Harrieth Wagner

Human VEGF gene expression in skeletal muscle: effect of acute normoxic and hypoxic exercise.

Vascular endothelial growth factor (VEGF) is involved in extracellular matrix changes and endothelial cell proliferation, both of which are precursors to new capillary growth. Angiogenesis is a vital adaptation to exercise training, and the exercise-induced reduction in intracellular[Formula: see text] has been proposed as a stimulus for this process. Thus we studied muscle cell[Formula: see text] [myoglobin[Formula: see text]([Formula: see text])] during exercise in normoxia and in hypoxia (12% O2) and studied the mRNA levels of VEGF in six untrained subjects after a single bout of exercise by quantitative Northern analysis. Single-leg knee extension provided the acute exercise stimulus: a maximal test followed by 30 min at 50% of the peak work rate achieved in this graded test. Because peak work rate was not affected by hypoxia, the absolute and relative work rates were identical in hypoxia and normoxia. Three pericutaneous needle biopsies were collected from the vastus lateralis muscle, one at rest and then the others at 1 h after exercise in normoxia or hypoxia. At rest (control), VEGF mRNA levels were very low (0.38 ± 0.04 VEGF/18S). After exercise in normoxia or hypoxia, VEGF mRNA levels were much greater (16.9 ± 6.7 or 7.1 ± 1.8 VEGF/18S, respectively). In contrast, there was no measurable basic fibroblast growth factor mRNA response to exercise at this 1-h postexercise time point. Magnetic resonance spectroscopy of myoglobin confirmed a reduction in[Formula: see text] in hypoxia (3.8 ± 0.3 mmHg) compared with normoxia (7.2 ± 0.6 mmHg) but failed to reveal a relationship between [Formula: see text] during exercise and VEGF expression. This VEGF mRNA increase in response to acute exercise supports the concept that VEGF is involved in exercise-induced skeletal muscle angiogenesis but questions the importance of a reduced cellular [Formula: see text]as a stimulus for this response.Vascular endothelial growth factor (VEGF) is involved in extracellular matrix changes and endothelial cell proliferation, both of which are precursors to new capillary growth. Angiogenesis is a vital adaptation to exercise training, and the exercise-induced reduction in intracellular PO2 has been proposed as a stimulus for this process. Thus we studied muscle cell PO2 [myoglobin PO2 (MbPO2)] during exercise in normoxia and in hypoxia (12% O2) and studied the mRNA levels of VEGF in six untrained subjects after a single bout of exercise by quantitative Northern analysis. Single-leg knee extension provided the acute exercise stimulus: a maximal test followed by 30 min at 50% of the peak work rate achieved in this graded test. Because peak work rate was not affected by hypoxia, the absolute and relative work rates were identical in hypoxia and normoxia. Three pericutaneous needle biopsies were collected from the vastus lateralis muscle, one at rest and then the others at 1 h after exercise in normoxia or hypoxia. At rest (control), VEGF mRNA levels were very low (0.38 +/- 0.04 VEGF/18S). After exercise in normoxia or hypoxia, VEGF mRNA levels were much greater (16.9 +/- 6.7 or 7.1 +/- 1.8 VEGF/18S, respectively). In contrast, there was no measurable basic fibroblast growth factor mRNA response to exercise at this 1-h postexercise time point. Magnetic resonance spectroscopy of myoglobin confirmed a reduction in MbPO2 in hypoxia (3.8 +/- 0.3 mmHg) compared with normoxia (7.2 +/- 0.6 mmHg) but failed to reveal a relationship between MbPO2 during exercise and VEGF expression. This VEGF mRNA increase in response to acute exercise supports the concept that VEGF is involved in exercise-induced skeletal muscle angiogenesis but questions the importance of a reduced cellular PO2 as a stimulus for this response.

Pulmonary extravascular fluid accumulation in recreational climbers: a prospective study

BACKGROUND High altitude pulmonary oedema (HAPE) that is severe enough to require urgent medical care is infrequent. We hypothesised that subclinical HAPE is far more frequent than suspected during even modest climbs of average effort. METHODS We assessed 262 consecutive climbers of Monte Rosa (4559 m), before ascent and about 24 h later on the summit 1 h after arriving, by clinical examination, electrocardiography, oximetry, spirometry, carbon monoxide transfer, and closing volume. A chest radiograph was taken at altitude. FINDINGS Only one climber was evacuated for HAPE, but 40 (15%) of 262 climbers had chest rales or interstitial oedema on radiograph after ascent. Of 37 of these climbers, 34 (92%) showed increased closing volume. Of the 197 climbers without oedema, 146 (74%) had an increase in closing volume at altitude. With no change in vital capacity, forced expiratory volume in 1 s and forced expiratory flow at 25-75% of forced vital capacity increased slightly at altitude, without evidence of oedema. If we assume that an increased closing volume at altitude indicates increased pulmonary extravascular fluid, our data suggest that three of every four healthy, recreational climbers have mild subclinical HAPE shortly after a modest climb. INTERPRETATION The risk of HAPE might not be confined to a small group of genetically susceptible people, but likely exists for most climbers if the rate of ascent and degree of physical effort are great enough, especially if lung size is normal or low.

Intercostal muscle blood flow limitation in athletes during maximal exercise

We investigated whether, during maximal exercise, intercostal muscle blood flow is as high as during resting hyperpnoea at the same work of breathing. We hypothesized that during exercise, intercostal muscle blood flow would be limited by competition from the locomotor muscles. Intercostal (probe over the 7th intercostal space) and vastus lateralis muscle perfusion were measured simultaneously in ten trained cyclists by near‐infrared spectroscopy using indocyanine green dye. Measurements were made at several exercise intensities up to maximal (WRmax) and subsequently during resting isocapnic hyperpnoea at minute ventilation levels up to those at WRmax. During resting hyperpnoea, intercostal muscle blood flow increased linearly with the work of breathing (R2= 0.94) to 73.0 ± 8.8 ml min−1 (100 g)−1 at the ventilation seen at WRmax (work of breathing ∼550–600 J min−1), but during exercise it peaked at 80% WRmax (53.4 ± 10.3 ml min−1 (100 g)−1), significantly falling to 24.7 ± 5.3 ml min−1 (100 g)−1 at WRmax. At maximal ventilation intercostal muscle vascular conductance was significantly lower during exercise (0.22 ± 0.05 ml min−1 (100 g)−1 mmHg−1) compared to isocapnic hyperpnoea (0.77 ± 0.13 ml min−1 (100 g)−1 mmHg−1). During exercise, both cardiac output and vastus lateralis muscle blood flow also plateaued at about 80% WRmax (the latter at 95.4 ± 11.8 ml min−1 (100 g)−1). In conclusion, during exercise above 80% WRmax in trained subjects, intercostal muscle blood flow and vascular conductance are less than during resting hyperpnoea at the same minute ventilation. This suggests that the circulatory system is unable to meet the demands of both locomotor and intercostal muscles during heavy exercise, requiring greater O2 extraction and likely contributing to respiratory muscle fatigue.

HUMAN RESPIRATORY MUSCLE BLOOD FLOW MEASURED BY NEAR-INFRARED SPECTROSCOPY AND INDOCYANINE GREEN

Measurement of respiratory muscle blood flow (RMBF) in humans has important implications for understanding patterns of blood flow distribution during exercise in healthy individuals and those with chronic disease. Previous studies examining RMBF in humans have required invasive methods on anesthetized subjects. To assess RMBF in awake subjects, we applied an indicator-dilution method using near-infrared spectroscopy (NIRS) and the light-absorbing tracer indocyanine green dye (ICG). NIRS optodes were placed on the left seventh intercostal space at the apposition of the costal diaphragm and on an inactive control muscle (vastus lateralis). The primary respiratory muscles within view of the NIRS optodes include the internal and external intercostals. Intravenous bolus injection of ICG allowed for cardiac output (by the conventional dye-dilution method with arterial sampling), RMBF, and vastus lateralis blood flow to be quantified simultaneously. Esophageal and gastric pressures were also measured to calculate the work of breathing and transdiaphragmatic pressure. Measurements were obtained in five conscious humans during both resting breathing and three separate 5-min bouts of constant isocapnic hyperpnea at 27.1 +/- 3.2, 56.0 +/- 6.1, and 75.9 +/- 5.7% of maximum minute ventilation as determined on a previous maximal exercise test. RMBF progressively increased (9.9 +/- 0.6, 14.8 +/- 2.7, 29.9 +/- 5.8, and 50.1 +/- 12.5 ml 100 ml(-1) min(-1), respectively) with increasing levels of ventilation while blood flow to the inactive control muscle remained constant (10.4 +/- 1.4, 8.7 +/- 0.7, 12.9 +/- 1.7, and 12.2 +/- 1.8 ml 100 ml(-1) min(-1), respectively). As ventilation rose, RMBF was closely and significantly correlated with 1) cardiac output (r = 0.994, P = 0.006), 2) the work of breathing (r = 0.995, P = 0.005), and 3) transdiaphragmatic pressure (r = 0.998, P = 0.002). These data suggest that the NIRS-ICG technique provides a feasible and sensitive index of RMBF at different levels of ventilation in humans.

HIF and VEGF relationships in response to hypoxia and sciatic nerve stimulation in rat gastrocnemius

To determine if hypoxia-inducible factor-1 (HIF-1) may regulate skeletal muscle vascular endothelial growth factor (VEGF) expression in response to exercise or hypoxia, rats underwent 1h sciatic nerve electrical stimulation (ES), hypoxic exposure (H) or combined stimuli. HIF-1alpha protein levels increased six-fold with maximal (8V) ES with or without H. Similar HIF-1alpha increases occurred with sub-maximal (6V and 4V) ES plus H, but not in sub-maximal ES or H alone. VEGF mRNA and protein levels increased three-fold in sub-maximal ES or H alone, six-fold in sub-maximal ES plus H, 6.3-fold with maximal ES, and 6.5-fold after maximal ES plus H. These data suggest: (1) intracellular hypoxia during normoxic exercise may exceed that during 8% oxygen breathing at rest and is more effective in stimulating HIF-1alpha; (2) HIF-1 may be an important regulator of exercise-induced VEGF transcription; and (3) breathing 8% O(2) does not alter HIF-1alpha expression in skeletal muscle, implying that exercise-generated signals contribute to the regulation of HIF-1alpha and/or VEGF.

Frontal cerebral cortex blood flow, oxygen delivery and oxygenation during normoxic and hypoxic exercise in athletes

Non‐technical summary  Exercise capacity is limited at high altitude where hypoxia (i.e. decreased amount of inspired oxygen resulting in decreased oxygen in the blood) is present, but it is unknown whether a reduction in the oxygen delivered to the brain constitutes the signal to the brain to prematurely terminate exercise. We show that during hypoxic exercise equivalent to exercise at ∼4000 m above sea‐level, the oxygen delivered to the brain during intense exercise is ∼60% less than that delivered to the brain at comparable exercise intensity at sea‐level. These results show that reduction in the oxygen delivered to the brain could constitute the signal to limit maximal exercise capacity in hypoxia, and help us understand better why exercise capacity is limited at high altitude. Moreover, a plausible mechanism of exercise limitation in patients who present decreased oxygen in the blood during exercise due to pulmonary and/or cardiac disease is revealed.

Does gender affect human pulmonary gas exchange during exercise

Women may experience greater pulmonary gas exchange impairment during exercise than men. To test this we used the multiple inert gas elimination technique to study eight women and seven men matched for age, height and V̇O2 max (∼48 ml kg−1 min−1) during normoxic and hypoxic (inspired PO2= 95 Torr) cycle exercise. Resting lung function was similar between the sexes, except for a lower carbon monoxide diffusing capacity (DLCO) in women (P < 0.05). Arterial PO2,PCO2 and alveolar–arterial O2 difference (A−aDO2) were not significantly different in men and women. Despite a lower diffusing capacity for O2 (DLO2) in women, the ratio DLO2/βQ̇ (which estimates pulmonary end‐capillary diffusion equilibrium) was similar between men and women and estimates of diffusion limitation during hypoxic exercise were not different between the sexes. Ventilation–perfusion inequality (described by the second moment of the perfusion distribution, logSD) increased during both normoxic and hypoxic exercise. Surprisingly, logSD values were slightly lower for women under all conditions (P < 0.05), but this did not significantly affect gas exchange. These data indicate that these active women, despite a lower DLCO and DLO2, do not experience greater exercise‐induced abnormalities in gas exchange than men matched for age, height, aerobic capacity and lung size. Possibly fitness level and lung size are more important in determining whether or not pulmonary gas exchange impairment occurs during exercise than sex per se.

The contribution of intrapulmonary shunts to the alveolar‐to‐arterial oxygen difference during exercise is very small

Exercise is well known to cause arterial to fall and the alveolar–arterial difference (Aa ) to increase. Until recently, the physiological basis for this was considered to be mostly ventilation/perfusion / inequality and alveolar–capillary diffusion limitation. Recently, arterio‐venous shunting through dilated pulmonary blood vessels has been proposed to explain a significant part of the Aa during exercise. To test this hypothesis we determined venous admixture during 5 min of near‐maximal, constant‐load, exercise in hypoxia (in inspired O2 fraction, , 0.13), normoxia (, 0.21) and hyperoxia (, 1.0) undertaken in balanced order on the same day in seven fit cyclists (, 61.3 ± 2.4 ml kg−1 min−1; mean ±s.e.m.). Venous admixture reflects three causes of hypoxaemia combined: true shunt, diffusion limitation and / inequality. In hypoxia, venous admixture was 22.8 ± 2.5% of the cardiac output; in normoxia it was 3.5 ± 0.5%; in hyperoxia it was 0.5 ± 0.2%. Since only true shunt accounts for venous admixture while breathing 100% O2, the present study suggests that shunt accounts for only a very small portion of the observed venous admixture, Aa and hypoxaemia during heavy exercise.

Contribution of respiratory muscle blood flow to exercise‐induced diaphragmatic fatigue in trained cyclists

We investigated whether the greater degree of exercise‐induced diaphragmatic fatigue previously reported in highly trained athletes in hypoxia (compared with normoxia) could have a contribution from limited respiratory muscle blood flow. Seven trained cyclists completed three constant load 5 min exercise tests at inspired O2 fractions () of 0.13, 0.21 and 1.00 in balanced order. Work rates were selected to produce the same tidal volume, breathing frequency and respiratory muscle load at each (63 ± 1, 78 ± 1 and 87 ± 1% of normoxic maximal work rate, respectively). Intercostals and quadriceps muscle blood flow (IMBF and QMBF, respectively) were measured by near‐infrared spectroscopy over the left 7th intercostal space and the left vastus lateralis muscle, respectively, using indocyanine green dye. The mean pressure time product of the diaphragm and the work of breathing did not differ across the three exercise tests. After hypoxic exercise, twitch transdiaphragmatic pressure fell by 33.3 ± 4.8%, significantly (P < 0.05) more than after both normoxic (25.6 ± 3.5% reduction) and hyperoxic (26.6 ± 3.3% reduction) exercise, confirming greater fatigue in hypoxia. Despite lower leg power output in hypoxia, neither cardiac output nor QMBF (27.6 ± 1.2 l min−1 and 100.4 ± 8.7 ml (100 ml)−1 min−1, respectively) were significantly different compared with normoxia (28.4 ± 1.9 l min−1 and 94.4 ± 5.2 ml (100 ml)−1 min−1, respectively) and hyperoxia (27.8 ± 1.6 l min−1 and 95.1 ± 7.8 ml (100 ml)−1 min−1, respectively). Neither IMBF was different across hypoxia, normoxia and hyperoxia (53.6 ± 8.5, 49.9 ± 5.9 and 52.9 ± 5.9 ml (100 ml)−1 min−1, respectively). We conclude that when respiratory muscle energy requirement is not different between normoxia and hypoxia, diaphragmatic fatigue is greater in hypoxia as intercostal muscle blood flow is not increased (compared with normoxia) to compensate for the reduction in , thus further compromising O2 supply to the respiratory muscles.

Peripheral oxygen transport and utilization in rats following continued selective breeding for endurance running capacity

Untrained rats selectively bred for either high (HCR) or low (LCR) treadmill running capacity previously demonstrated divergent physiological traits as early as the seventh generation (G7). We asked whether continued selective breeding to generation 15 (G15) would further increase the divergence in skeletal muscle capillarity, morphometry, and oxidative capacity seen previously at G7. At G15, mean body weight was significantly lower (P < 0.001) in the HCR rats (n = 11; 194 +/- 3 g) than in LCR (n = 12; 259 +/- 9 g) while relative medial gastrocnemius muscle mass was not different (0.23 +/- 0.01 vs. 0.22 +/- 0.01% total body weight). Normoxic (Fi(O(2)) = 0.21) Vo(2max) was 50% greater (P < 0.001) in HCR despite the lower absolute muscle mass, and skeletal muscle O(2) conductance (measured in hypoxia; Fi(O(2)) = 0.10) was 49% higher in HCR (P < 0.001). Muscle oxidative enzyme activities were significantly higher in HCR (citrate synthase: 16.4 +/- 0.4 vs. 14.0 +/- 0.6; beta-hydroxyacyl-CoA dehydrogenase: 5.2 +/- 0.2 vs. 4.2 +/- 0.2 mmol.kg(-1).min(-1)). HCR rats had approximately 36% more total muscle fibers and also 36% more capillaries in the medial gastrocnemius. Because average muscle fiber area was 35% smaller, capillary density was 36% higher in HCR, but capillary-to-fiber ratio was the same. Compared with G7, G15 HCR animals showed 38% greater total fiber number with an additional 25% decrease in mean fiber area. These data suggest that many of the skeletal muscle structural and functional adaptations enabling greater O(2) utilization in HCR at G7 continue to progress following additional selective breeding for endurance capacity. However, the largest changes at G15 relate to O(2) delivery to skeletal muscle and not to the capacity of skeletal muscle to use O(2).