Rho-kinase inhibition reduces systolic blood pressure and forearm vascular resistance in healthy older adults: a double-blind, randomized, placebo-controlled pilot study.

Rho-kinase has been implicated in the development of hypertension in preclinical studies and may contribute to age-related blood pressure elevation. This study tested the hypothesis that Rho-kinase contributes to elevated systolic blood pressure (SBP) in healthy older adults. Young (18-30 years, 6F/6M) and older (60-80 years, 7F/6M) adults were enrolled in a double-blind, placebo-controlled crossover study using intravenous fasudil infusion to inhibit Rho-kinase. Fasudil lowered SBP in older adults compared to placebo (saline) (2-h post-infusion: 125 ± 4 vs. 133 ± 4 mmHg, P < 0.05), whereas fasudil had no impact on SBP in young adults. Immediately following fasudil infusion, there was a transient reduction in mean arterial pressure (MAP) in young adults that was no longer evident 1-h post-infusion. In older adults, MAP remained lower throughout the fasudil visit compared to placebo (2-h post-infusion: 93 ± 3 vs. 100 ± 3 mmHg, P < 0.05) such that age-related differences in SBP and MAP were abolished. Aortic stiffness (carotid-femoral pulse wave velocity) was not altered by fasudil when central MAP was included as a covariate in analyses. Fasudil reduced forearm vascular resistance in older (2-h post-infusion: 3.3 ± 0.4 vs. 4.8 ± 0.6 mmHg/ml/min, P < 0.05) but not young (4.0 ± 0.6 vs. 3.8 ± 0.5 mmHg/ml/min) adults, which was accompanied by an increase in brachial artery diameter only in older adults. Brachial artery flow-mediated dilation was not affected by fasudil in either group. These findings indicate that Rho-kinase inhibition reduces SBP in healthy older but not young adults, which is associated with a concomitant reduction in forearm vascular resistance.

Daily blueberry consumption for 12 weeks improves endothelial function in postmenopausal women with above-normal blood pressure through reductions in oxidative stress: a randomized controlled trial.

Estrogen-deficient postmenopausal women have oxidative stress-mediated suppression of endothelial function that is exacerbated by high blood pressure. Previous research suggests blueberries may improve endothelial function through reductions in oxidative stress, while also exerting other cardiovascular benefits. The objective of this study was to examine the efficacy of blueberries to improve endothelial function and blood pressure in postmenopausal women with above-normal blood pressure, and to identify potential mechanisms for improvements in endothelial function. A randomized, double-blind, placebo-controlled, parallel-arm clinical trial was performed, where postmenopausal women aged 45-65 years with elevated blood pressure or stage 1-hypertension (total n = 43, endothelial function n = 32) consumed 22 g day-1 of freeze-dried highbush blueberry powder or placebo powder for 12 weeks. Endothelial function was assessed at baseline and 12 weeks through ultrasound measurement of brachial artery flow-mediated dilation (FMD) normalized to shear rate area under the curve (FMD/SRAUC) before and after intravenous infusion of a supraphysiologic dose of ascorbic acid to evaluate whether FMD improvements were mediated by reduced oxidative stress. Hemodynamics, arterial stiffness, cardiometabolic blood biomarkers, and plasma (poly)phenol metabolites were assessed at baseline and 4, 8, and 12 weeks, and venous endothelial cell protein expression was assessed at baseline and 12 weeks. Absolute FMD/SRAUC was 96% higher following blueberry consumption compared to baseline (p < 0.05) but unchanged in the placebo group (p > 0.05), and changes from baseline to 12 weeks were greater in the blueberry group than placebo (+1.09 × 10-4 ± 4.12 × 10-5vs. +3.82 × 10-6 ± 1.59 × 10-5, p < 0.03, respectively). The FMD/SRAUC response to ascorbic acid infusion was lower (p < 0.05) at 12 weeks compared to baseline in the blueberry group with no change in the placebo group (p > 0.05). The sum of plasma (poly)phenol metabolites increased at 4, 8, and 12 weeks in the blueberry group compared to baseline, and were higher than the placebo group (all p < 0.05). Increases in several plasma flavonoid and microbial metabolites were also noted. No major differences were found for blood pressure, arterial stiffness, blood biomarkers, or endothelial cell protein expression following blueberry consumption. These findings suggest daily consumption of freeze-dried blueberry powder for 12 weeks improves endothelial function through reduced oxidative stress in postmenopausal women with above-normal blood pressure. The clinical trial registry number is NCT03370991 (https://clinicaltrials.gov).

Emerging mitochondrial signaling mechanisms in cardio-oncology: beyond oxidative stress.

Many anticancer therapies (CTx) have cardiotoxic side effects that limit their therapeutic potential and cause long-term cardiovascular complications in cancer survivors. This has given rise to the field of cardio-oncology, which recognizes the need for basic, translational, and clinical research focused on understanding the complex signaling events that drive CTx-induced cardiovascular toxicity. Several CTx agents cause mitochondrial damage in the form of mitochondrial DNA deletions, mutations, and suppression of respiratory function and ATP production. In this review, we provide a brief overview of the cardiovascular complications of clinically used CTx agents and discuss current knowledge of local and systemic secondary signaling events that arise in response to mitochondrial stress/damage. Mitochondrial oxidative stress has long been recognized as a contributor to CTx-induced cardiotoxicity; thus, we focus on emerging roles for mitochondria in epigenetic regulation, innate immunity, and signaling via noncoding RNAs and mitochondrial hormones. Because data exploring mitochondrial secondary signaling in the context of cardio-oncology are limited, we also draw upon clinical and preclinical studies, which have examined these pathways in other relevant pathologies.

Rho-kinase inhibition improves haemodynamic responses and circulating ATP during hypoxia and moderate intensity handgrip exercise in healthy older adults.

Skeletal muscle haemodynamics and circulating adenosine triphosphate (ATP) responses during hypoxia and exercise are blunted in older (OA) vs. young (YA) adults, which may be associated with impaired red blood cell (RBC) ATP release. Rho-kinase inhibition improves deoxygenation-induced ATP release from OA isolated RBCs. We tested the hypothesis that Rho-kinase inhibition (via fasudil) in vivo would improve local haemodynamic and ATP responses during hypoxia and exercise in OA. Healthy YA (25 ± 3 years; n = 12) and OA (65 ± 5 years; n = 13) participated in a randomized, double-blind, placebo-controlled, crossover study on two days (≥5 days between visits). A forearm deep venous catheter was used to administer saline/fasudil and sample venous plasma ATP ([ATP]V ). Forearm vascular conductance (FVC) and [ATP]V were measured at rest, during isocapnic hypoxia (80% SpO2${S_{{\rm{p}}{{\rm{O}}_{\rm{2}}}}}$ ), and during graded rhythmic handgrip exercise that was similar between groups (5, 15 and 25% maximum voluntary contraction (MVC)). Isolated RBC ATP release was measured during normoxia/hypoxia. With saline, ΔFVC was lower (P < 0.05) in OA vs. YA during hypoxia (∼60%) and during 15 and 25% MVC (∼25-30%), and these impairments were abolished with fasudil. Similarly, [ATP]V and ATP effluent responses from normoxia to hypoxia and rest to 25% MVC were lower in OA vs. YA and improved with fasudil (P < 0.05). Isolated RBC ATP release during hypoxia was impaired in OA vs. YA (∼75%; P < 0.05), which tended to improve with fasudil in OA (P = 0.082). These data suggest Rho-kinase inhibition improves haemodynamic responses to hypoxia and moderate intensity exercise in OA, which may be due in part to improved circulating ATP. KEY POINTS: Skeletal muscle blood flow responses to hypoxia and exercise are impaired with age. Blunted increases in circulating ATP, a vasodilator, in older adults may contribute to age-related impairments in haemodynamics. Red blood cells (RBCs) are a primary source of circulating ATP, and treating isolated RBCs with a Rho-kinase inhibitor improves age-related impairments in deoxygenation-induced RBC ATP release. In this study, treating healthy older adults systemically with the Rho-kinase inhibitor fasudil improved blood flow and circulating ATP responses during hypoxia and moderate intensity handgrip exercise compared to young adults, and also tended to improve isolated RBC ATP release. Improved blood flow regulation with fasudil was also associated with increased skeletal muscle oxygen delivery during hypoxia and exercise in older adults. This is the first study to demonstrate that Rho-kinase inhibition can significantly improve age-related impairments in haemodynamic and circulating ATP responses to physiological stimuli, which may have therapeutic implications.

Endothelial dysfunction as a complication of anti-cancer therapy.

Recent strides in anti-cancer therapeutics have improved longevity and led to a growing population of cancer survivors, who are increasingly likely to die of other causes. Treatment-induced cardiotoxicity is a complication of several therapeutic agents with acute and long-term consequences for cancer patients. Vascular endothelial dysfunction is a precursor and hallmark of ischemic coronary disease and may play a role in anti-cancer therapy-induced cardiotoxicity. This review summarizes clinical evidence for endothelial dysfunction following anti-cancer therapy and extends the discussion to include the impact of therapeutic agents on conduit arteries and the microcirculation. We highlight the role of innate immune system activation and cross-talk between inflammation and oxidative stress as pathogenic mechanisms underlying anti-cancer therapy-induced vascular toxicity. Understanding the impact of anti-cancer agents on the vascular endothelium will inform therapeutic approaches to prevent or reverse treatment-induced cardiotoxicity and may serve as an important tool to predict, monitor, and prevent adverse cardiovascular outcomes in patients undergoing treatment.

ATP and acetylcholine interact to modulate vascular tone and α1-adrenergic vasoconstriction in humans.

The vascular endothelium senses and integrates numerous inputs to regulate vascular tone. Recent evidence reveals complex signal processing within the endothelium, yet little is known about how endothelium-dependent stimuli interact to regulate blood flow. We tested the hypothesis that combined stimulation of the endothelium with adenosine triphosphate (ATP) and acetylcholine (ACh) elicits greater vasodilation and attenuates α1-adrenergic vasoconstriction compared with combination of ATP or ACh with the endothelium-independent dilator sodium nitroprusside (SNP). We assessed forearm vascular conductance (FVC) in young adults (6 women, 7 men) during local intra-arterial infusion of ATP, ACh, or SNP alone and in the following combinations: ATP + ACh, SNP + ACh, and ATP + SNP, wherein the second dilator was coinfused after attaining steady state with the first dilator. By design, each dilator evoked a similar response when infused separately (ΔFVC, ATP: 48 ± 4; ACh: 57 ± 6; SNP: 53 ± 6 mL·min-1·100 mmHg-1; P ≥ 0.62). Combined infusion of the endothelium-dependent dilators evoked greater vasodilation than combination of either dilator with SNP (ΔFVC from first dilator, ATP + ACh: 45 ± 9 vs. SNP + ACh: 18 ± 7 and ATP + SNP: 26 ± 4 mL·min-1·100 mmHg-1, P < 0.05). Phenylephrine was subsequently infused to evaluate α1-adrenergic vasoconstriction. Phenylephrine elicited less vasoconstriction during infusion of ATP or ACh versus SNP (ΔFVC, -25 ± 3 and -29 ± 4 vs. -48 ± 3%; P < 0.05). The vasoconstrictor response to phenylephrine was further diminished during combined infusion of ATP + ACh (-13 ± 3%; P < 0.05 vs. ATP or ACh alone) and was less than that observed when either dilator was combined with SNP (SNP + ACh: -26 ± 3%; ATP + SNP: -31 ± 4%; both P < 0.05 vs. ATP + ACh). We conclude that endothelium-dependent agonists interact to elicit vasodilation and limit α1-adrenergic vasoconstriction in humans.NEW & NOTEWORTHY The results of this study highlight the vascular endothelium as a critical site for integration of vasomotor signals in humans. To our knowledge, this is the first study to demonstrate that combined stimulation of the endothelium with ATP and ACh results in enhanced vasodilation compared with combination of either ATP or ACh with an endothelium-independent dilator. Furthermore, we show that ATP and ACh interact to modulate α1-adrenergic vasoconstriction in human skeletal muscle in vivo.

Comprehensive assessment of cardiovascular structure and function and disease risk in middle-aged ultra-endurance athletes.

BACKGROUND AND AIMS: Recent studies suggest that long-term endurance training may be damaging to the heart, thus increasing cardiovascular disease (CVD) risk. However, studies utilizing cardiac imaging are conflicting and lack measures of central and peripheral vascular structure and function, which are also independently predictive of CVD events. METHODS: We performed a comprehensive assessment of cardiovascular structure and function in long-term (≥ 10 years) ultra-endurance athletes (ATH, 14 M/11 F, 50 ± 1 y) and physically active controls (CON, 9 M/9 F, 49 ± 2 y). RESULTS: As expected, left ventricular mass and end-diastolic volume (echocardiography) were greater in ATH vs CON, whereas there was no difference in cardiac function at rest. Coronary artery calcium scores (computed tomography) were not statistically different between groups. There was no evidence of myocardial fibrosis (contrast magnetic resonance imaging) in any subject. Aortic stiffness (carotid-femoral pulse wave velocity) was lower in ATH vs CON (6.2 ± 0.2 vs 6.9 ± 0.2 m/s, p < 0.05), whereas carotid intima-media thickness (ultrasound) was not different between groups. Peripheral vascular endothelial function (flow-mediated vasodilation of the brachial artery) and microvascular function (peak blood velocity) in response to 5 min of forearm ischemia were not different between groups. Furthermore, there was no difference in 10-year coronary heart disease risk (ATH; 2.3 ± 0.5 vs CON; 1.6 ± 0.2%, p > 0.05). CONCLUSIONS: Our data indicate that middle-aged ultra-endurance ATH do not have marked signs of widespread cardiovascular dysfunction or elevated CHD risk compared to CON meeting physical activity guidelines.

KIR channel activation links local vasodilatation with muscle fibre recruitment during exercise in humans.

KEY POINTS: During exercise, blood flow to working skeletal muscle increases in parallel with contractile activity such that oxygen delivery is sufficient to meet metabolic demand. K+ released from active skeletal muscle fibres could facilitate vasodilatation in proportion to the degree of muscle fibre recruitment. Once released, K+ stimulates inwardly rectifying K+ (KIR ) channels on the vasculature to elicit an increase in blood flow. In the present study, we demonstrate that KIR channels mediate the rapid vasodilatory response to an increase in exercise intensity. We also show that KIR channels augment vasodilatation during exercise which demands greater muscle fibre recruitment independent of the total amount of work performed. These results suggest that K+ plays a key role in coupling the magnitude of vasodilatation to the degree of contractile activity. Ultimately, the findings from this study help us understand the signalling mechanisms that regulate muscle blood flow in humans. ABSTRACT: Blood flow to active skeletal muscle is augmented with greater muscle fibre recruitment. We tested whether activation of inwardly rectifying potassium (KIR ) channels underlies vasodilatation with elevated muscle fibre recruitment when work rate is increased (Protocol 1) or held constant (Protocol 2). We assessed forearm vascular conductance (FVC) during rhythmic handgrip exercise under control conditions and during local inhibition of KIR channels (intra-arterial BaCl2 ). In Protocol 1, healthy volunteers performed mild handgrip exercise for 3 min, then transitioned to moderate intensity for 30 s. BaCl2 eliminated vasodilatation during the first contraction at the moderate workload (ΔFVC, BaCl2 : -1 ± 17 vs. control: 30 ± 28 ml min-1  100 mmHg-1 ; n = 9; P = 0.004) and attenuated the 30 s area under the curve by 56 ± 14% (n = 9; P < 0.0001). In Protocol 2, participants performed two exercise bouts in which muscle fibre recruitment was manipulated while total contractile work was held constant via reciprocal changes in contraction frequency: (1) low fibre recruitment, with contractions at 12.5% maximal voluntary contraction once every 4 s and (2) high fibre recruitment, with contractions at 25% maximal voluntary contraction once every 8 s. Under control conditions, steady-state FVC was augmented in high vs. low fibre recruitment (211 ± 90 vs. 166 ± 73 ml min-1 ⋅100 mmHg-1 ; n = 10; P = 0.0006), whereas BaCl2 abolished the difference between high and low fibre recruitment (134 ± 59 vs. 134 ± 63 ml min-1  100 mmHg-1 ; n = 10; P = 0.85). These findings demonstrate that KIR channel activation is a key mechanism linking local vasodilatation with muscle fibre recruitment during exercise.

Elevated extracellular potassium prior to muscle contraction reduces onset and steady-state exercise hyperemia in humans.

The increase in interstitial potassium (K+) during muscle contractions is thought to be a vasodilatory signal that contributes to exercise hyperemia. To determine the role of extracellular K+ in exercise hyperemia, we perfused skeletal muscle with K+ before contractions, such that the effect of any endogenously-released K+ would be minimized. We tested the hypothesis that local, intra-arterial infusion of potassium chloride (KCl) at rest would impair vasodilation in response to subsequent rhythmic handgrip exercise in humans. In 11 young adults, we determined forearm blood flow (FBF) (Doppler ultrasound) and forearm vascular conductance (FVC) (FBF/mean arterial pressure) during 4 min of rhythmic handgrip exercise at 10% of maximal voluntary contraction during 1) control conditions, 2) infusion of KCl before the initiation of exercise, and 3) infusion of sodium nitroprusside (SNP) as a control vasodilator. Infusion of KCl or SNP elevated resting FVC similarly before the onset of exercise (control: 39 ± 6 vs. KCl: 81 ± 12 and SNP: 82 ± 13 ml·min-1·100 mmHg-1; both P < 0.05 vs. control). Infusion of KCl at rest diminished the hyperemic (ΔFBF) and vasodilatory (ΔFVC) response to subsequent exercise by 22 ± 5% and 30 ± 5%, respectively (both P < 0.05 vs. control), whereas SNP did not affect the change in FBF ( P = 0.74 vs. control) or FVC ( P = 0.61 vs. control) from rest to steady-state exercise. These findings implicate the K+ ion as an essential vasodilator substance contributing to exercise hyperemia in humans. NEW & NOTEWORTHY Our findings support a significant and obligatory role for potassium signaling in the local vasodilatory and hyperemic response to exercise in humans.