Assessing the transport rate of hyperpolarized pyruvate and lactate from the intra- to the extracellular space.
The use of [1-(13) C]pyruvate hyperpolarized by means of dynamic nuclear polarization provides a direct way to track the metabolic transformations of this metabolite in vivo and in cell cultures. The identification of the intra- and extracellular contributions to the (13) C NMR resonances is not straightforward. In order to obtain information about the rate of pyruvate and lactate transport through the cellular membrane, we set up a method that relies on the sudden 'quenching' of the extracellular metabolites' signal. The paramagnetic Gd-tetraazacyclododecane triacetic acid (Gd-DO3A) complex was used to dramatically decrease the longitudinal relaxation time constants of the (13) C-carboxylate resonances of both pyruvate and lactate. When Gd-DO3A was added to an MCF-7 cellular culture, which had previously received a dose of hyperpolarized [1-(13) C]pyruvate, the contributions of the extracellular pyruvate and lactate signals were deleted. From the analysis of the decay curves of the (13) C-carboxylate resonances of pyruvate and lactate it was possible to extract information about the exchange rate of the two metabolites across the cellular membrane. In particular, it was found that, in the reported experimental conditions, the lactate transport from the intra- to the extracellular space is not much lower than the rate of lactate formation. The method reported herein is non-destructive and it could be translated to in vivo studies. It opens a route for the use of hyperpolarized pyruvate to assess altered activity of carboxylate transporter proteins that may occur in pathological conditions. Copyright © 2016 John Wiley & Sons, Ltd.
Department of Molecular Biotechnology and Health Sciences, University of Torino, Turin, Italy.
Francesca
F
Reineri
Department of Molecular Biotechnology and Health Sciences, University of Torino, Turin, Italy.
Valeria
V
Daniele
Department of Molecular Biotechnology and Health Sciences, University of Torino, Turin, Italy.
Eleonora
E
Cavallari
Department of Molecular Biotechnology and Health Sciences, University of Torino, Turin, Italy.
Silvio
S
Aime
D001693:Biological Transport, Active / Q000502:physiology*; D066241:Carbon-13 Magnetic Resonance Spectroscopy; D002462:Cell Membrane / Q000378:metabolism*; D003287:Contrast Media / Q000493:pharmacokinetics; D005110:Extracellular Space / Q000378:metabolism; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism; D019344:Lactic Acid / Q000378:metabolism*; D061986:MCF-7 Cells; D008657:Metabolic Clearance Rate; D009942:Organometallic Compounds / Q000493:pharmacokinetics; D066244:Proton Magnetic Resonance Spectroscopy; D019289:Pyruvic Acid / Q000378:metabolism*; D019275:Radiopharmaceuticals / Q000493:pharmacokinetics
D003287:Contrast Media; D009942:Organometallic Compounds; D019275:Radiopharmaceuticals; C090600:gadobutrol; D019344:Lactic Acid; D019289:Pyruvic Acid
Gd(III) complexes; Hyperpolarization; dynamic nuclear polarization; monocarboxylate transporter; pyruvate; relaxometry