Minute‐Scale Repeated Metabolic Probing of Living Cells Enabled by Rapid Parahydrogen‐Based Hyperpolarization
Abstract
ABSTRACT Hyperpolarized (HP) 13 C nuclear magnetic resonance (NMR) spectroscopy enables real‐time observation of metabolic fluxes but is typically limited to single‐shot measurements due to complex preparation procedures and low experimental throughput. Here, we established a rapid and experimentally accessible workflow for temporally controlled HP measurements in living cells. Using SABRE‐SHEATH at 0.4 µT, we achieved 7.7% ± 0.2% 13 C polarization of 50 mM [1‐ 13 C]pyruvate within 60 s. Simple 1:50 dilution with phosphate‐buffered D 2 O yielded cell‐compatible solutions that retained 5.3% ± 0.4% polarization of 1.3 mM pyruvate without multi‐step purification. Combined with a simplified agarose bead immobilization approach, this enabled four injections of HP pyruvate into the same HeLa cell population within 7 min. Under rapid (≈2 min) reinjection intervals, the pyruvate‐to‐lactate conversion progressively declined, whereas stable metabolic conversion was maintained at 20 min intervals with intermittent cell medium perfusion. These findings demonstrate that rapid repeated substrate delivery can transiently exhaust cellular metabolic conversion capacity. This experimentally accessible operating mode enables the study of short‐term metabolic dynamics that remain obscured in conventional single‐shot HP‐NMR or long timescale thermally polarized NMR measurements.
Article Details
Authors (18)
Philipp R. Groß
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany
Stefan Petersen
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany
Zirun Wang
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany
Behnam Shamshiri
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany
Henri de Maissin
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany
Sebastian Lucas
NVision Imaging Technologies GmbH Ulm Germany
Oliver Gorka
Faculty of Medicine, University of Freiburg
Lisa Heß
Institute of Molecular Medicine and Cell Research Faculty of Medicine University of Freiburg Freiburg Germany
Irene Marco‐Rius
Institute For Bioengineering of Catalonia (IBEC) Barcelona Institute of Science and Technology (BIST) Barcelona Spain
Lluís Mangas‐Florencio
Institute For Bioengineering of Catalonia (IBEC) Barcelona Institute of Science and Technology (BIST) Barcelona Spain
André F. Martins
Department of Preclinical Imaging and Radiopharmacy Werner Siemens Imaging Center Eberhard Karls University Tübingen Tübingen Germany
Philipp Boehm‐Sturm
Charité 3R – Replace, Reduce, Refine Charité – Universitätsmedizin Berlin, Corporate Member of Freie Universität Berlin and Humboldt‐Universität zu Berlin Berlin Germany
Maxim Zaitsev
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany
Robert Zeiser
Jan‐Bernd Hövener
Sektion Biomedizinsche Bildgebung Molecular Imaging North Competence Center MOINCC Klinik Für Radiologie und Neuroradiologie University Hospital Schleswig‐Holstein Kiel University Kiel Germany
Thomas Reinheckel
German Cancer Consortium (DKTK), Partner Site Freiburg a Partnership Between DKFZ and University Medical Center Freiburg Heidelberg Germany
Olaf Gross
Faculty of Medicine, University of Freiburg
Andreas B. Schmidt
Division of Medical Physics Department of Radiology Medical Center and Faculty of Medicine Freiburg Germany