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Metabolism of [3-13C]pyruvate and [3-13C]propionate in normal and ischaemic rat heart in vivo: 1H- and 13C-NMR studies.

ABSTRACT: The oxidation of [3-13C]pyruvate and [3-13C]propionate was studied in vivo in infused rats. The infused [3-13C]pyruvate was quickly converted to [3-13C]lactate in the blood, and the [3-13C]lactate formed was well metabolized in both normoxic and ischaemic hearts. Large differences (200-600%) in the 13C enrichment of alanine (C-3) and acetyl-CoA (C-2) compared with lactate (C-3) were found in both normoxic and ischaemic hearts, suggesting that the extracellular [3-13C]lactate preferentially entered a region of the cytoplasm which specifically transfers the labelled pyruvate (formed from [3-13C]lactate) to the mitochondria. The highly enriched mitochondrial pyruvate gave high enrichment in alanine and acetyl-CoA, which was detected by 1H- and 13C-NMR spectroscopy. Ischaemia increased 13C incorporation into the main cytoplasmic lactate pool and decreased 13C incorporation into citric acid cycle intermediates, mainly decreasing the pyruvate anaplerosis. Isoprenaline-induced ischaemia of the heart caused only a slight decrease in pyruvate oxidation. In contrast to the decreased anaplerosis of pyruvate, the anaplerosis of propionate (and propionyl-carnitine) increased significantly in ischaemic hearts, which may contribute to the protective effect of propionyl-carnitine seen in ischaemia. In addition, we found that [3-13C]propionate preferentially labelled aspartate C-3 in rat heart, suggesting incomplete randomization of label in the succinyl-CoA-malate span of the citric acid cycle. These data show that proton observed 13C edited spectroscopic methods, i.e. heteronuclear spin-echo and the one-dimensional heteronuclear multiple quantum coherence sequence, can be successfully used to study heart metabolism in vivo.

SUBMITTER: Sumegi B

PROVIDER: S-EPMC1136229 | biostudies-other | 1995 Nov

REPOSITORIES: biostudies-other

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Project description:Metabolic reprogramming facilitates cancer cell growth, so quantitative metabolic flux measurements could produce useful biomarkers. However, current methods to analyze flux in vivo provide either a steady-state overview of relative activities (infusion of (13)C and analysis of extracted metabolites) or a dynamic view of a few reactions (hyperpolarized (13)C spectroscopy). Moreover, although hyperpolarization has successfully quantified pyruvate-lactate exchanges, its ability to assess mitochondrial pyruvate metabolism is unproven in cancer. Here, we combined (13)C hyperpolarization and isotopomer analysis to quantify multiple fates of pyruvate simultaneously. Two cancer cell lines with divergent pyruvate metabolism were incubated with thermally polarized [3-(13)C]pyruvate for several hours, then briefly exposed to hyperpolarized [1-(13)C]pyruvate during acquisition of NMR spectra using selective excitation to maximize detection of H[(13)C]O3(-) and [1-(13)C]lactate. Metabolites were then extracted and subjected to isotopomer analysis to determine relative rates of pathways involving [3-(13)C]pyruvate. Quantitation of hyperpolarized H[(13)C]O3(-) provided a single definitive metabolic rate, which was then used to convert relative rates derived from isotopomer analysis into quantitative fluxes. This revealed that H[(13)C]O3(-) appearance reflects activity of pyruvate dehydrogenase rather than pyruvate carboxylation followed by subsequent decarboxylation reactions. Glucose substantially altered [1-(13)C]pyruvate metabolism, enhancing exchanges with [1-(13)C]lactate and suppressing H[(13)C]O3(-) formation. Furthermore, inhibiting Akt, an oncogenic kinase that stimulates glycolysis, reversed these effects, indicating that metabolism of pyruvate by both LDH and pyruvate dehydrogenase is subject to the acute effects of oncogenic signaling on glycolysis. The data suggest that combining (13)C isotopomer analyses and dynamic hyperpolarized (13)C spectroscopy may enable quantitative flux measurements in living tumors.

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Metabolism of [3-13C]pyruvate and [3-13C]propionate in normal and ischaemic rat heart in vivo: 1H- and 13C-NMR studies.

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