Lung capillary endothelial cells (ECs) are a critical target of oxygen toxicity and play a central role in the pathogenesis of hyperoxic lung injury. To determine mechanisms and time course of EC activation in normobaric hyperoxia, we measured endothelial concentration of reactive oxygen species (ROS) and cytosolic calcium ([Ca(2+)](i)) by in situ imaging of 2',7'-dichlorofluorescein (DCF) and fura 2 fluorescence, respectively, and translocation of the small GTPase Rac1 by immunofluorescence in isolated perfused rat lungs. Endothelial DCF fluorescence and [Ca(2+)](i) increased continuously yet reversibly during a 90-min interval of hyperoxic ventilation with 70% O(2), demonstrating progressive ROS generation and second messenger signaling. ROS formation increased exponentially with higher O(2) concentrations. ROS and [Ca(2+)](i) responses were blocked by the mitochondrial complex I inhibitor rotenone, whereas inhibitors of NAD(P)H oxidase and the intracellular Ca(2+) chelator BAPTA predominantly attenuated the late phase of the hyperoxia-induced DCF fluorescence increase after > 30 min. Rac1 translocation in lung capillary ECs was barely detectable at normoxia but was prominent after 60 min of hyperoxia and could be blocked by rotenone and BAPTA. We conclude that hyperoxia induces ROS formation in lung capillary ECs, which initially originates from the mitochondrial electron transport chain but subsequently involves activation of NAD(P)H oxidase by endothelial [Ca(2+)](i) signaling and Rac1 activation. Our findings demonstrate rapid activation of ECs by hyperoxia in situ and identify mechanisms that may be relevant in the initiation of hyperoxic lung injury.

These data suggest that CD95/Apo1/Fas is directly involved in cell death after myocardial ischemia. The CD95 system might thus represent a novel target for therapeutic prevention of postischemic cell death in the heart.

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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 (WR max ) and subsequently during resting isocapnic hyperpnoea at minute ventilation levels up to those at WR max . During resting hyperpnoea, intercostal muscle blood flow increased linearly with the work of breathing (R 2 = 0.94) to 73.0 ± 8.8 ml min −1 (100 g) −1 at the ventilation seen at WR max (work of breathing ∼550-600 J min −1 ), but during exercise it peaked at 80% WR max (53.4 ± 10.3 ml min −1 (100 g) −1 ), significantly falling to 24.7 ± 5.3 ml min −1 (100 g)at WR max . 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% WR max (the latter at 95.4 ± 11.8 ml min −1 (100 g) −1 ). In conclusion, during exercise above 80% WR max 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 O 2 extraction and likely contributing to respiratory muscle fatigue.

During intense exercise in COPD, restriction of intercostal muscle perfusion but preservation of quadriceps muscle blood flow along with attainment of a plateau in cardiac output represents the inability of the circulatory system to satisfy the energy demands of locomotor and respiratory muscles.

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.Abstract During maximal hypoxic exercise, a reduction in cerebral oxygen delivery may constitute a signal to the central nervous system to terminate exercise. We investigated whether the rate of increase in frontal cerebral cortex oxygen delivery is limited in hypoxic compared to normoxic exercise. We assessed frontal cerebral cortex blood flow using near-infrared spectroscopy and the light-absorbing tracer indocyanine green dye, as well as frontal cortex oxygen saturation (S tO 2 %) in 11 trained cyclists during graded incremental exercise to the limit of tolerance (maximal work rate, WR max ) in normoxia and acute hypoxia (inspired O 2 fraction (F IO 2 ), 0.12). In normoxia, frontal cortex blood flow and oxygen delivery increased (P < 0.05) from baseline to sub-maximal exercise, reaching peak values at near-maximal exercise (80% WR max : 287 ± 9 W; 81 ± 23% and 75 ± 22% increase relative to baseline, respectively), both leveling off thereafter up to WR max (382 ± 10 W). Frontal cortex S tO 2 % did not change from baseline (66 ± 3%) throughout graded exercise. During hypoxic exercise, frontal cortex blood flow increased (P = 0.016) from baseline to sub-maximal exercise, peaking at 80% WR max (213 ± 6 W; 60 ± 15% relative increase) before declining towards baseline at WR max (289 ± 5 W). Despite this, frontal cortex oxygen delivery remained unchanged from baseline throughout graded exercise, being at WR max lower than at comparable loads (287 ± 9 W) in normoxia (by 58 ± 12%; P = 0.01). Frontal cortex S tO 2 % fell from baseline (58 ± 2%) on light and moderate exercise in parallel with arterial oxygen saturation, but then remained unchanged to exhaustion (47 ± 1%). Thus, during maximal, but not light to

Abstract-During myocardial reperfusion, polymorphonuclear neutrophil (PMN) adhesion involving the intercellular adhesion molecule-1 (ICAM-1) may lead to aggravation and prolongation of reperfusion injury. We studied the role of early tumor necrosis factor-␣ (TNF-␣) cleavage and nuclear factor-B (NF-B) activation on ICAM-1 expression and venular adhesion of PMN in isolated hearts after ischemia (15 minutes) and reperfusion (30 to 480 minutes). NF-B activation (electromobility shift assay) was found after 30 minutes of reperfusion and up to 240 minutes. ICAM-1 mRNA, assessed by Northern blot, increased during the same interval. Functional effect of newly synthesized adhesion molecules was found by quantification (in situ fluorescence microscopy) of PMN, given as bolus after ischemia, which became adherent to small coronary venules (10 to 50 m in diameter). After 480 minutes of reperfusion, ICAM-1-dependent PMN adhesion increased 2.5-fold compared with PMN adhesion obtained during acute reperfusion.To study the influence of NF-B on PMN adhesion, we inhibited NF-B activation by transfection of NF-B decoy oligonucleotides into isolated hearts using HJV-liposomes. Decoy NF-B but not control oligonucleotides blocked ICAM-1 upregulation and inhibited the subacute increase in PMN adhesion. Similar effects were obtained using BB 1101 (10 g), an inhibitor of TNF-␣ cleavage enzyme. These data suggest that ischemia and reperfusion in isolated hearts cause liberation of TNF-␣, activation of NF-B, and upregulation of ICAM-1, an adhesion molecule involved in inflammatory response after ischemia and reperfusion. (Circ Res. 1999;84:392-400.)

Emerging evidence indicates that, besides dyspnea relief, an improvement in locomotor muscle oxygen delivery may also contribute to enhanced exercise tolerance following normoxic heliox (replacement of inspired nitrogen by helium) administration in patients with chronic obstructive pulmonary disease (COPD). Whether blood flow redistribution from intercostal to locomotor muscles contributes to this improvement currently remains unknown. Accordingly, the objective of this study was to investigate whether such redistribution plays a role in improving locomotor muscle oxygen delivery while breathing heliox at near-maximal [75% peak work rate (WR(peak))], maximal (100%WR(peak)), and supramaximal (115%WR(peak)) exercise in COPD. Intercostal and vastus lateralis muscle perfusion was measured in 10 COPD patients (FEV(1) = 50.5 ± 5.5% predicted) by near-infrared spectroscopy using indocyanine green dye. Patients undertook exercise tests at 75 and 100%WR(peak) breathing either air or heliox and at 115%WR(peak) breathing heliox only. Patients did not exhibit exercise-induced hyperinflation. Normoxic heliox reduced respiratory muscle work and relieved dyspnea across all exercise intensities. During near-maximal exercise, quadriceps and intercostal muscle blood flows were greater, while breathing normoxic heliox compared with air (35.8 ± 7.0 vs. 29.0 ± 6.5 and 6.0 ± 1.3 vs. 4.9 ± 1.2 ml·min(-1)·100 g(-1), respectively; P < 0.05; mean ± SE). In addition, compared with air, normoxic heliox administration increased arterial oxygen content, as well as oxygen delivery to quadriceps and intercostal muscles (from 47 ± 9 to 60 ± 12, and from 8 ± 1 to 13 ± 3 mlO(2)·min(-1)·100 g(-1), respectively; P < 0.05). In contrast, normoxic heliox had neither an effect on systemic nor an effect on quadriceps or intercostal muscle blood flow and oxygen delivery during maximal or supramaximal exercise. Since intercostal muscle blood flow did not decrease by normoxic heliox administration, blood flow redistribution from intercostal to locomotor muscles does not represent a likely mechanism of improvement in locomotor muscle oxygen delivery. Our findings might not be applicable to patients who hyperinflate during exercise.