Claire Wary

Human skeletal muscle intracellular oxygenation: the impact of ambient oxygen availability

Intracellular oxygen (O2) availability and the impact of ambient hypoxia have far reaching ramifications in terms of cell signalling and homeostasis; however, in vivo cellular oxygenation has been an elusive variable to assess. Within skeletal muscle the extent to which myoglobin desaturates (deoxy‐Mb) and the extent of this desaturation in relation to O2 availability provide an endogenous probe for intracellular O2 partial pressure (P  iO 2 ). By combining proton nuclear magnetic resonance spectroscopy (1H NMRS) at a high field strength (4 T), assessing a large muscle volume in a highly efficient coil, and extended signal averaging (30 min) we assessed the level of skeletal muscle deoxy‐Mb in 10 healthy men (30 ± 4 years) at rest in both normoxia and hypoxia (10% O2). In normoxia there was an average deoxy‐Mb signal of 9 ± 1%, which, when converted to P  iO 2 using an O2/Mb half‐saturation (P50) of 3.2 mmHg, revealed an P  iO 2 of 34 ± 6 mmHg. In ambient hypoxia the deoxy‐Mb signal rose to 13 ± 3% (P  iO 2 = 23 ± 6 mmHg). However, intersubject variation in the defence of arterial oxygenation (S  aO 2 ) in hypoxia (S  aO 2 range: 86–67%) revealed a significant relationship between the changes in S  aO 2 and P  iO 2(r2= 0.5). These data are the first to document resting intracellular oxygenation in human skeletal muscle, highlighting the relatively high P  iO 2 values that contrast markedly with those previously recorded during exercise (∼2–5 mmHg). Additionally, the impact of ambient hypoxia on P  iO 2 and the relationship between changes in S  aO 2 and P  iO 2 stress the importance of the O2 cascade from air to cell that ultimately effects O2 availability and O2 sensing at the cellular level.

Determination of skeletal muscle perfusion using arterial spin labeling NMRI: validation by comparison with venous occlusion plethysmography.

T1‐based determination of perfusion was performed with the high temporal and spatial resolution that monitoring of exercise physiology requires. As no data were available on the validation of this approach in human muscles, T1‐based NMRI of perfusion was compared to standard strain‐gauge venous occlusion plethysmography performed simultaneously within a 4 T magnet. Two different situations were investigated in 21 healthy young volunteers: 1) a 5‐min ischemia of the leg, or 2) a 2–3 min ischemic exercise consisting of a plantar flexion on an amagnetic ergometer. Leg perfusion was monitored over 5–15 min of the recovery phase, after the air‐cuff arterial occlusion had been released. The interesting features of the sequence were the use of a saturation‐recovery module for the introduction of a T1 modulation and of single‐shot spin echo for imaging. Spatial resolution was 1.7 × 2.0 mm and temporal resolution was 2 s. For data analysis, ROIs were traced on different muscles and perfusion was calculated from the differences in muscle signal intensity in successive images. To allow comparison with the global measurement of perfusion by plethysmography, the T1‐based NMR measurements in exercising muscles were rescaled to the leg cross‐section. The perfusion measurements obtained by plethysmography and NMRI were in close agreement with a correlation coefficient between 0.87 and 0.92. This indicates that pulsed arterial techniques provide determination of muscle perfusion not only with superior spatial and temporal resolution but also with exactitude. Magn Reson Med 46:305–311, 2001.

Muscle glycogenosis due to phosphoglucomutase 1 deficiency.

To the Editor: Muscle glycogen storage diseases are rare inborn diseases caused by errors of metabolism associated with either dynamic, exercise-related symptoms or permanent muscle weakness. The m...

Forelimb Treatment in a Large Cohort of Dystrophic Dogs Supports Delivery of a Recombinant AAV for Exon Skipping in Duchenne Patients

Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disorder caused by mutations in the dystrophin gene, without curative treatment yet available. Our study provides, for the first time, the overall safety profile and therapeutic dose of a recombinant adeno-associated virus vector, serotype 8 (rAAV8) carrying a modified U7snRNA sequence promoting exon skipping to restore a functional in-frame dystrophin transcript, and injected by locoregional transvenous perfusion of the forelimb. Eighteen Golden Retriever Muscular Dystrophy (GRMD) dogs were exposed to increasing doses of GMP-manufactured vector. Treatment was well tolerated in all, and no acute nor delayed adverse effect, including systemic and immune toxicity was detected. There was a dose relationship for the amount of exon skipping with up to 80% of myofibers expressing dystrophin at the highest dose. Similarly, histological, nuclear magnetic resonance pathological indices and strength improvement responded in a dose-dependent manner. The systematic comparison of effects using different independent methods, allowed to define a minimum threshold of dystrophin expressing fibers (>33% for structural measures and >40% for strength) under which there was no clear-cut therapeutic effect. Altogether, these results support the concept of a phase 1/2 trial of locoregional delivery into upper limbs of nonambulatory DMD patients.

Influence of vascular filling and perfusion on BOLD contrast during reactive hyperemia in human skeletal muscle

Mechanisms generating BOLD contrast are complex and depend on parameters that are prone to large variations, in particular in skeletal muscle. Here, we simultaneously measured perfusion by ASL, and BOLD response in the calf muscle of 6 healthy volunteers during post‐ischemic reactive hyperemia. We tested whether the relation between the two was altered for varying degrees of leg vascular replenishment induced by prior positioning of the leg at different heights relative to the heart. We found that the BOLD response depended on perfusion, but also on the degree of repletion of leg blood vessels. We conclude that simultaneous determination of perfusion by ASL is important to identify the mechanisms underlying BOLD contrast in the skeletal muscle. Magn Reson Med, 2006.

Antioxidants and aging: NMR-based evidence of improved skeletal muscle perfusion and energetics

We sought to examine the potential role of oxidative stress on skeletal muscle function with advancing age. Nuclear magnetic resonance (NMR) was employed to simultaneously assess muscle perfusion (arterial spin labeling) and energetics ((31)P NMR spectroscopy) in the lower leg of young (26 + or - 5 yr, n = 6) and older (70 + or - 5 yr, n = 6) healthy volunteers following the consumption of either placebo (PL) or an oral antioxidant (AO) cocktail (vitamins C and E and alpha-lipoic acid), previously documented to decrease plasma free radical concentration. NMR measurements were made during and after 5 min of moderate intensity (approximately 5 W) plantar flexion exercise. AO administration significantly improved end-exercise perfusion (AO, 50 + or - 5, and PL, 43 + or - 4 ml x 100 g(-1) x min(-1)) and postexercise perfusion area under the curve (AO, 1,286 + or - 236, and PL, 866 + or - 144 ml/100 g) in older subjects, whereas AO administration did not alter hemodynamics in the young group. Concomitantly, muscle oxidative capacity (time constant of phosphocreatine recovery, tau) was improved following AO in the older (AO, 43 + or - 1, and PL, 51 + or - 7 s) but not the young (AO, 54 + or - 5, and PL, 48 + or - 7 s) group. These findings support the concept that oxidative stress may be partially responsible for the age-related decline in skeletal muscle perfusion during physical activity and reveal a muscle metabolic reserve capacity in the elderly that is accessible under conditions of improved perfusion.

Functional assessment of skeletal muscle in intact mice lacking myostatin by concurrent NMR imaging and spectroscopy

Inhibiting myostatin (mstn) causes spectacular increase in muscle mass, spurring research for therapeutic approaches against neuromuscular disorders. Yet, possible functional deterioration and compromised force production have been reported in isolated muscle of null mstn(−/−) mice. We analyzed vascular and metabolic response to repeated electro-stimulated exercise in vivo in mstn−/− mice compared with FVB wild-type controls (WT), using interleaved multi-parametric functional nuclear magnetic resonance (NMR) imaging and spectroscopy. At steady-state exercise, specific force of plantar flexion, phosphocreatine consumption measured by phosphorus spectroscopy and maximum perfusion measured by arterial spin-labeled (ASL) NMR imaging were identical in both groups. After exercise, phosphorus spectroscopy revealed reduced oxidative mitochondrial capacity in mstn−/−, whereas early recovery perfusion was identical and oxygen extraction, estimated from the blood oxygen level-dependent (BOLD) contrast, was decreased when compared with WT. Hyperemia was prolonged, indicating specific regulation of the perfusional response in mstn−/− mice. Histology showed an increased proportion of type IIb fibers in hypertrophied muscles, but the distribution of capillary contacts per fiber between oxidative and glycolytic fibers was unaltered in mstn−/− compared with WT. These integrated results formed coherent evidence of a congruous, non-pathologic shift toward a more glycolytic metabolism in this model of mstn−/−.

Abnormal response to cortical activation in early stages of Huntington disease

We wished to identify noninvasive in vivo biomarkers of brain energy deficit in Huntington disease.

Multigap parallel‐plate bracelet resonator frequency determination and applications

A loop made from a length of transmission line with alternate electrical discontinuities (gaps) on the two conductors behaves as a resonator whose resonant frequency is a function of the transmission line parameters as well as its form and the number (n) of gaps. For a given loop diameter, one can thus design a wide range of resonant frequency by choice of n and the characteristic impedance Zc of the line. The analysis is illustrated by a parallel‐plate transmission line resonator in the form of a bracelet. Experimental resonant frequencies obtained from resonators of various dimensions are in good agreement with calculated values. Two resonators were tested on a 3 T NMR imager: A 11 cm diameter 1H surface coil, yielding results comparable to those obtained from a lumped capacitor coil and a 13 cm diameter volume coil to perform in vivo muscle 13C spectroscopy without 1H decoupling.

Quantitative NMRI and NMRS identify augmented disease progression after loss of ambulation in forearms of boys with Duchenne muscular dystrophy.

Quantitative NMRI and 31P NMRS indices are reported in the forearms of 24 patients with Duchenne muscular dystrophy (DMD) (6–18 years, 14 non‐ambulant) amenable to exon 53 skipping therapy and in 12 age‐matched male controls (CONT). Examinations carried out at 3 T comprised multi‐slice 17‐echo measurements of muscle water T2 and heterogeneity, three‐point Dixon imaging of fat fraction in flexor and extensor muscles (FLEX, EXT), and non‐localised spectroscopy of phosphate metabolites. We studied four imaging indices, eight metabolic ratios combining ATP, phosphocreatine, phosphomonoesters and phosphodiesters, the cytosolic inorganic phosphate (Pia) and an alkaline (Pib) pool present in dystrophic muscle, and average pH. All indices differed between DMD and CONT, except for muscle water T2. Measurements were outside the 95th percentile of age‐matched CONT values in over 65% of cases for percentage fat signal (%F), and in 78–100% of cases for all spectroscopic indices. T2 was elevated in one‐third of FLEX measurements, whereas %pixels > 39 ms and T2 heterogeneity were abnormal in one‐half of the examinations. The FLEX muscles had higher fat infiltration and T2 than EXT muscle groups. All indices, except pH, correlated with patient age, although the correlation was negative for T2. However, in non‐ambulant patients, the correlation with years since loss of ambulation was stronger than the correlation with age, and the slope of evolution per year was steeper after loss of ambulation. All indices except Pi/gATP differed between ambulant and non‐ambulant patients; however, T2 and %pixels > 39 ms were highest in ambulant patients, possibly owing to the greater extent of inflammatory processes earlier in the disease. All other indices were worse in non‐ambulant subjects. Quantitative measurements obtained from patients at different disease stages covered a broad range of abnormalities that evolved with the disease, and metabolic indices were up to 10‐fold above normal from the onset, thus establishing a variety of potential markers for future therapy. Copyright