Résumé :
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Myostatin negatively regulates muscle growth. Deletion of the myostatin gene results in spectacular increase in muscle mass, and opened the path to therapeutic approaches. Yet improvement in muscle strength does not necessarily match the observed increase in muscle mass. If function is to be preserved in hypertrophic muscle, adequate oxygen supply and substrate utilization should also be maintained. Multi-parametric-functional (mpf) NMR, initially developed for humans, explores these aspects in vivo and non-invasively but remains challenging in mouse because of the small size of the animal. We proposed a NMR-gated approach for mouse. The set-up, including a repeated exercise protocol was developed to simultaneously assess muscle perfusion, BOLD, phosphorus metabolism, and isometric force in response to electrical stimulation. It was applied to investigate myostatin knock-out mice (mstn-/-). Mstn-/-mice [8] (n=6) were compared to wild-type (WT, n=10). Anesthetized animals were installed inside a Bruker Biospec 4T NMR system with custom-designed coils (actively decoupled 1H wholebody transmission and surface reception, 31P leg-volumetric acquisition). Contractions were induced through subcutaneous silver electrodes, and force measurement was collected through a custombuilt ergometer. Pulsed arterial spin labeled RARE images (PASL-NMRI) using the SATIR variant [9] acquired every 10s were interleaved with 4 successive 31P-NMR spectra (TR= 2.5s), using the Bruker MultiScanControl tool. Acquisitions were triggered to the electro-stimulation, to allow perfusion measurement even during exercise. Mice were subjected to an "exercise (2 min) recovery (10 min)" protocol, repeated 12 times, with the aim of summing 31P data across exercises, in order to compensate for intrinsically low signal to noise ratio (S/N), but to fit phosphocreatine resynthesis (mono-exponential constant PCr) to high temporal resolution data. First; we evaluated our approach. Owing to high time resolution and improved S/N obtained after gated summation of successive exercises, we could measure PCr for each animal with an averaged CV of 15%. The results of inter-animal summation showed that PCr reached a steady-state after the first exercise at 88 +/-10s. Steady state was also reached after the 2nd bout of exercise, for all variables describing force, perfusion, and BOLD. Using this gated approach, we compared results from WT and mstn-/-mice. The force developed by mstn-/-was not different from WT when normalized for the 36% increase in muscle cross sectional area (2386+119 vs 2456+145 mN.s/cm2 resp). Depletion of PCr during exercise was identical (mstn-/-63%+11 vs WT 58%+13), as was endexercise pH. Maximal perfusion at exercise was also the same, but time courses of recovery of mstn-/-were radically different. Hyperemic perfusion was extremely prolonged, while Pcr recovery was also significantly delayed (P<.015). The present work describes preliminary findings in mstn-/-animals, showing that although muscle of these mice develops force matched to its mass, recovery from exercise is much slower, as compared to the WT, in terms of oxidative phosphorylation, while oxygen supply to the muscle is also prolonged. Adapting the mpf-NMR approach to mice makes it possible to non invasively investigate the interplay of muscle oxygen supply and energetics in vivo after exercise to improve our understanding of function in these models.
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