In vivo 2H/13C flux analysis in metabolism research

In vivo 13C magnetic resonance spectroscopy (MRS) enables the investigation of cerebral metabolic compartmentation while, e.g. infusing 13C-labeled glucose. Metabolic flux analysis of 13C turnover previously yielded quantitative information of glutamate and glutamine metabolism in humans and rats, while the application to in vivo mouse brain remains exceedingly challenging. In the present study, 13C direct detection at 14.1 T provided highly resolved in vivo spectra of the mouse brain while infusing [1,6-13C2]glucose for up to 5 h. 13C incorporation to glutamate and glutamine C4, C3, and C2 and aspartate C3 were detected dynamically and fitted to a two-compartment model: flux estimation of neuron-glial metabolism included tricarboxylic acid cycle (TCA) flux in astrocytes (Vg = 0.16 ± 0.03 µmol/g/min) and neurons (VTCAn = 0.56 ± 0.03 µmol/g/min), pyruvate carboxylase activity (VPC = 0.041 ± 0.003 µmol/g/min) and neurotransmission rate (VNT = 0.084 ± 0.008 µmol/g/min), resulting in a cerebral metabolic rate of glucose (CMRglc) of 0.38 ± 0.02 µmol/g/min, in excellent agreement with that determined with concomitant 18F-fluorodeoxyglucose positron emission tomography (18FDG PET).We conclude that modeling of neuron-glial metabolism in vivo is accessible in the mouse brain from 13C direct detection with an unprecedented spatial resolution under [1,6-13C2]glucose infusion.

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Novel biological insights through metabolomics and 13C-flux analysis

Current Opinion in Microbiology ◽

10.1016/j.mib.2009.08.003 ◽

2009 ◽

Vol 12 (5) ◽

pp. 553-558 ◽

Cited By ~ 99

Author(s):

Nicola Zamboni ◽

Uwe Sauer

Keyword(s):

Flux Analysis ◽

13C Flux Analysis

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Cardiac-specific VEGFB overexpression reduces lipoprotein lipase activity and improves insulin action in rat heart

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00219.2021 ◽

2021 ◽

Author(s):

Rui Shang ◽

Nathaniel Lal ◽

ChaeSyng Lee ◽

Yajie Zhai ◽

Karanjit Puri ◽

...

Keyword(s):

Insulin Sensitivity ◽

Lipoprotein Lipase ◽

Insulin Action ◽

Insulin Signalling ◽

Isolated Heart ◽

Cardiac Metabolism ◽

Metabolic Reprogramming ◽

Flux Analysis

Cardiac muscle utilizes multiple sources of energy including glucose and fatty acid (FA). The heart cannot synthesize FA and relies on obtaining it from other sources, with lipoprotein lipase (LPL) breakdown of lipoproteins suggested to be a key source of FA for cardiac use. Recent work has indicated that cardiac vascular endothelial growth factor B (VEGFB) overexpression expands the coronary vasculature and facilitates metabolic reprogramming that favours glucose utilization. We wanted to explore whether this influence of VEGFB on cardiac metabolism involves regulation of LPL activity with consequent effects on lipotoxicity and insulin signalling. The transcriptomes of rats with and without cardiomyocyte-specific overexpression of human VEGFB were compared by using RNA-sequencing. Isolated perfused hearts or cardiomyocytes incubated with heparin were used to enable measurement of LPL activity. Untargeted metabolomic analysis was performed for quantification of cardiac lipid metabolites. Cardiac insulin sensitivity was evaluated using fast-acting insulin. Isolated heart and cardiomyocytes were used to determine transgene-encoded VEGFB isoform secretion patterns and mitochondrial oxidative capacity using high-resolution respirometry and extracellular flux analysis. In vitro, primary transgenic cardiomyocytes incubated overnight and thus exposed to abundantly secreted VEGFB isoforms in the absence of any in vivo confounding regulators of cardiac metabolism demonstrated higher basal oxygen consumption. In the whole heart, VEGFB overexpression induced an angiogenic response that was accompanied by limited cardiac LPL activity through multiple mechanisms. This was associated with a lowered accumulation of lipid intermediates, diacylglycerols and lysophosphatidylcholine, that are known to influence insulin action. In response to exogenous insulin, transgenic hearts demonstrated increased insulin sensitivity. In conclusion, the interrogation of VEGFB function on cardiac metabolism uncovered an intriguing and previously unappreciated effect to lower LPL activity and prevent lipid metabolite accumulation to improve insulin action. VEGFB could be a potential cardioprotective therapy to treat metabolic disorders, for example diabetes.

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2H and 13C metabolic flux analysis elucidates in vivo thermodynamics of the ED pathway in Zymomonas mobilis

Metabolic Engineering ◽

10.1016/j.ymben.2019.05.006 ◽

2019 ◽

Vol 54 ◽

pp. 301-316 ◽

Cited By ~ 18

Author(s):

Tyler B. Jacobson ◽

Paul A. Adamczyk ◽

David M. Stevenson ◽

Matthew Regner ◽

John Ralph ◽

...

Keyword(s):

Metabolic Flux Analysis ◽

Zymomonas Mobilis ◽

Metabolic Flux ◽

Flux Analysis ◽

13C Metabolic Flux Analysis

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4-Octyl itaconate inhibits aerobic glycolysis by targeting GAPDH to exert anti-inflammatory effects

Nature Communications ◽

10.1038/s41467-019-13078-5 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 25

Author(s):

Shan-Ting Liao ◽

Chao Han ◽

Ding-Qiao Xu ◽

Xiao-Wei Fu ◽

Jun-Song Wang ◽

...

Keyword(s):

Aerobic Glycolysis ◽

Glycolytic Enzyme ◽

Cysteine Residue ◽

Glycolytic Flux ◽

Flux Analysis ◽

Activated Macrophages ◽

A Cell ◽

Anti Inflammatory ◽

Heptelidic Acid

Abstract Activated macrophages switch from oxidative phosphorylation to aerobic glycolysis, similar to the Warburg effect, presenting a potential therapeutic target in inflammatory disease. The endogenous metabolite itaconate has been reported to regulate macrophage function, but its precise mechanism is not clear. Here, we show that 4-octyl itaconate (4-OI, a cell-permeable itaconate derivative) directly alkylates cysteine residue 22 on the glycolytic enzyme GAPDH and decreases its enzyme activity. Glycolytic flux analysis by U13C glucose tracing provides evidence that 4-OI blocks glycolytic flux at GAPDH. 4-OI thereby downregulates aerobic glycolysis in activated macrophages, which is required for its anti-inflammatory effects. The anti-inflammatory effects of 4-OI are replicated by heptelidic acid, 2-DG and reversed by increasing wild-type (but not C22A mutant) GAPDH expression. 4-OI protects against lipopolysaccharide-induced lethality in vivo and inhibits cytokine release. These findings show that 4-OI has anti-inflammatory effects by targeting GAPDH to decrease aerobic glycolysis in macrophages.

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