Impaired angiogenesis and peripheral muscle microcirculation loss contribute to exercise intolerance in pulmonary arterial hypertension.

RATIONALE: Pulmonary arterial hypertension (PAH) is characterized by significant exercise intolerance, which is multifactorial and involves skeletal muscle alterations. There is growing evidence that microRNAs (miRs) are involved in PAH pathogenesis. OBJECTIVES: We hypothesized that miR-126, an endothelial-specific, proangiogenic miR, is down-regulated in the peripheral muscles of patients with PAH, which would account for skeletal muscle microcirculation loss and exercise intolerance. MEASUREMENTS AND MAIN RESULTS: Patients with PAH displayed decreases in exercise capacity ([Formula: see text]o2max) and microcirculation loss on quadriceps muscle biopsy (in CD31(+) immunofluorescence experiments) compared to control subjects. Exercise capacity correlated with muscle capillarity (r = 0.84, P < 0.01). At the cellular level, vascular endothelial growth factor (VEGF) and VEGF receptor 2 expression were similar in both groups. Conversely, PAH was associated with a 60% decrease in miR-126 expression in a quantitative reverse transcriptase polymerase chain reaction experiment (P < 0.01), resulting in up-regulation of its targeted protein, Sprouty-related, EVH1 domain-containing protein 1 (SPRED-1), and a marked decrease in the downstream effectors of the VEGF pathway, p-Raf/Raf and p-ERK/ERK, as determined by immunoblot analysis. Using freshly isolated CD31(+) cells from human quadriceps biopsies, we found that the down-regulation of miR-126 in PAH triggered the activation of SPRED-1, impairing the angiogenic response (Matrigel assay). These abnormalities were reversed by treating the PAH cells with miR-126 mimic, whereas inhibition of miR-126 (antagomir) in healthy CD31(+) cells fully mimicked the PAH phenotype. Finally, miR-126 down-regulation in skeletal muscle of healthy rats decreased muscle capillarity in immunofluorescence assays (P < 0.05) and exercise tolerance in treadmill tests (P < 0.05), whereas miR-126 up-regulation increased them in monocrotaline PAH rats. CONCLUSIONS: We demonstrate for the first time that exercise intolerance in PAH is associated with skeletal muscle microcirculation loss and impaired angiogenesis secondary to miR-126 down-regulation.

Repeatability and responsiveness of exercise tests in pulmonary arterial hypertension.

Exercise tolerance in pulmonary arterial hypertension (PAH) is most commonly assessed by the 6-min walk test (6MWT). Whether endurance exercise tests are more responsive than the 6MWT remains unknown. 20 stable PAH patients (mean±sd age 53±15 years and mean pulmonary arterial pressure 44±16 mmHg) already on PAH monotherapy completed the 6MWT, the endurance shuttle walk test (ESWT) and the cycle endurance test (CET) before and after the addition of sildenafil citrate 20 mg three times daily or placebo for 28 days in a randomised double-blind crossover setting. Pre- or post-placebo tests were used to assess repeatability of each exercise test, whereas pre- or post-sildenafil citrate tests were used to assess their responsiveness. Sildenafil citrate led to placebo-corrected changes in exercise capacity of 18±25 m (p = 0.02), 58±235 s (p = 0.58) and 29±77 s (p = 0.09) for the 6MWT, the ESWT and the CET, respectively. The 6MWT was associated with a lower coefficient of variation between repeated measures (3% versus 18% versus 13%), resulting in a higher standardised response mean compared with endurance tests (0.72, 0.25 and 0.38 for the 6MWT, the ESWT and the CET, respectively). The 6MWT had the best ability to capture changes in exercise capacity when sildenafil citrate was combined with patients' baseline monotherapy, supporting its use as an outcome measure in PAH.

Signal transducers and activators of transcription-3/pim1 axis plays a critical role in the pathogenesis of human pulmonary arterial hypertension.

BACKGROUND: Pulmonary artery hypertension (PAH) is a proliferative disorder associated with enhanced pulmonary artery smooth muscle cell proliferation and suppressed apoptosis. The sustainability of this phenotype required the activation of a prosurvival transcription factor like signal transducers and activators of transcription-3 (STAT3) and nuclear factor of activated T cell (NFAT). Because these factors are implicated in several physiological processes, their inhibition in PAH patients could be associated with detrimental effects. Therefore, a better understanding of the mechanism accounting for their expression/activation in PAH pulmonary artery smooth muscle cells is of great therapeutic interest. METHODS AND RESULTS: Using multidisciplinary and translational approaches, we demonstrated that STAT3 activation in both human and experimental models of PAH accounts for the expression of both NFATc2 and the oncoprotein kinase Pim1, which trigger NFATc2 activation. Because Pim1 expression correlates with the severity of PAH in humans and is confined to the PAH pulmonary artery smooth muscle cell, Pim1 was identified as an attractive therapeutic target for PAH. Indeed, specific Pim1 inhibition in vitro decreases pulmonary artery smooth muscle cell proliferation and promotes apoptosis, all of which are sustained by NFATc2 inhibition. In vivo, tissue-specific inhibition of Pim1 by nebulized siRNA reverses monocrotaline-induced PAH in rats, whereas Pim1 knockout mice are resistant to PAH development. CONCLUSION: We demonstrated for the first time that inhibition of the inappropriate activation of STAT3/Pim1 axis is a novel, specific, and attractive therapeutic strategy to reverse PAH.

Peripheral muscle dysfunction in idiopathic pulmonary arterial hypertension.

BACKGROUND: A majority of patients with idiopathic pulmonary arterial hypertension (IPAH) display persistent exercise intolerance despite new specific therapies. Whether patients with IPAH exhibit peripheral muscle dysfunction that may contribute to this limitation remains unknown. The hypothesis that the muscles of patients with IPAH are weaker and display morphological changes compared with those of control subjects and that those changes partly correlate with their exercise capacity was tested. OBJECTIVE: To characterise quadriceps function, morphology and the enzymatic profile of patients with IPAH. METHODS: Exercise capacity, limb muscle cross-sectional area by CT scan, quadriceps strength by maximal voluntary contraction and non-volitional magnetic stimulation of the femoral nerve (quadriceps twitch; TWq), and muscle morphology and enzymatic profile by quadriceps biopsy of 10 patients with IPAH were compared with those of 10 matched controls subjects. RESULTS: Patients with IPAH displayed a lower proportion of type I muscle fibres (p=0.05), a lower maximal voluntary contraction (p=0.05) and TWq (p=0.01), and an increased muscular phosphofructokinase/3-hydroxyacyl-CoA-dehydrogenase ratio (p=0.05). They also tended to have lower thigh muscle cross-sectional area (p=0.15). Maximal oxygen uptake correlated with quadriceps strength (R(2)=0.42, p=0.04), and oxygen uptake at anaerobic threshold correlated with muscle oxidative capacity assessed by oxidative enzyme level for citrate synthase (R(2)=0.45, p=0.05) and 3-hydroxyacyl-CoA-dehydrogenase (R(2)=0.86, p<0.01), and type I fibre capillarity (R(2)=0.57, p=0.02). CONCLUSION: Patients with IPAH present significant peripheral muscle changes that partly correlated with their exercise capacity.

Impact of preinduced quadriceps fatigue on exercise response in chronic obstructive pulmonary disease and healthy subjects.

Exercise intolerance in chronic obstructive pulmonary disease (COPD) results from a complex interaction between central (ventilatory) and peripheral (limb muscles) components of exercise limitation. The purpose of this study was to evaluate the influence of quadriceps muscle fatigue on exercise tolerance and ventilatory response during constant-workrate cycling exercise testing (CWT) in patients with COPD and healthy subjects. Fifteen patients with COPD and nine age-matched healthy subjects performed, 7 days apart, two CWTs up to exhaustion at 80% of their predetermined maximal work capacity. In a randomized order, one test was performed with preinduced quadriceps fatigue and the other in a fresh state. Quadriceps fatigue was produced by electrostimulation-induced contractions and quantified by maximal voluntary contraction and potentiated twitch force (TwQ(pot)). Endurance time and ventilatory response during CWT were compared between fatigued and fresh state. Endurance time significantly decreased in the fatigued state compared with the fresh condition in COPD (356 +/- 69 s vs. 294 +/- 45 s, P < 0.05) and controls (450 +/- 74 s vs. 340 +/- 45 s, P < 0.05). Controls showed significantly higher ventilation and end-exercise dyspnea scores in the fatigued condition, whereas, in COPD, fatigue did not influence ventilation or dyspnea during exercise. The degree of ventilatory limitation, as expressed by the Ve/maximum voluntary ventilation ratio, was similar in both conditions in patients with COPD. We conclude that it is possible to induce quadriceps fatigue by local electrostimulation-induced contractions. Our findings demonstrate that peripheral muscle fatigue is an additional important factor, besides intense dyspnea, that limits exercise tolerance in COPD.

Impaired Skeletal Muscle Oxygenation and Exercise Tolerance in Pulmonary Hypertension.

BACKGROUND: Limb muscle dysfunction is documented in pulmonary arterial hypertension (PAH), but little is known regarding muscle oxygen (O2) supply and its possible effects on exercise tolerance in PAH. METHODS: Ten patients with PAH and 10 matched controls underwent progressive maximal cardiopulmonary exercise test, voluntary and nonvolitional dominant quadriceps muscle strength measures, and nondominant quadriceps biopsy to assess maximal oxygen uptake, muscle function, and lower limb fiber type and capillarity, respectively. Both groups then performed normoxic and hyperoxic submaximal intensity exercise protocol at the same absolute workload during which muscle O2 supply was assessed by measuring changes in myoglobin-deoxyhemoglobin level (Δ[Mb-HHb]) and tissue oxygenation index in the dominant quadriceps using near-infrared spectroscopy. Changes in cardiac output, estimated systemic O2 delivery, and systemic O2 saturation were also assessed noninvasively throughout both submaximal exercises. RESULTS: Patients with PAH displayed lower maximal oxygen uptake (P < 0.01), skeletal muscle strength (P < 0.05), and capillarity (P = 0.01). Throughout the normoxic submaximal exercise protocol, Δ[Mb-HHb] (P < 0.01) was higher whereas changes in tissue oxygenation index (P < 0.01) and systemic O2 saturation (P = 0.01) were lower in patients with PAH compared with those in controls. Conversely, changes in cardiac output and estimated systemic O2 delivery were similar between groups. Muscle oxygenation remained unchanged with O2 supplementation. Among variables known to influence tissue oxygenation, only quadriceps capillarity density correlated with Δ[Mb-HHb] (r = -0.66, P < 0.01), which in turn correlated with maximal oxygen uptake (r = -0.64, P < 0.01), 6-min walked distance (r = -0.74, P = 0.01), and both voluntary (r = -0.46, P = 0.04) and nonvolitional (r = -0.50, P = 0.02) quadriceps strength. CONCLUSIONS: Capillary rarefaction within the skeletal muscle influences exercise tolerance and quadriceps strength at least partly through impaired muscle oxygen supply in PAH.

Tolerance and physiological correlates of neuromuscular electrical stimulation in COPD: a pilot study.

RATIONALE: Neuromuscular electrical stimulation (NMES) of the lower limbs is an emerging training strategy in patients with COPD. The efficacy of this technique is related to the intensity of the stimulation that is applied during the training sessions. However, little is known about tolerance to stimulation current intensity and physiological factors that could determine it. Our goal was to find potential physiological predictors of the tolerance to increasing NMES stimulation intensity in patients with mild to severe COPD. METHODS: 20 patients with COPD (FEV1 = 54±14% pred.) completed 2 supervised NMES sessions followed by 5 self-directed sessions at home and one final supervised session. NMES was applied simultaneously to both quadriceps for 45 minutes, at a stimulation frequency of 50 Hz. Spirometry, body composition, muscle function and aerobic capacity were assessed at baseline. Cardiorespiratory responses, leg discomfort, muscle fatigue and markers of systemic inflammation were assessed during or after the last NMES session. Tolerance to NMES was quantified as the increase in current intensity from the initial to the final NMES session (ΔInt). RESULTS: Mean ΔInt was 12±10 mA. FEV1, fat-free-mass, quadriceps strength, aerobic capacity and leg discomfort during the last NMES session positively correlated with ΔInt (r = 0.42 to 0.64, all p≤0.06) while post/pre NMES IL-6 ratio negatively correlated with ΔInt (r = -0.57, p = 0.001). FEV1, leg discomfort during last NMES session and post/pre IL-6 ratio to NMES were independent factors of variance in ΔInt (r2 = 0.72, p = 0.001). CONCLUSION: Lower tolerance to NMES was associated with increasing airflow obstruction, low tolerance to leg discomfort during NMES and the magnitude of the IL-6 response after NMES. TRIAL REGISTRATION: ClinicalTrials.gov NCT00809120.

Alternatives to the six-minute walk test in pulmonary arterial hypertension.

INTRODUCTION: The physiological response during the endurance shuttle walk test (ESWT), the cycle endurance test (CET) and the incremental shuttle walk test (ISWT) remains unknown in PAH. We tested the hypothesis that endurance tests induce a near-maximal physiological demand comparable to incremental tests. We also hypothesized that differences in respiratory response during exercise would be related to the characteristics of the exercise tests. METHODS: Within two weeks, twenty-one PAH patients (mean age: 54(15) years; mean pulmonary arterial pressure: 42(12) mmHg) completed two cycling exercise tests (incremental cardiopulmonary cycling exercise test (CPET) and CET) and three field tests (ISWT, ESWT and six-minute walk test (6MWT)). Physiological parameters were continuously monitored using the same portable telemetric device. RESULTS: Peak oxygen consumption (VO(2peak)) was similar amongst the five exercise tests (p = 0.90 by ANOVA). Walking distance correlated markedly with the VO(2peak) reached during field tests, especially when weight was taken into account. At 100% exercise, most physiological parameters were similar between incremental and endurance tests. However, the trends overtime differed. In the incremental tests, slopes for these parameters rose steadily over the entire duration of the tests, whereas in the endurance tests, slopes rose sharply from baseline to 25% of maximum exercise at which point they appeared far less steep until test end. Moreover, cycling exercise tests induced higher respiratory exchange ratio, ventilatory demand and enhanced leg fatigue measured subjectively and objectively. CONCLUSION: Endurance tests induce a maximal physiological demand in PAH. Differences in peak respiratory response during exercise are related to the modality (cycling vs. walking) rather than the progression (endurance vs. incremental) of the exercise tests.

Pathogenesis of hyperinflation in chronic obstructive pulmonary disease.

Chronic obstructive pulmonary disease (COPD) is a preventable and treatable lung disease characterized by airflow limitation that is not fully reversible. In a significant proportion of patients with COPD, reduced lung elastic recoil combined with expiratory flow limitation leads to lung hyperinflation during the course of the disease. Development of hyperinflation during the course of COPD is insidious. Dynamic hyperinflation is highly prevalent in the advanced stages of COPD, and new evidence suggests that it also occurs in many patients with mild disease, independently of the presence of resting hyperinflation. Hyperinflation is clinically relevant for patients with COPD mainly because it contributes to dyspnea, exercise intolerance, skeletal muscle limitations, morbidity, and reduced physical activity levels associated with the disease. Various pharmacological and nonpharmacological interventions have been shown to reduce hyperinflation and delay the onset of ventilatory limitation in patients with COPD. The aim of this review is to address the more recent literature regarding the pathogenesis, assessment, and management of both static and dynamic lung hyperinflation in patients with COPD. We also address the influence of biological sex and obesity and new developments in our understanding of hyperinflation in patients with mild COPD and its evolution during progression of the disease.

Functional and muscular effects of neuromuscular electrical stimulation in patients with severe COPD: a randomized clinical trial.

BACKGROUND: The mechanisms through which neuromuscular electrical stimulation (NMES) training may improve limb muscle function and exercise tolerance in COPD are poorly understood. We investigated the functional and muscular effects of NMES in advanced COPD. METHODS: Twenty of 22 patients with COPD were randomly assigned to NMES (n = 12) or sham (n = 8) training in a double-blind controlled study. NMES was performed on quadriceps and calf muscles, at home, 5 days per week for 6 weeks. Quadriceps and calf muscle cross-sectional area (CSA), quadriceps force and endurance, and the shuttle-walking distance with cardiorespiratory measurements were assessed before and after training. Quadriceps biopsy specimens were obtained to explore the insulin-like growth factor-1/AKT signaling pathway (70-kDa ribosomal S6 kinase [p70S6K] , atrogin-1). RESULTS: NMES training improved muscle CSA (P < .05), force, and endurance (P < .03) when compared with sham training. Phosphorylated p70S6K levels (anabolism) were increased after NMES as compared with sham (P = .03), whereas atrogin-1 levels (catabolism) were reduced (P = .01). Changes in quadriceps strength and ventilation during walking contributed independently to variations in walking distance after training (r = 0.77, P < .001). Gains in walking distance were related to the ability to tolerate increasing current intensities during training (r = 0.95, P < .001). CONCLUSIONS: In patients with severe COPD, NMES improved muscle CSA. This was associated with a more favorable muscle anabolic to catabolic balance. Improvement in walking distance after NMES training was associated with gains in muscle strength, reduced ventilation during walking, and the ability to tolerate higher stimulation intensity. TRIAL REGISTRY: ClinicalTrials.gov; No.: NCT00874965; URL: www.clinicaltrials.gov.

Effect of bisoprolol on respiratory function and exercise capacity in chronic obstructive pulmonary disease.

Cardioselective ß blockers are considered to have little impact on lung function at rest in patients with chronic obstructive pulmonary disease (COPD). However, their effects on dynamic hyperinflation, an important mechanism contributing to symptoms and exercise tolerance in patients with COPD, have not been evaluated. Twenty-seven patients with moderate to severe COPD (forced expiratory volume in 1 second 52 ± 13% predicted) completed pulmonary function tests, echocardiography, maximal exercise tests, and cycle endurance tests at baseline. Inspiratory capacity was measured at 2-minute intervals during the cycle endurance test to quantify dynamic hyperinflation. Pulmonary function and cycle endurance testing were repeated after 14 days of bisoprolol 10 mg/day and 14 days of placebo in a randomized, double-blind, placebo-controlled, crossover trial. The extent of dynamic hyperinflation at peak isotime exercise with bisoprolol and placebo was compared. Peak isotime was defined as the latest time point that was reached during the 2 cycle endurance tests. Changes in inspiratory capacity from rest to peak isotime were different with bisoprolol compared to placebo (-0.50 ± 0.35 vs -0.41 ± 0.33 L, p = 0.01). Exercise duration tended to be lower with bisoprolol compared to placebo (305 ± 125 vs 353 ± 172 seconds, p = 0.11). The magnitude of change in exercise duration between the bisoprolol and placebo conditions was correlated with the magnitude of change in inspiratory capacity (r = 0.57, p <0.01). In conclusion, bisoprolol was associated with modest worsening dynamic hyperinflation, whereas exercise duration remained unchanged in patients with moderate to severe COPD. The magnitude of these effects was small and should not contraindicate the use of bisoprolol in patients with COPD.

Assessment of daily life physical activities in pulmonary arterial hypertension.

BACKGROUND: In pulmonary arterial hypertension (PAH), the six-minute walk test (6MWT) is believed to be representative of patient's daily life physical activities (DL(PA)). Whether DL(PA) are decreased in PAH and whether the 6MWT is representative of patient's DL(PA) remain unknown. METHODS: 15 patients with idiopathic PAH (IPAH) and 10 patients with PAH associated with limited systemic sclerosis (PAH-SSc) were matched with 15 healthy control subjects and 10 patients with limited systemic sclerosis without PAH. Each subject completed a 6MWT. The mean number of daily steps and the mean energy expenditure and duration of physical activities >3 METs were assessed with a physical activity monitor for seven consecutive days and used as markers of DL(PA). RESULTS: The mean number of daily steps and the mean daily energy expenditure and duration of physical activities >3 METs were all reduced in PAH patients compared to their controls (all p<0.05). The mean number of daily steps correlated with the 6MWT distance for both IPAH and PAH-SSc patients (r = 0.76, p<0.01 and r = 0.85, p<0.01), respectively. CONCLUSION: DL(PA) are decreased in PAH and correlate with the 6MWT distance. Functional exercise capacity may thus be a useful surrogate of DL(PA) in PAH.

Effects of a rehabilitation program on skeletal muscle function in idiopathic pulmonary arterial hypertension.

INTRODUCTION: A majority of patients with idiopathic pulmonary arterial hypertension (IPAH) display persistent exercise intolerance despite current therapies. Whether a rehabilitation program elicits favorable changes in muscle function which would partly explain improvements in exercise tolerance of IPAH patients remains unknown. We performed this study to assess the effect of a 12-week rehabilitation program on skeletal muscle characteristics and exercise tolerance in patients with IPAH. METHODS: Exercise capacity measured by the 6-minute walk test and by the cycle endurance test (CET), limb muscle cross-sectional area, quadriceps function by maximal voluntary contraction and magnetic stimulation (potentiated twitches), and molecular muscle characteristics by quadriceps biopsy of 5 IPAH patients were assessed before and after a 12-week rehabilitation program. RESULTS: Following training, improvements in all patients were observed for the 6-minute walk test distance, from 441 (75) to 499 (85) m, P = .01, and the CET time, from 429 (239) to 633 (380) seconds, P = .16. Minute ventilation assessed at isotime during CET decreased by 15(11)%, P = .05. This was related to both decreased carbon dioxide output and (Equation is included in full-text article.)E/(Equation is included in full-text article.)co2. These improvements were associated with decreased type IIx fiber proportion, 31(8)% to 23(10)%, P = .05. DISCUSSION: Peripheral muscle characteristic improvements may contribute to the clinical benefit observed following a rehabilitation program in IPAH.