Hyperpolarized 13C Magnetic Resonance and Its Use in Metabolic Assessment of Cultured Cells and Perfused Organs

Abstract

Diseased tissue is often characterized by abnormalities in intermediary metabolism. Observing these alterations in situ may lead to an improved understanding of pathological processes and novel ways to monitor these processes non-invasively in human patients. Although 13C is a stable isotope safe for use in animal models of disease as well as human subjects, its utility as a metabolic tracer has largely been limited to ex vivo analyses employing analytical techniques like mass spectrometry or nuclear magnetic resonance spectroscopy. Neither of these techniques is suitable for non-invasive metabolic monitoring, and the low abundance and poor gyromagnetic ratio of conventional 13C make it a poor nucleus for imaging. However, the recent advent of hyperpolarization methods, particularly dynamic nuclear polarization (DNP), make it possible to enhance the spin polarization state of 13C by many orders of magnitude, resulting in a temporary amplification of the signal sufficient for monitoring kinetics of enzyme-catalyzed reactions in living tissue through magnetic resonance spectroscopy or magnetic resonance imaging. Here we review DNP techniques to monitor metabolism in cultured cells, perfused hearts, and perfused livers, focusing on our experiences with hyperpolarized [1-13C]pyruvate. We present detailed approaches to optimize the DNP procedure, streamline biological sample preparation, and maximize detection of specific metabolic activities. We also discuss practical aspects in the choice of metabolic substrates for hyperpolarization studies, and outline some of the current technical and conceptual challenges in the field, including efforts to use hyperpolarization to quantify metabolic rates in vivo.

Construction and 13C hyperpolarization efficiency of a 180 GHz dissolution dynamic nuclear polarization system

“…In this paper, we report on the development and 13 C hyperpolarization efficiency of a homebuilt dynamic nuclear polarization (DNP) system operating at 6.423 T and 1.4 K. The DNP hyperpolarizer system was assembled on a wide-bore superconducting magnet, equipped with a standard continuous-flow cryostat, and a 180 GHz microwave source with 120 mW power output and wide 4 GHz frequency tuning range. At 6.423 T and 1.4 K, solid-state 13 C polarization P levels of 64% and 31% were achieved for 3M [1][2][3][4][5][6][7][8][9][10][11][12][13] C] sodium acetate samples in 1:1 v/v glycerol:water glassing matrix doped with 15 mM trityl OX063 and 40 mM 4-oxo-TEMPO, respectively.…”

“…In this paper, we report on the development and 13 C hyperpolarization efficiency of a homebuilt dynamic nuclear polarization (DNP) system operating at 6.423 T and 1.4 K. The DNP hyperpolarizer system was assembled on a wide-bore superconducting magnet, equipped with a standard continuous-flow cryostat, and a 180 GHz microwave source with 120 mW power output and wide 4 GHz frequency tuning range. At 6.423 T and 1.4 K, solid-state 13 C polarization P levels of 64% and 31% were achieved for 3M [1][2][3][4][5][6][7][8][9][10][11][12][13] C] sodium acetate samples in 1:1 v/v glycerol:water glassing matrix doped with 15 mM trityl OX063 and 40 mM 4-oxo-TEMPO, respectively. Upon dissolution which takes about 15 s to complete, liquid-state 13 C NMR signal enhancements as high as 240,000-fold (P=21%) were recorded in a nearby high resolution 13 C NMR spectrometer at 1 T and 297 K. Considering the relatively lower cost of our homebuilt DNP system and the relative simplicity of its design, the dissolution DNP setup reported here could be feasibly adapted for in vitro or in vivo hyperpolarized 13 C NMR or MRI at least in the pre-clinical setting.…”

“…[1,2] Meanwhile, the magnetic moment of electrons (γ=28,020 MHz/T) dwarfs that of any nuclear spin by a factor of several hundred or thousand-fold. As a result, when cooled at cryogenic temperatures close to 1 K under high magnetic field, the Boltzmann-dictated polarization level of electrons can approach to 100% whereas an ensemble of typical low-γ nuclei such as 13 C spins (γ=10.7 MHz/T) would only have a tiny fraction of a percent thermal polarization even under these conditions. [2,3] Dynamic nuclear polarization (DNP) is a technique, originally used in nuclear scattering and particle physics experiments in the 1960s, that takes advantage of the high Boltzmann electron polarization under those conditions and transfers the high electron spin magnetization or polarization to the target nuclear spins.…”

“…This simple but innovative technique has provided the necessary bridge to take advantage of the superb NMR signal sensitivity by DNP to chemical and biomedical research. In particular, the use of hyperpolarized 13 C biomolecules such as 13 C pyruvate allows NMR and MRI studies that can track the metabolism in cells in vitro and tissues in vivo in real-time with unprecedented sensitivity and superb specificity. [7][8][9][10][11][12][13] In addition to 13 C spins, the liquid-state NMR sensitivity of other low-γ nuclei with relatively long T 1 such as 15 N, [14] 6 Li, [15] 89 Y, [16,17] and 107,109 Ag [18] have also been shown to be greatly enhanced by dissolution DNP.…”

“…In particular, the use of hyperpolarized 13 C biomolecules such as 13 C pyruvate allows NMR and MRI studies that can track the metabolism in cells in vitro and tissues in vivo in real-time with unprecedented sensitivity and superb specificity. [7][8][9][10][11][12][13] In addition to 13 C spins, the liquid-state NMR sensitivity of other low-γ nuclei with relatively long T 1 such as 15 N, [14] 6 Li, [15] 89 Y, [16,17] and 107,109 Ag [18] have also been shown to be greatly enhanced by dissolution DNP. With this capability, hyperpolarized 13 C magnetic resonance in particular offers a promising diagnostic molecular imaging method in which endogenous metabolic probes can be used to track metabolism of normal and diseased tissues such as cancer, without the drawbacks of other imaging methods such as the radioactivity in positron emission tomography (PET) or the large background signal in conventional in vivo proton NMR and MRI.…”