Abstract WP268: Roles And Regulation Of Ketogenesis In Cultured Astroglia And Neurons Under Hypoxia

Stroke ◽  
2013 ◽  
Vol 44 (suppl_1) ◽  
Author(s):  
Shinichi Takahashi ◽  
Takuya Iizumi ◽  
Takato Abe ◽  
Norihiro Suzuki
Keyword(s):  
Alternative Energy ◽  
Ketone Bodies ◽  
Cell Damage ◽  
Tca Cycle ◽  
Neural Cells ◽  
Lactate Oxidation ◽  
Energy Substrates ◽  
The Tca Cycle ◽  
A Cell ◽  
Neuronal Energy Metabolism

Purpose: Although exogenous ketone bodies (KBs), acetoacetate (AA) and β-hydroxybutyrate (BHB) can serve as alternative energy substrates in neural cells under starvation, the exact roles and the regulation of ketogenesis in the brain remain uncertain. The present study examined the ketogenic capacity of cultured rat astroglia under hypoxia and possible roles and the regulation of KBs in neuronal energy metabolism. Methods: Primary neurons and secondary astroglia were prepared from SD rats. Palmitic acid (PA) and L-carnitine (LC) were added to the nutrient medium. AA and BHB produced and released in the medium during 24h were measured using the cyclic thio-NADH method. 14 C-labeled acid-soluble products (i.e., KBs) and 14 CO 2 produced from [1- 14 C]PA were measured to assess the role of PA and KBs as energy substrates in the TCA cycle. [U- 14 C]lactate or [1- 14 C]BHB was used to compare the oxidative metabolism of the end-products of glycolysis with those of the β-oxidation of fatty acid. Some cells were placed in a hypoxic chamber (1%O 2 ) for 12h-24h to evaluate the effects of hypoxia and re-oxygenation on KB metabolism. Results: PA (100 μM) and LC (1 mM) induced higher KB production (mean ± SD in pmol/μg /24h, n=6) in astroglia (AA: 40.8 ± 1.3, BHB: 9.6 ± 1.4) than in neurons (AA: 0.3 ± 1.5, BHB: 2.2 ± 2.7), while CO 2 production from PA was less than 5% of the KB production in astroglia. KB production was augmented by AICAR (500 μM), a cell-permeable AMPK activator (AA: 66.8 ± 8.7, BHB: 12.7 ± 0.9) as well as hypoxia (AA: 59.4 ± 5.1, BHB: 30.0 ± 14.4) in astroglia. [1- 14 C]BHB oxidation (mean ± SD in pmol/μg/h, n=4) in neurons (2.9 ± 0.6) was about 1.5 times as high as that in astroglia (1.8 ± 0.1). [U- 14 C]lactate oxidation in astroglia (14.0 ± 4.6) was not affected by addition of BHB (13.3 ± 4.7) in astroglia, while that in neurons (21.5 ± 2.8) was reduced by 15% (18.1 ± 1.2, P <0.05). Conclusions: Astroglia responded to hypoxia by enhancing KB production in the presence of PA and LC, and KBs produced by astroglia might serve as a neuronal energy substrate for the TCA cycle in place of lactate, since pyruvate dehydrogenase is susceptible to ischemia. The activation of astroglial ketogenesis may reduce ischemic cell damage after stroke.

137 ROLE OF GLUCOSE AND FATTY ACID METABOLISM IN PORCINE EARLY EMBRYO DEVELOPMENT

2008 ◽  
Vol 20 (1) ◽  
pp. 149 ◽  
Author(s):  
R. G. Sturmey ◽  
H. J. Leese
Keyword(s):  
Energy Source ◽  
Fatty Acid ◽  
Glucose Metabolism ◽  
Embryo Development ◽  
Alternative Energy ◽  
Tca Cycle ◽  
Early Embryo ◽  
Metabolic Energy ◽  
The Tca Cycle

Glucose metabolism plays an important role in the preimplantation development of porcine embryos in vitro. As in mammalian species generally, a proportion of glucose consumed is converted to lactate by aerobic glycolysis generating small amounts of ATP, with the remainder oxidized by the TCA cycle. However, a striking feature of the porcine early embryo is the large amount of lipid present as triglyceride (TG), which represents an alternative energy source. The TG is metabolized via β-oxidation, producing acetyl Co A, which in turn is oxidized by the TCA cycle. This sequence of reactions requires a constant supply of carbohydrate to provide oxaloacetate (OA) to prime the TCA cycle. The provision of OA from pyruvate arising from glycolysis may represent an alternative role for glucose in early pig embryo development. We have therefore sought to determine the importance of interplay between glucose and TG metabolism in porcine embryos in vitro. Porcine embryos were generated in vitro by fertilization of in vitro-matured oocytes collected from abattoir-derived ovaries. Oocytes were matured in defined maturation medium and embryos cultured in NCSU23. Glucose consumption, lactate production, and TG content of single porcine blastocysts cultured throughout development in the presence of methyl palmoxirate (MP), an inhibitor of TG metabolism, were measured as described by Sturmey RG and Leese HJ 2003 (Reprod. 126, 197–204). The capacity of zygotes to form blastocysts when cultured with OA in place of glucose in the presence or absence of MP and the amount of TG in blastocysts grown in either glucose or OA-containing medium were then determined (6 replicates). When TG metabolism was inhibited, porcine blastocysts consumed significantly more glucose (32 � 9 pmol/embryo/h v. 11 � 1 pmol/embryo/h; P < 0.05; n = 34) and produced higher amounts of lactate (35 � 4 pmol/embryo/h v. 10 � 0.8 pmol/embryo/h; P < 0.01; n = 34). Blastocyst rates did not differ significantly between embryos grown in the presence of glucose or OA, and in blastocysts grown in OA-NCSU the TG content was significantly reduced (155 � 8 ng v. 240 � 12 ng; P = 0.015; n = 41). All embryos cultured in OA-containing medium in the presence of MP failed to develop beyond the zygote stage. The data support the notion that porcine embryos can use endogenous TG as a metabolic energy source. When this is prevented by chemical inhibition, the embryo upregulates glycolysis and glucose oxidation as an alternate means of generating ATP. When cultured in medium containing OA, a compound that cannot generate ATP per se, embryo development was similar to controls, again suggesting the ability to use endogenous energy stores, a proposition reinforced by a significant fall in the levels of TG in the presence of OA. However, by inhibiting β-oxidation in the absence of glucose, porcine embryos were unable to develop. The relationship between TG and glucose metabolism by porcine embryos is analogous to the glucose/fatty acid cycle in whole animals where glucose and TG can be used as energy sources, but in a reciprocal manner. The data also demonstrate the plasticity of energy metabolism by porcine early embryos.

scholarly journals Waste Not, Want Not: Lactate Oxidation Fuels the TCA Cycle

2017 ◽  
Vol 26 (6) ◽  
pp. 803-804 ◽  
Author(s):  
Inmaculada Martínez-Reyes ◽  
Navdeep S. Chandel
Keyword(s):  
Tca Cycle ◽  
Lactate Oxidation ◽  
The Tca Cycle

Substrate metabolism of isolated jejunal epithelium: conservation of three-carbon units

1986 ◽  
Vol 250 (2) ◽  
pp. C191-C198 ◽  
Author(s):  
R. T. Mallet ◽  
J. K. Kelleher ◽  
M. J. Jackson
Keyword(s):  
Ketone Bodies ◽  
Tca Cycle ◽  
Substrate Metabolism ◽  
Metabolic Substrates ◽  
The Tca Cycle ◽  
Carbon Compounds ◽  
Consumption Rates ◽  
14Co2 Production ◽  
Tca Cycle Intermediates ◽  
Labeling Pattern

This study characterizes the substrate metabolism of isolated jejunal epithelial cells. Utilization of substrates was assessed by spectrophotometric assay. Significant quantities of glucose, glutamine, and ketone bodies were consumed in a 1-h period; lactate and ammonia were produced. [U-14C]glucose was metabolized in this medium to approximately three moles of lactate per mole of CO2. The pattern of tricarboxylic acid (TCA) cycle metabolism was analyzed utilizing media containing different concentrations of potential metabolic substrates and trace quantities of [14C]- succinate. O2 consumption rates indicated that glutamine can serve as an energy source in the absence of other substrates. Relative 14CO2 production from [1,4-14C]succinate versus [2,3-14C]succinate, which estimates flux of TCA cycle intermediates to products other than CO2, was increased more than twofold when glutamine was the only major substrate available. Alanine was produced from TCA cycle intermediates. Analysis of the citrate labeling pattern in the presence of [2,3-14C] succinate suggested that carbon from the TCA cycle does not form a significant fraction of acetyl-CoA used for citrate synthesis and that glutamine carbon was not completely oxidized to CO2. These findings suggest that glucose and glutamine are converted to three-carbon compounds by the jejunal epithelium.

scholarly journals DDRE-01. METABOLIC PLASTICITY AND HETEROGENEITY IN IDH1MUT CELL LINES PRODUCES RESISTANCE TO GLUTAMINASE INHIBITION BY CB839

2020 ◽  
Vol 3 (Supplement_1) ◽  
pp. i6-i6
Author(s):  
Mioara Larion ◽  
Victor Ruiz-Rodado
Keyword(s):  
Cell Lines ◽  
Metabolic Profiling ◽  
Cellular Proliferation ◽  
Lactate Production ◽  
Tca Cycle ◽  
Heterogeneous Landscape ◽  
The Tca Cycle ◽  
A Cell ◽  
Metabolic Tracing ◽  
Mutant Idh1

Abstract BACKGROUND Mutant IDH1 (IDH1mut) gliomas have characteristic genetic and metabolic profiles and exhibit phenotype that is distinct from their wild-type counterparts. The glutamine/glutamate pathway has been hypothesized as a selective therapeutic target in IDH1mut gliomas. However, little information exists on the contribution of this pathway to the formation of D-2-hydroxyglutarate (D-2HG), a hallmark of IDHmut cells, and the metabolic consequences of inhibiting this pathway. METHODS We employed an untargeted metabolic profiling approach in order to detect metabolic changes arising from glutaminase (GLS) inhibition treatment. Subsequently, 13C metabolic tracing analysis through a combined Nuclear Magnetic Resonance and Liquid Chromatography-Mass Spectrometry approach, we explored the fate of glutamine and glucose under treatment with CB839 a glutaminase-GLS-inhibitor and their respective contributions to D-2HG formation. RESULTS AND CONCLUSIONS The effects of CB839 on cellular proliferation differed among the cell lines tested, leading to designations of GLS-inhibition super-sensitive, -sensitive or -resistant. Our data indicates a decrease in the production of downstream metabolites of glutamate, including those involved in the TCA cycle, when treating the sensitive cells with CB839 (glutaminase -GLS- inhibitor). Notably, CB839-sensitive IDH1mutcells respond to GLS inhibition by upregulating glycolysis and lactate production. In contrast, CB839-resistantIDH1mut cell lines do not rely only on glutamine for the sustenance of TCA cycle. In these cells, glucose contribution to TCA is enough to compensate the downregulation of glutamine-derived TCA metabolites. This investigation reveals that the glutamine/glutamate pathway contributes differentially to D-2HG in a cell-line dependent fashion on a panel of IDHmut cell lines. Further, these results demonstrate that there is a heterogeneous landscape of IDH1mut metabolic phenotypes. This underscores the importance of detailed metabolic profiling of IDH1mut patients prior to the decision to target glutamine/glutamate pathway clinically.

Glucose alleviates ammonia-induced inhibition of short-chain fatty acid metabolism in rat colonic epithelial cells

2003 ◽  
Vol 285 (1) ◽  
pp. G105-G114 ◽  
Author(s):  
John D. Cremin ◽  
Mark D. Fitch ◽  
Sharon E. Fleming
Keyword(s):  
Fatty Acid ◽  
Epithelial Cells ◽  
Short Chain Fatty Acid ◽  
Ketone Bodies ◽  
Tca Cycle ◽  
Chain Fatty Acid ◽  
Short Chain ◽  
Colonic Epithelial Cells ◽  
The Tca Cycle

Ammonia decreased metabolism by rat colonic epithelial cells of butyrate and acetate to CO2 and ketones but increased oxidation of glucose and glutamine. Ammonia decreased cellular concentrations of oxaloacetate for all substrates evaluated. The extent to which butyrate carbon was oxidized to CO2 after entering the tricarboxylic acid (TCA) cycle was not significantly influenced by ammonia, suggesting there was no major shift toward efflux of carbon from the TCA cycle. Ammonia reduced entry of butyrate carbon into the TCA cycle, and the proportion of CoA esterified with acetate and butyrate correlated positively with the production of CO2 and ketone bodies. Also, ammonia reduced oxidation of propionate but had no effect on oxidation of 3-hydroxybutyrate. Inclusion of glucose, lactate, or glutamine with butyrate and acetate counteracted the ability of ammonia to decrease their oxidation. In rat colonocytes, it appears that ammonia suppresses short-chain fatty acid (SCFA) oxidation by inhibiting a step before or during their activation. This inhibition is alleviated by glucose and other energy-generating compounds. These results suggest that ammonia may only affect SCFA metabolism in vivo when glucose availability is compromised.

scholarly journals Modulation of cerebral ketone metabolism following traumatic brain injury in humans

2018 ◽  
Vol 40 (1) ◽  
pp. 177-186 ◽  
Author(s):  
Adriano Bernini ◽  
Mojgan Masoodi ◽  
Daria Solari ◽  
John-Paul Miroz ◽  
Laurent Carteron ◽  
...  
Keyword(s):  
Traumatic Brain Injury ◽  
Brain Injury ◽  
Alternative Energy ◽  
Metabolic Response ◽  
Ketone Bodies ◽  
Cerebral Microdialysis ◽  
Interstitial Tissue ◽  
Energy Substrates ◽  
Medium Chain ◽  
Total Brain

Adaptive metabolic response to injury includes the utilization of alternative energy substrates – such as ketone bodies (KB) – to protect the brain against further damage. Here, we examined cerebral ketone metabolism in patients with traumatic brain injury (TBI; n = 34 subjects) monitored with cerebral microdialysis to measure total brain interstitial tissue KB levels (acetoacetate and β-hydroxybutyrate). Nutrition – from fasting vs. stable nutrition state – was associated with a significant decrease of brain KB (34.7 [10th–90th percentiles 10.7–189] µmol/L vs. 13.1 [6.5–64.3] µmol/L, p < 0.001) and blood KB (668 [168.4–3824.9] vs. 129.4 [82.6–1033.8] µmol/L, p < 0.01). Blood KB correlated with brain KB (Spearman’s rho 0.56, p = 0.0013). Continuous feeding with medium-chain triglycerides-enriched enteral nutrition did not increase blood KB, and provided a modest increase in blood and brain free medium chain fatty acids. Higher brain KB at the acute TBI phase correlated with age and brain lactate, pyruvate and glutamate, but not brain glucose. These novel findings suggest that nutritional ketosis was the main determinant of cerebral KB metabolism following TBI. Age and cerebral metabolic distress contributed to brain KB supporting the hypothesis that ketones might act as alternative energy substrates to glucose. Further studies testing KB supplementation after TBI are warranted.

scholarly journals A comprehensive analysis of myocardial substrate preference emphasizes the need for a synchronized fluxomic/metabolomic research design

2017 ◽  
Vol 312 (6) ◽  
pp. H1215-H1223 ◽  
Author(s):  
Mukundan Ragavan ◽  
Alexander Kirpich ◽  
Xiaorong Fu ◽  
Shawn C. Burgess ◽  
Lauren M. McIntyre ◽  
...  
Keyword(s):  
Myocardial Metabolism ◽  
Ketone Bodies ◽  
Tca Cycle ◽  
Oxidative Capacity ◽  
Total Carbohydrate ◽  
Atp Production ◽  
The Tca Cycle ◽  
Isotopomer Analysis ◽  
Myocardial Substrate ◽  
Correct Estimation

The heart oxidizes fatty acids, carbohydrates, and ketone bodies inside the tricarboxylic acid (TCA) cycle to generate the reducing equivalents needed for ATP production. Competition between these substrates makes it difficult to estimate the extent of pyruvate oxidation. Previously, hyperpolarized pyruvate detected propionate-mediated activation of carbohydrate oxidation, even in the presence of acetate. In this report, the optimal concentration of propionate for the activation of glucose oxidation was measured in mouse hearts perfused in Langendorff mode. This study was performed with a more physiologically relevant perfusate than the previous work. Increasing concentrations of propionate did not cause adverse effects on myocardial metabolism, as evidenced by unchanged O2 consumption, TCA cycle flux, and developed pressures. Propionate at 1 mM was sufficient to achieve significant increases in pyruvate dehydrogenase flux (3×), and anaplerosis (6×), as measured by isotopomer analysis. These results further demonstrate the potential of propionate as an aid for the correct estimation of total carbohydrate oxidative capacity in the heart. However, liquid chromotography/mass spectroscopy-based metabolomics detected large changes (~30-fold) in malate and fumarate pool sizes. This observation leads to a key observation regarding mass balance in the TCA cycle; flux through a portion of the cycle can be drastically elevated without changing the O2 consumption.

Liquid-liquid extraction of succinate using a dicopper cryptate

2020 ◽  
Author(s):  
Riccardo Mobili ◽  
Sonia La Cognata ◽  
Francesca Merlo ◽  
Andrea Speltini ◽  
Massimo Boiocchi ◽  
...  
Keyword(s):  
Aqueous Phase ◽  
Visible Spectrum ◽  
Tca Cycle ◽  
Residual Concentration ◽  
Waste Products ◽  
Liquid Liquid Extraction ◽  
The Tca Cycle ◽  
Quantitative Extraction ◽  
Uv Visible

<div> <p>The extraction of the succinate dianion from a neutral aqueous solution into dichloromethane is obtained using a lipophilic cage-like dicopper(II) complex as the extractant. The quantitative extraction exploits the high affinity of the succinate anion for the cavity of the azacryptate. The anion is effectively transferred from the aqueous phase, buffered at pH 7 with HEPES, into dichloromethane. A 1:1 extractant:anion adduct is obtained. Extraction can be easily monitored by following changes in the UV-visible spectrum of the dicopper complex in dichloromethane, and by measuring the residual concentration of succinate in the aqueous phase by HPLC−UV. Considering i) the relevance of polycarboxylates in biochemistry, as e.g. normal intermediates of the TCA cycle, ii) the relevance of dicarboxylates in the environmental field, as e.g. waste products of industrial processes, and iii) the recently discovered role of succinate and other dicarboxylates in pathophysiological processes including cancer, our results open new perspectives for research in all contexts where selective recognition, trapping and extraction of polycarboxylates is required. </p> </div>

scholarly journals Mitochondrial Mistranslation in Brain Provokes a Metabolic Response Which Mitigates the Age-Associated Decline in Mitochondrial Gene Expression

2021 ◽  
Vol 22 (5) ◽  
pp. 2746
Author(s):  
Dimitri Shcherbakov ◽  
Reda Juskeviciene ◽  
Adrián Cortés Sanchón ◽  
Margarita Brilkova ◽  
Hubert Rehrauer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Metabolic Response ◽  
Mitochondrial Gene ◽  
Tca Cycle ◽  
Brain Mitochondria ◽  
Mitochondrial Gene Expression ◽  
Rna Seq ◽  
The Tca Cycle ◽  
Neurological Phenotype

Mitochondrial misreading, conferred by mutation V338Y in mitoribosomal protein Mrps5, in-vivo is associated with a subtle neurological phenotype. Brain mitochondria of homozygous knock-in mutant Mrps5V338Y/V338Y mice show decreased oxygen consumption and reduced ATP levels. Using a combination of unbiased RNA-Seq with untargeted metabolomics, we here demonstrate a concerted response, which alleviates the impaired functionality of OXPHOS complexes in Mrps5 mutant mice. This concerted response mitigates the age-associated decline in mitochondrial gene expression and compensates for impaired respiration by transcriptional upregulation of OXPHOS components together with anaplerotic replenishment of the TCA cycle (pyruvate, 2-ketoglutarate).

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