non-amino organic acids in biological samples. We developed a novel calibration curve-free GC–MS method based on isotope-coded derivatization for absolute non-targeted quantification of polar metabolites. A new R-based package MetabQ was created for automated data processing of GC–MS data files performing data extraction and calculation of absolute metabolite values. The new method requires metabolite response factors which should be calculated only once for each equipment, and was validated for metabolite quantification of different biological matrices. The method showed high reproducibility and accuracy, and does not require the use of calibration curves using standards to be analyzed in parallel with every sample batch. However, there is a small group of metabolites where their quantification required additional steps of correction due to their chemical instability. The introduced R package significantly increased the throughput in the data analysis process, extensively reducing the time required to perform the task manually. Our novel approach gives the potential to identify and quantify hundreds of metabolites, far exceeding the capabilities of any absolute quantitative targeted metabolite analysis, limited only by the size of the mass spectral library.

The recent history and the current state of relations between Russia and the European Union (EU) reflect the geopolitical competition between the two powers over their contested neighbourhood – Ukraine, Belarus and Moldova. Although prioritizing the EU as its major trade and political partner, Russia is also conscious of its strategic interests in the ‘near abroad’. While there is little discrimination in the political discourse of Russian elites in relation to the European Neighbourhood Policy (ENP), in reality, as popular opinion indicates, those neighbours who openly show their allegiances to the EU – Ukraine, Georgia and Moldova – have been increasingly categorized as hostile and unfriendly towards Russia. Public opinion remains strikingly congruent with and reflective of government foreign policy preferences.

Cells can synthesize fatty acids by ligating multiple acetyl units from acetyl-CoA. This is followed by desaturation and elongation reactions to produce a variety of fatty acids required for proper cellular functioning. Alternatively, exogenous lipid sources can contribute to cellular fatty acid pools. Here, we present a method based on incorporation of (13)C-carbon from labeled substrates into fatty acids and subsequent mass spectrometry analysis. The resulting labeling patterns can be used to determine (1) (13)C-enrichment of lipogenic acetyl-CoA, (2) the relative contributions of synthesis and uptake, and (3) absolute fatty acid fluxes. We begin by providing a background and general principles regarding the use of stable isotopes to study fatty acid metabolism. We then proceed with detailing procedures for sample preparation and both GC-MS and LC-MS analysis of isotope incorporation. Finally, we discuss the interpretation of the resulting fatty acid-labeling patterns.

We evaluated the quality and content of fish oil supplements in New Zealand. All encapsulated fish oil supplements marketed in New Zealand were eligible for inclusion. Fatty acid content was measured by gas chromatography. Peroxide values (PV) and anisidine values (AV) were measured and total oxidation values (Totox) calculated. Only 3 of 32 fish oil supplements contained quantities of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) that were equal or higher than labelled content, with most products tested (69%) containing

Cells can synthesize fatty acids by ligating multiple acetyl units from acetyl-CoA. This is followed by desaturation and elongation reactions to produce a variety of fatty acids required for proper cellular functioning. Alternatively, exogenous lipid sources can contribute to cellular fatty acid pools. Here, we present a method based on incorporation of (13)C-carbon from labeled substrates into fatty acids and subsequent mass spectrometry analysis. The resulting labeling patterns can be used to determine (1) (13)C-enrichment of lipogenic acetyl-CoA, (2) the relative contributions of synthesis and uptake, and (3) absolute fatty acid fluxes. We begin by providing a background and general principles regarding the use of stable isotopes to study fatty acid metabolism. We then proceed with detailing procedures for sample preparation and both GC-MS and LC-MS analysis of isotope incorporation. Finally, we discuss the interpretation of the resulting fatty acid-labeling patterns.

Bleaching (dinoflagellate symbiont loss) is one of the greatest threats facing coral reefs. The functional cnidarian-dinoflagellate symbiosis, which forms coral reefs, is based on the bi-directional exchange of nutrients. During thermal stress this exchange breaks down, however major gaps remain in our understanding of the roles of free metabolite pools in symbiosis and homeostasis. In this study we applied gas chromatography-mass spectrometry (GC-MS) to explore thermally induced changes in intracellular pools of amino and non-amino organic acids in each partner of the model sea anemone Aiptasia sp. and its dinoflagellate symbiont. Elevated temperatures (32°C for 6 d) resulted in symbiont photoinhibition and bleaching. Thermal stress induced distinct changes in the metabolite profiles of both partners, associated with alterations to central metabolism, oxidative state, cell structure, biosynthesis and signalling. Principally, we detected elevated pools of polyunsaturated fatty acids (PUFAs) in the symbiont, indicative of modifications to lipogenesis/lysis, membrane structure and nitrogen assimilation. In contrast, reductions of multiple PUFAs were detected in host pools, indicative of increased metabolism, peroxidation and/or reduced translocation of these groups. Accumulations of glycolysis intermediates were also observed in both partners, associated with photoinhibition and downstream reductions in carbohydrate metabolism. Correspondingly, we detected accumulations of amino acids and intermediate groups in both partners, with roles in gluconeogenesis and acclimation responses to oxidative stress. These data further our understanding of cellular responses to thermal stress in the symbiosis and generates hypotheses relating to the secondary roles of a number of compounds in homeostasis and heat stress resistance.

Oxygen and oxidative stress have become relevant components in clarifying the mechanism that weakens bacterial cells in parallel to the mode of action of bactericidal antibiotics. Given the importance of oxidative stress in the overall defense mechanism of bacteria and their apparent role in the antimicrobial mode of action, it is important to understand how bacteria respond to this stress at a metabolic level. The aim of this study was to determine the impact of oxygen on the metabolism of the facultative anaerobe Enterococcus faecalis using continuous culture, metabolomics and 13C-enrichment of metabolic intermediates. When E. faecalis was rapidly transitioned from anaerobic to aerobic growth, cellular metabolism was directed towards intracellular glutathione production and glycolysis was upregulated two-fold, which increased the supply of critical metabolite precursors (e.g. glycine and glutamate) for sulfur metabolism and glutathione biosynthesis as well as reducing power for cellular respiration in the presence of haemin. The ultimate metabolic response of E. faecalis to an aerobic environment was the upregulation of fatty acid metabolism and benzoate degradation, which was linked to important changes in the bacterial membrane composition as evidenced by changes in membrane fatty acid composition and the reduction of membrane-associated demethylmenaquinone. These key metabolic pathways associated with the response of E. faecalis to oxygen may represent potential new targets to increase the susceptibility of this bacterium to bactericidal drugs., This work was funded by the HRC (Health and Research Council of New Zealand) and the FCT (Portuguese Foundation for Science and Technology), with grant reference SFRH/BD/47016/2008.

One-thousand two-hundred and ninety university students from twenty-seven ethnic⧹national groups across six locations in the former Soviet Union (Novopolotsk in Byelorussia, Kharkov in the Ukraine, Moscow and Nizniy Novgorod in Russia, Ufa in the Bashkir Autonomous Republic and Ulan-Ude in the Buryat Autonomous Republic) participated in 1991–1992 in a survey investigating the existence of ingroup preference in inter-ethnic contact, ingroup consensus on an ethnic hierarchy of outgroups and the measure of intergroup consensus on an ethnic hierarchy among the ethnic⧹national groups in each location. Hypotheses about ingroup preference, ethnic hierarchies and consensus were derived from realistic group conflict theory, social identity theory and from the assumption that outgroup preferences reflect considerations of the status effects of intergroup contact. It appeared that the last type of hypothesis could explain most of the results.

Research in the United States and Europe has focused on the prejudice of majority groups towards minority groups, the implication somehow being that majority groups were more prejudiced than minority groups. In the former Soviet Union, ethnic environments were more complex; the same ethnic group could be a majority in one region but a minority in others. Using a sample of 1,459 first‐ and fourth‐year university students from eight regions of the former USSR, this study focuses on Russian, Tatar and Ukrainian respondents (n = 821) to test the hypothesis that the status of an ethnic group (majority/minority) or in‐group bias explains members’ prejudice. According to in‐group bias, all ethnic groups are equally prejudiced, minority and majority alike, whereas group status posits that groups in a majority position are more prejudiced. Findings show that group status has greater impact on prejudice than does in‐group bias. This applies, however, only to Russians. Interpretations of the findings rest o...

The legends of Figures 1 and 2 were swapped. The correct version of those figure legends are available below: Figure 1. Growth curves of C. albicans cells grown in the presence (red) or absence (blue) of phenylethyl alcohol (15 mM). Figure 2. The morphology of C. albicans cells incubated in the presence of different concentrations of phenylethyl alcohol in minimal mineral medium (MM) at 37°C for 12 h. The Images were taken by Nomarksi contrast microscopy with 800× magnification.