Reliability of the passive leg movement assessment of vascular function in men.

NEW FINDINGS: What is the central question of this study? Use of the passive leg movement (PLM) test, a non-invasive assessment of microvascular function, is on the rise. However, PLM reliability in men has not been adequately investigated, nor has such reliability data, in men, been compared to the most commonly employed vascular function assessment, flow-mediated vasodilation (FMD). What is the main finding and its importance? PLM is a reliable method to assess vascular function in men, and is comparable to values previously reported for PLM in women, and for FMD. Given the importance of vascular function as a predictor of cardiovascular disease risk, these data support the utility of PLM as a clinically relevant measurement. ABSTRACT: Although vascular function is an independent predictor of cardiovascular disease risk, and therefore has significant prognostic value, there is currently not a single clinically accepted method of assessment. The passive leg movement (PLM) assessment predominantly reflects microvascular endothelium-dependent vasodilation and can identify decrements in vascular function with advancing age and pathology. Reliability of the PLM model was only recently determined in women, and has not been adequately investigated in men. Twenty healthy men (age: 27 ± 2 year) were studied on three separate experimental days, resulting in three within-day and three between-day trials. The hyperemic response to PLM was assessed with Doppler ultrasound, and expressed as the absolute peak in leg blood flow (LBFpeak ), change from baseline to peak (ΔLBFpeak ), absolute area under the curve (LBFAUC ), and change in AUC from baseline (ΔLBFAUC ). PLM-induced hyperemia yielded within-day coefficients of variation (CV) from 10.9 to 22.9%, intraclass correlation coefficients (ICC) from 0.82 to 0.90, standard error of the measurement (SEM) from 8.3 to 17.2%, and Pearson's correlation coefficients (r) from 0.56 to 0.81. Between-day assessments of PLM hyperemia resulted in CV from 14.4 to 25%, ICC from 0.75 to 0.87, SEM from 9.8 to 19.8%, and r from 0.46 to 0.75. Similar to previous reports in women, the hyperemic responses to PLM in men display moderate-to-high reliability, and are comparable to reliability data for brachial artery flow mediated vasodilation. These positive reliability findings further support the utility of PLM as a clinical measurement of vascular function and cardiovascular disease risk.

Attenuated nitric oxide bioavailability in systemic sclerosis: Evidence from the novel assessment of passive leg movement.

NEW FINDINGS: What is the central question of this study? Do systemic sclerosis patients exhibit impaired nitric oxide-mediated vascular function of the lower limb and are these decrements correlated with plasma biomarkers for inflammation and oxidative stress? What is the main finding and its importance? Findings indicate impaired nitric oxide-mediated vascular function, linked to the incidence of digital ulcers and a milieu of inflammation and oxidative stress. However, the absence of significant correlations between individual biomarkers and blood flow responses suggests that the vasculopathy observed in systemic sclerosis may not be solely the result of derangements in the redox balance or inflammatory signalling. ABSTRACT: Systemic sclerosis (SSc) is an autoimmune disease characterized by vasculopathy, which may be the consequence of inflammation and oxidative stress that ultimately leads to a reduced nitric oxide (NO) bioavailability. Passive leg movement (PLM) is a novel methodology for assessing lower limb vascular function that is predominantly NO dependent. We combined this vascular assessment with a comprehensive panel of plasma biomarkers to assess the axis of inflammation, oxidative stress and NO in SSc patients (n = 12; 62 ± 11 years of age) compared with healthy control subjects (n = 17; 60 ± 16 years of age). The PLM-induced changes in leg blood flow (LBF; 191 ± 104 versus 327 ± 217 ml min-1 ) and LBF area under the curve (39 ± 104 versus 125 ± 131 ml) were reduced in SSc compared with control subjects. Stratification of patients according to history of digital ulcer (DU) formation revealed a further reduction in LBF area under the curve in DU (-13 ± 83 ml) versus non-DU (91 ± 102 ml) patients. Biomarkers of inflammation (C-reactive protein) and oxidative stress (malondialdehyde and protein carbonyl) were all elevated in SSc (C-reactive protein, 3299 ± 2372 versus 984 ± 565 ng ml-1 ; malondialdehyde, 3.2 ± 1.1 versus 1.1 ± 0.7 µm; and protein carbonyl, 0.15 ± 0.05 versus 0.12 ± 0.03 nmol mg-1 ), and C-reactive protein was further elevated in patients with a history of DU (4551 ± 2752 versus 2047 ± 1019 ng ml-1 ) compared with non-DU, although these were not individually correlated with changes in LBF. These findings of impaired NO-mediated vascular function, linked to DU and a milieu of inflammation and oxidative stress, suggest that redox balance plays an important, but not necessarily deterministic, role in the vascular pathophysiology of SSc.

Single passive leg movement assessment of vascular function: contribution of nitric oxide.

Broxterman RM, Trinity JD, Gifford JR, Kwon OS, Kithas AC, Hydren JR, Nelson AD, Morgan DE, Jessop JE, Bledsoe AD, Richardson RS. Single passive leg movement assessment of vascular function: contribution of nitric oxide. J Appl Physiol 123: 1468-1476, 2017. First published August 31, 2017; doi:10.1152/japplphysiol.00533.2017.-The assessment of passive leg movement (PLM)-induced leg blood flow (LBF) and vascular conductance (LVC) is a novel approach to assess vascular function that has recently been simplified to only a single PLM (sPLM), thereby increasing the clinical utility of this technique. As the physiological mechanisms mediating the robust increase in LBF and LVC with sPLM are unknown, we tested the hypothesis that nitric oxide (NO) is a major contributor to the sPLM-induced LBF and LVC response. In nine healthy men, sPLM was performed with and without NO synthase inhibition by intra-arterial infusion of NG-monomethyl-l-arginine (l-NMMA). Doppler ultrasound and femoral arterial pressure were used to determine LBF and LVC, which were characterized by the peak change (ΔLBFpeak and ΔLVCpeak) and area under the curve (LBFAUC and LVCAUC). l-NMMA significantly attenuated ΔLBFpeak [492 ± 153 (l-NMMA) vs. 719 ± 238 (control) ml/min], LBFAUC [57 ± 34 (l NMMA) vs. 147 ± 63 (control) ml], ΔLVCpeak [4.7 ± 1.1 (l-NMMA) vs. 8.0 ± 3.0 (control) ml·min-1·mmHg-1], and LVCAUC [0.5 ± 0.3 (l-NMMA) vs. 1.6 ± 0.9 (control) ml/mmHg]. The magnitude of the NO contribution to LBF and LVC was significantly correlated with the magnitude of the control responses ( r = 0.94 for ΔLBFpeak, r = 0.85 for LBFAUC, r = 0.94 for ΔLVCpeak, and r = 0.95 for LVCAUC). These data establish that the sPLM-induced hyperemic and vasodilatory response is predominantly (~65%) NO-mediated. As such, sPLM appears to be a promising, simple, in vivo assessment of NO-mediated vascular function and NO bioavailability. NEW & NOTEWORTHY Passive leg movement (PLM), a novel assessment of vascular function, has been simplified to a single PLM (sPLM), thereby increasing the clinical utility of this technique. However, the role of nitric oxide (NO) in mediating the robust sPLM hemodynamic responses is unknown. This study revealed that sPLM induces a hyperemic and vasodilatory response that is predominantly NO-mediated and, as such, appears to be a promising simple, in vivo, clinical assessment of NO-mediated vascular function and, therefore, NO bioavailability.

CORP: Ultrasound assessment of vascular function with the passive leg movement technique.

As dysfunction of the vascular system is an early, modifiable step in the progression of many cardiovascular diseases, there is demand for methods to monitor the health of the vascular system noninvasively in clinical and research settings. Validated by very good agreement with more technical assessments of vascular function, like intra-arterial drug infusions and flow-mediated dilation, the passive leg movement (PLM) technique has emerged as a powerful, yet relatively simple, test of peripheral vascular function. In the PLM technique, the change in leg blood flow elicited by the passive movement of the leg through a 90° range of motion is quantified with Doppler ultrasound. This relatively easy-to-learn test has proven to be ≤80% dependent on nitric oxide bioavailability and is especially adept at determining peripheral vascular function across the spectrum of cardiovascular health. Indeed, multiple reports have documented that individuals with decreased cardiovascular health such as the elderly and those with heart failure tend to exhibit a substantially blunted PLM-induced hyperemic response (~50 and ~85% reduction, respectively) compared with populations with good cardiovascular health such as young individuals. As specific guidelines have not yet been put forth, the purpose of this Cores of Reproducibility in Physiology (CORP) article is to provide a comprehensive reference for the assessment and interpretation of vascular function with PLM with the aim to increase reproducibility and consistency among studies and facilitate the use of PLM as a research tool with clinical relevance.

Single passive leg movement-induced hyperemia: a simple vascular function assessment without a chronotropic response.

Passive leg movement (PLM)-induced hyperemia is a novel approach to assess vascular function, with a potential clinical role. However, in some instances, the varying chronotropic response induced by PLM has been proposed to be a potentially confounding factor. Therefore, we simplified and modified the PLM model to require just a single PLM (sPLM), an approach that may evoke a peripheral hemodynamic response, allowing a vascular function assessment, but at the same time minimizing central responses. To both characterize and assess the utility of sPLM, in 12 healthy subjects, we measured heart rate (HR), stroke volume, cardiac output (CO), mean arterial pressure (MAP), leg blood flow (LBF), and calculated leg vascular conductance (LVC) during both standard PLM, consisting of passive knee flexion and extension performed at 1 Hz for 60 s, and sPLM, consisting of only a single passive knee flexion and extension over 1 s. During PLM, MAP transiently decreased (5 ± 1 mmHg), whereas both HR and CO increased from baseline (6.0 ± 1.1 beats/min, and 0.8 ± 0.01 l/min, respectively). Following sPLM, MAP fell similarly (5 ± 2 mmHg; P = 0.8), but neither HR nor CO responses were identifiable. The peak LBF and LVC response was similar for PLM (993 ± 189 ml/min; 11.9 ± 1.5 ml·min-1·mmHg-1, respectively) and sPLM (878 ± 119 ml/min; 10.9 ± 1.6 ml·min-1·mmHg-1, respectively). Thus sPLM represents a variant of the PLM approach to assess vascular function that is more easily performed and evokes a peripheral stimulus that induces a significant hyperemia, but does not generate a potentially confounding, chronotropic response, which may make sPLM more useful clinically. NEW & NOTEWORTHY: Using the single passive leg movement (PLM) technique, a variant of the vascular function assessment PLM, we have identified a novel peripheral vascular assessment method that is more easily performed than PLM, which, by not evoking potentially confounding central hemodynamic responses, may be more useful clinically.

Nitric oxide-mediated vascular function in sepsis using passive leg movement as a novel assessment: a cross-sectional study.

Post-cuff occlusion flow-mediated dilation (FMD) is a proposed indicator of nitric oxide (NO) bioavailability and vascular function. FMD is reduced in patients with sepsis and may be a marker of end organ damage and mortality. However, FMD likely does not solely reflect NO-mediated vasodilation, is technically challenging, and often demonstrates poor reproducibility. In contrast, passive leg movement (PLM), a novel methodology to assess vascular function, yields a hyperemic response that is predominately NO-dependent, reproducible, and easily measured. This study evaluated PLM as an approach to assess NO-mediated vascular function in patients with sepsis. We hypothesized that PLM-induced hyperemia, quantified by the increase in leg blood flow (LBF), would be attenuated in sepsis. In a cross-sectional study, 17 subjects in severe sepsis or septic shock were compared with 16 matched healthy controls. Doppler ultrasound was used to assess brachial artery FMD and the hyperemic response to PLM in the femoral artery. FMD was attenuated in septic compared with control subjects (1.1 ± 1.7% vs. 6.8 ± 1.3%; values are means ± SD). In terms of PLM, baseline LBF (196 ± 33 ml/min vs. 328 ± 20 ml/min), peak change in LBF from baseline (133 ± 28 ml/min vs. 483 ± 86 ml/min), and the LBF area under the curve (16 ± 8.3 vs. 143 ± 33) were all significantly attenuated in septic subjects. Vascular function, as assessed by both FMD and PLM, is attenuated in septic subjects compared with controls. These data support the concept that NO bioavailability is attenuated in septic subjects, and PLM appears to be a novel and feasible approach to assess NO-mediated vascular function in sepsis.

In vivo evidence of an age-related increase in ATP cost of contraction in the plantar flexor muscles.

Impaired skeletal muscle efficiency potentially contributes to the age-related decline in exercise capacity and may explain the altered haemodynamic response to exercise in the elderly. Thus we examined whether (i) the ATP cost of contraction increases with age, and (ii) this results in altered convective O(2) delivery to maintain microvascular oxygenation in the calf muscle. To this aim, we used an integrative experimental approach combining (31)P-MRS (magnetic resonance spectroscopy), Doppler ultrasound imaging and NIRS (near-IR spectroscopy) during dynamic plantar flexion exercise at 40% of WR(max) (maximal power output) in 20 healthy young and 20 older subjects matched for physical activity. The ATP cost of contraction was significantly higher in the old (7.2±4.1 mM/min per W) compared with the young (2.4±1.9 mM/min per W; P<0.05) and this was only significantly correlated with the plantar flexion WR(max) value in the old subjects (r=-0.52; P<0.05). Even when differences in power output were taken into account, end-exercise blood flow (old, 259±168 ml/min per W and young, 134±40 ml/min per W; P<0.05) and convective O(2) delivery (old, 0.048±0.031 l/min per W and young, 0.026±0.008 l/min per W; P<0.05) were greater in the old in comparison with the young subjects. In contrast, the NIRS oxyhaemoglobin, deoxyhaemoglobin and microvascular oxygenation indices were not significantly different between the groups (P>0.05). Therefore the present study reveals that, although the peripheral haemodynamic responses to plantar flexion exercise appear to be appropriate, the elevated energy cost of contraction and associated reduction in the WR(max) value in this muscle group may play a role in limiting exercise capacity with age.

Mitochondrial function and increased convective O2 transport: implications for the assessment of mitochondrial respiration in vivo.

Although phosphorus magnetic resonance spectroscopy (31P-MRS)-based evidence suggests that in vivo peak mitochondrial respiration rate in young untrained adults is limited by the intrinsic mitochondrial capacity of ATP synthesis, it remains unknown whether a large, locally targeted increase in convective O2 delivery would alter this interpretation. Consequently, we examined the effect of superimposing reactive hyperemia (RH), induced by a period of brief ischemia during the last minute of exercise, on oxygen delivery and mitochondrial function in the calf muscle of nine young adults compared with free-flow conditions (FF). To this aim, we used an integrative experimental approach combining 31P-MRS, Doppler ultrasound imaging, and near-infrared spectroscopy. Limb blood flow [area under the curve (AUC), 1.4 ± 0.8 liters in FF and 2.5 ± 0.3 liters in RH, P < 0.01] and convective O2 delivery (AUC, 0.30 ± 0.16 liters in FF and 0.54 ± 0.05 liters in RH, P < 0.01), were significantly increased in RH compared with FF. RH was also associated with significantly higher capillary blood flow (P < 0.05) and faster tissue reoxygenation mean response times (70 ± 15 s in FF and 24 ± 15 s in RH, P < 0.05). This resulted in a 43% increase in estimated peak mitochondrial ATP synthesis rate (29 ± 13 mM/min in FF and 41 ± 14 mM/min in RH, P < 0.05) whereas the phosphocreatine (PCr) recovery time constant in RH was not significantly different (P = 0.22). This comprehensive assessment of local skeletal muscle O2 availability and utilization in untrained subjects reveals that mitochondrial function, assessed in vivo by 31P-MRS, is limited by convective O2 delivery rather than an intrinsic mitochondrial limitation.

The effect of higher ATP cost of contraction on the metabolic response to graded exercise in patients with chronic obstructive pulmonary disease.

To better understand the metabolic implications of a higher ATP cost of contraction in chronic obstructive pulmonary disease (COPD), we used (31)P-magnetic resonance spectroscopy ((31)P-MRS) to examine muscle energetics and pH in response to graded exercise. Specifically, in six patients and six well-matched healthy controls, we determined the intracellular threshold for pH (T(pH)) and inorganic phosphate-to-phosphocreatine ratio (T(Pi/PCr)) during progressive dynamic plantar flexion exercise with work rate expressed as both absolute and relative intensity. Patients with COPD displayed a lower peak power output (WRmax) compared with controls (controls 25 ± 4 W, COPD 15 ± 5 W, P = 0.01) while end-exercise pH (controls 6.79 ± 0.15, COPD 6.76 ± 0.21, P = 0.87) and PCr consumption (controls 82 ± 10%, COPD 70 ± 18%, P = 0.26) were similar between groups. Both T(pH) and T(Pi/PCr) occurred at a significantly lower absolute work rate in patients with COPD compared with controls (controls: 14.7 ± 2.4 W for T(pH) and 15.3 ± 2.4 W for T(Pi/PCr); COPD: 9.7 ± 4.5 W for T(pH) and 10.0 ± 4.6 W for T(Pi/PCr), P < 0.05), but these thresholds occurred at the same percentage of WRmax (controls: 63 ± 11% WRmax for T(pH) and 67 ± 18% WRmax for T(Pi/PCr); COPD: 59 ± 9% WRmax for T(pH) and 61 ± 12% WRmax for T(Pi/PCr), P > 0.05). Indexes of mitochondrial function, the PCr recovery time constant (controls 42 ± 7 s, COPD 45 ± 11 s, P = 0.66) and the PCr resynthesis rate (controls 105 ± 21%/min, COPD 91 ± 31%/min, P = 0.43) were similar between groups. In combination, these results reveal that when energy demand is normalized to WRmax, as a consequence of higher ATP cost of contraction, patients with COPD display the same metabolic pattern as healthy subjects, suggesting that skeletal muscle energy production is well preserved in these patients.

Evidence that a higher ATP cost of muscular contraction contributes to the lower mechanical efficiency associated with COPD: preliminary findings.

Impaired metabolism in peripheral skeletal muscles potentially contributes to exercise intolerance in chronic obstructive pulmonary disease (COPD). We used (31)P-magnetic resonance spectroscopy ((31)P-MRS) to examine the energy cost and skeletal muscle energetics in six patients with COPD during dynamic plantar flexion exercise compared with six well-matched healthy control subjects. Patients with COPD displayed a higher energy cost of muscle contraction compared with the controls (control: 6.1 ± 3.1% of rest·min(-1)·W(-1), COPD: 13.6 ± 8.3% of rest·min(-1)·W(-1), P = 0.01). Although, the initial phosphocreatine resynthesis rate was also significantly attenuated in patients with COPD compared with controls (control: 74 ± 17% of rest/min, COPD: 52 ± 13% of rest/min, P = 0.04), when scaled to power output, oxidative ATP synthesis was similar between groups (6.5 ± 2.3% of rest·min(-1)·W(-1) in control and 7.8 ± 3.9% of rest·min(-1)·W(-1) in COPD, P = 0.52). Therefore, our results reveal, for the first time that in a small subset of patients with COPD a higher ATP cost of muscle contraction may substantially contribute to the lower mechanical efficiency previously reported in this population. In addition, it appears that some patients with COPD have preserved mitochondrial function and normal energy supply in lower limb skeletal muscle.

Ultrasound assessment of flow-mediated dilation.

Developed in 1992, the flow-mediated dilation test is now the most commonly used noninvasive assessment of vascular endothelial function in humans. Since its inception, scientists have refined their understanding of the physiology, analysis, and interpretation of this measurement. Recently, a significant growth of knowledge has added to our understanding and implementation of this clinically relevant research methodology. Therefore, this tutorial provides timely insight into recent advances and practical information related to the ultrasonic assessment of vascular endothelial function in humans.

Multiparametric NMR-based assessment of skeletal muscle perfusion and metabolism during exercise in elderly persons: preliminary findings.

BACKGROUND: Aging is associated with a decline in exercise capacity that may be attributable to maladaptations in both skeletal muscle perfusion and metabolism; yet very little is known regarding the real-time, within-muscle interplay between these parameters during physical activity. Therefore, we utilized an unique nuclear magnetic resonance sequence to concomitantly examine changes in lower leg skeletal muscle perfusion and metabolism. METHODS: In young (26+/-5 years, n=6) and older (70+/-5 years, n=6) healthy volunteers, arterial spin labeling measurements of muscle perfusion were combined with 31 Phosphorous (31P) nuclear magnetic resonance spectroscopy to monitor high-energy phosphate metabolites during and after 5 minutes of moderate-intensity (approximately 5W) plantar flexion exercise. RESULTS: Compared with young, end-exercise perfusion was diminished in older participants (43+/-10 mL/100 g/minute, old; 60+/-7 mL/100 g.minute, young), accompanied by greater phosphocreatine (PCr) depletion (-28%+/-12%, old; -19%+/-7%, young) and elevated inorganic phosphate/PCr (0.41+/-0.2, old; 0.24+/-0.09, young); yet the time constant for PCr recovery (tau, an index of muscle oxidative capacity) was similar between groups (51+/-17 seconds, old; 48+/-7 seconds, young). CONCLUSIONS: Together, these preliminary data provide evidence of an age-related decline in tissue perfusion and increased "metabolic stress" during exercise but demonstrate that overall oxidative capacity in the elderly does not appear negatively affected by this relatively hypoperfused state.

Skeletal muscle oxidative metabolism in sedentary humans: 31P-MRS assessment of O2 supply and demand limitations.

Previously, it was demonstrated in exercise-trained humans that phosphocreatine (PCr) recovery is significantly altered by fraction of inspired O2 (FI(O2)), suggesting that in this population under normoxic conditions, O2 availability limits maximal oxidative rate. Haseler LJ, Hogan ML, and Richardson RS. J Appl Physiol 86: 2013-2018, 1999. To further elucidate these population-specific limitations to metabolic rate, we used 31P-magnetic resonance spectroscopy to study the exercising human gastrocnemius muscle under conditions of varied FI(O2) in sedentary subjects. To test the hypothesis that PCr recovery from submaximal exercise in sedentary subjects is not limited by O2 availability, but rather by their mitochondrial capacity, six sedentary subjects performed three bouts of 6-min steady-state submaximal plantar flexion exercise followed by 5 min of recovery while breathing three different FI(O2) (0.10, 0.21, and 1.00). PCr recovery time constants were significantly longer in hypoxia (47.0 +/- 3.2 s), but there was no difference between hyperoxia (31.8 +/- 1.9 s) and normoxia (30.0 +/- 2.1 s) (mean +/- SE). End-exercise pH was not significantly different across treatments. These results suggest that the maximal muscle oxidative rate of these sedentary subjects, unlike their exercise-trained counterparts, is limited by mitochondrial capacity and not O2 availability in normoxia. Additionally, the significant elongation of PCr recovery in these subjects in hypoxia illustrates the reliance on O2 supply at the other end of the O2 availability spectrum in both sedentary and active populations.