Vasodilator Response Research Articles

Renal Dysfunction in Patients with Liver Cirrhosis

AbstractLiver cirrhosis is a major health problem that can affect people of different ages. It induces pivotal hemodynamic and metabolic systemic disturbances along with other organs dysfunction. Renal dysfunction in cirrhotic patients is not uncommon, and subtle renal impairment is an early and very frequent finding. Liver cirrhosis can afflict kidney functions through different mechanisms. Renal vasoconstriction is usually the initial response of splanchnic vasodilation and decreased effective renal plasma flow. This induces a reduction of intraglomerular pressure leading to stimulation of renin-angiotensin system to maintain the glomerular filtration rate. Other causes of renal dysfunction include electrolytes and acid-base disturbances, systemic inflammation, bile cast nephropathy, and intra-abdominal hypertension. Loss of renal reserve is usually the earliest manifestation of kidney dysfunction in cirrhotic patients. This makes the kidney supersensitive to any subsequent hemodynamic or metabolic abnormalities. Proper assessment of kidney function is one of the major challenges in cirrhotic patients. The use of serum creatinine and creatinine-based equations is inaccurate and can overestimate kidney function. Hepato-renal syndrome (HRS) is a life-threatening disorder. In the last decade, there was significant progress in understanding the mechanism of this mysterious disorder. In this article, we are focusing on different mechanisms of kidney dysfunction in cirrhotic patients and the major diagnostic and therapeutic challenges.

Persistent vascular dysfunction following an acute nonpharmacological reduction in blood pressure in hypertensive patients.

Vascular dysfunction, an independent risk factor for cardiovascular disease, often persists in patients with hypertension, despite improvements in blood pressure control induced by antihypertensive medications. As some of these medications may directly affect vascular function, this study sought to comprehensively examine the impact of reducing blood pressure, by a nonpharmacological approach (5 days of sodium restriction), on vascular function in 22 hypertensive individuals (14 men/8 women, 50 ± 10 years). Following a 2-week withdrawal of antihypertensive medications, two 5-day dietary phases, liberal sodium (liberal sodium, 200 mmol/day) followed by restricted sodium (restricted sodium, 10 mmol/day), were completed. Resting blood pressure was assessed and vascular function, at both the conduit and microvascular levels, was evaluated by brachial artery flow-mediated dilation (FMD), reactive hyperemia, progressive handgrip exercise, and passive leg movement (PLM). Despite a sodium restriction-induced fall in blood pressure (liberal sodium: 141 ± 14/85 ± 9; restricted sodium 124 ± 12/79 ± 9 mmHg, P < 0.01 for both SBP and DBP), FMD (liberal sodium: 4.6 ± 1.8%; restricted sodium: 5.1 ± 2.1%, P = 0.27), and reactive hyperemia (liberal sodium: 548 ± 201; restricted sodium: 615 ± 206 ml, P = 0.08) were not altered. Similarly, brachial artery vasodilation during handgrip exercise was not different between conditions (liberal sodium: Δ0.36 ± 0.19 mm; restricted sodium: Δ0.42 ± 0.18 mm, P = 0.16). Lastly, PLM-induced changes in peak blood flow (liberal sodium: 5.3 ± 2.5; restricted sodium: 5.8 ± 3.6 ml/min per mmHg, P = 0.30) and the total vasodilatory response [liberal sodium: 2 (0.9-2.5) vs. restricted sodium: 1.7 (1.1-2.6) ml/min per mmHg; P = 0.5] were also not different between conditions. Thus vascular dysfunction, at both the conduit and microvascular levels, persists in patients with hypertension even when blood pressure is acutely reduced by a nonpharmacological approach.

PERMISSIVE ROLE OF ANGIOTENSIN II IN MANTENANCE OF ENDOTHELIUM-DEPENDENT VASODILATION IN CEREBRAL MICROCIRCULATION

Objective: A number of data suggest, rather controversial, permissive role of angiotensin II (ANGII) in maintaining vasodilator responses in microcirculation, via its interaction with AT1 receptors (AT1R). This study aimed to determine the role of ANGII and AT1R in the mechanisms of flow-induced dilation (FID) in middle cerebral arteries (MCA) and incidence of oxidative stress at the functional, cellular and molecular level in the cerebral vasculature and in the serum of Sprague-Dawley (SD) rats fed high salt diet (HS). Design and method: Eleven-week old, male SD rats were randomly assigned to control group (CTRL, 0.4% NaCl in rat chow); Losartan group (rats on a standard diet given the AT1R blocker losartan (1 mg/mL) in drinking water); HS group (7 days 4% NaCl in rat chow) or HS+ANGII group (7 days HS with 3 days ANGII administration via osmotic minipumps (100 ng/kg/min on days 4–7)). FID of MCA was determined in absence/presence of the NOS inhibitor L-NAME, the non-selective cyclooxygenases inhibitor indomethacin, and the superoxide dismutase mimetic TEMPOL. Gene and protein expression of antioxidative enzymes and enzymes involved in FID mechanisms were determined by RT-qPCR and western blot. Vascular NO and superoxide/ROS levels were assessed by direct fluorescence. Serum systemic oxidative stress parameters was measured by spectrophotometry. All experimental procedures conformed to the EU Directive 2010/63/EU. Results: HS or AT1R blockade impaired FID and increased oxidative stress. Supplementation of ANGII restored the mechanisms of FID in MCA of rats fed HS diet to the ones present in CTRL rats without affecting arterial blood pressure by decreasing systemic oxidative stress and decreasing superoxide/ROS levels and increased NO bioavailability in the vascular wall of HS treated rats. The AT1R blockade resulted in significantly attenuated endothelium-dependent dilation, changed mechanisms of FID, increased vascular and systemic oxidative stress and decreased expression/ activity of antioxidative enzymes. Conclusions: ANGII is crucial in the preservation of vascular oxidative stress-antioxidant system balance and via AT1R, in maintenance of physiological vasodilation mechanisms of MCA. Suppression of ANGII is a key link between HS intake and development of endothelial dysfunction, and impaired regulation of cerebral blood flow.

Increased contribution of KCa channels to muscle contraction induced vascular and blood flow responses in sedentary and exercise trained ZFDM rats

In resistance arteries, endothelium-dependent hyperpolarization (EDH)-mediated vasodilatation is depressed in diabetes. We hypothesized that downregulation of KCa channel derived EDH reduces exercise-induced vasodilatation and blood flow redistribution in diabetes. To test this hypothesis, we evaluated vascular function in response to hindlimb muscle contraction, and the contribution of KCa channels in anaesthetised ZFDM, metabolic disease rats with type 2 diabetes. We also tested whether exercise training ameliorated the vascular response. Using in vivo microangiography, the hindlimb vasculature was visualized before and after rhythmic muscle contraction (0.5s tetanus every 3s, 20 times) evoked by sciatic nerve stimulation (40Hz). Femoral blood flow of the contracting hindlimb was simultaneously measured by an ultrasonic flowmeter. The contribution of KCa channels was investigated in the presence and absence of apamin and charybdotoxin. We found that vascular and blood flow responses to muscle contraction were significantly impaired at the level of small artery segments in ZFDM fa/fa rats compared to its lean control fa/+ rats. The contribution of KCa channels was also smaller in fa/fa than in fa/+ rats. Low-intensity exercise training for 12weeks in fa/fa rats demonstrated minor changes in the vascular and blood flow response to muscle contraction. However, the KCa-derived component in the response to muscle contraction was much greater in exercise trained than in sedentary fa/fa rats. These data suggest that exercise training increases the contribution of KCa channels among endothelium-dependent vasodilatory mechanisms to maintain vascular and blood flow responses to muscle contraction in this metabolic disease rat model. KEY POINTS: Microvascular dysfunction in type 2 diabetes impairs blood flow redistribution during exercise and limits the performance of skeletal muscle and may cause early fatigability. Endothelium-dependent hyperpolarization (EDH), which mediates vasodilatation in resistance arteries, is known to be depressed in animals with diabetes. Here, we report that low-intensity exercise training in ZFDM rats increased the KCa channel-derived component in the vasodilator responses to muscle contraction compared to that in sedentary rats, partly as a result of the increase in KCNN3 expression. These results suggest that low-intensity exercise training improves blood flow redistribution in contracting skeletal muscle in metabolic disease with diabetes via upregulation of EDH.

Effects of acute administration of trimethylamine N-oxide on endothelial function: a translational study

Elevated circulating levels of nutrient-derived trimethylamine N-oxide (TMAO) have been associated with the onset and progression of cardiovascular disease by promoting athero-thrombosis. However, in conditions like bariatric surgery (Roux-en-Y gastric bypass, RYGB), stable increases of plasma TMAO are associated with improved endothelial function and reduced cardiovascular morbidity and mortality, thus questioning whether a mechanistic relationship between TMAO and endothelial dysfunction exists. Herein, we translationally assessed the effects of acute TMAO exposure on endothelial dysfunction, thrombosis and stroke. After RYGB, fasting circulating levels of TMAO increased in patients and obese rats, in parallel with an improved gluco-lipid profile and higher circulating bile acids. The latter enhanced FXR-dependent signalling in rat livers, which may lead to higher TMAO synthesis post RYGB. In lean rats, acute TMAO injection (7 mg kg−1) 1.5-h before sacrifice and ex-vivo 30-min incubation of thoracic aortas with 10−6 M TMAO did not impair vasodilation in response to acetylcholine (Ach), glucagon-like peptide 1, or insulin. Similarly, in lean WT mice (n = 5–6), TMAO injection prior to subjecting mice to ischemic stroke or arterial thrombosis did not increase its severity compared to vehicle treated mice. Endothelial nitric oxide synthase (eNOS) activity and intracellular stress-activated pathways remained unaltered in aorta of TMAO-injected rats, as assessed by Western Blot. Pre-incubation of human aortic endothelial cells with TMAO (10−6 M) did not alter NO release in response to Ach. Our results indicate that increased plasmatic TMAO in the near-physiological range seems to be a neutral bystander to vascular function as translationally seen in patients after bariatric surgery or in healthy lean rodent models and in endothelial cells exposed acutely to TMAO.

The superoxide radical switch in the biology of nitric oxide and peroxynitrite.

The free radical nitric oxide (·NO) is a key mediator in different physiological processes such as vasodilation, neurotransmission, inflammation, and cellular immune responses, and thus preserving its bioavailability is essential. In several disease conditions, superoxide radical (O2·-) production increases and leads to the rapid "inactivation" of ·NO by a diffusion-controlled radical termination reaction that yields a potent and short-lived oxidant, peroxynitrite. This reaction not only limits ·NO bioavailability for physiological signal transduction but also can divert and switch the biochemistry of ·NO toward nitrooxidative processes. Indeed, since the early 1990s peroxynitrite (and its secondary derived species) has been linked to the establishment and progression of different acute and chronic human diseases and also to the normal aging process. Here, we revisit an earlier and classical review on the role of peroxynitrite in human physiology and pathology (Pacher P, Beckman J, Liaudet L. Physiol Rev 87: 315-424, 2007) and further integrate, update, and interpret the accumulated evidence over 30 years of research. Innovative tools and approaches for the detection, quantitation, and sub- or extracellular mapping of peroxynitrite and its secondary products (e.g., protein 3-nitrotyrosine) have allowed us to unambiguously connect the complex biochemistry of peroxynitrite with numerous biological outcomes at the physiological and pathological levels. Furthermore, our current knowledge of the ·NO/O2·- and peroxynitrite interplay at the cell, tissue, and organ levels is assisting in the discovery of therapeutic interventions for a variety of human diseases.

Parabiosis Improves Endothelial Dysfunction in Aged Female Mice

IntroductionThe study aims to investigate the effect of parabiosis method on endothelial dysfunction in naturally aging mice and determine the time projections for predicted improvement in the mentioned target group. MethodsThe balb/c mice were separated into six groups, these being; isochronic old, heterochronic old (HP-O), isochronic young, heterochronic young, young control, and old control. After parabiosis protocol, animals were sacrificed at the third, fifth, seventh, and ninth weeks, and their thoracic aortas were isolated. The vasodilatation and vasoconstriction responses of the vessels were detected using potassium chloride and phenylephrine, acetylcholine (ACh), and sodium nitroprusside. ResultsAging had a significant decreasing effect on maximum ACh relaxation responses (P < 0.01). However, in the HP-O group, the maximum ACh relaxation response in the third week was significantly lower (P < 0.05), but this effect disappeared in the ninth week. Maximum phenylephrine contraction responses were lower in the heterochronic parabiosis group (P < 0.05). ConclusionsACh responses increased at the end of the ninth week in the HP-O group, therefore, the parabiosis model may have an improving effect on endothelial dysfunction seen in aging.

The Vasodilatory Response to CGRP of the Anterior and Posterior Cerebral Circulation in Migraine.

IntroductionMigraine aura can be associated with headache or it may occur without one, which suggests an independent mechanism for the aura and for migraine headache. The role of CGRP in migraine headache is well established, but the connection between CGRP and the aura is still lacking an explanation. Exogenous CGRP can induce CGRP headaches and migraine auras in patients with migraine. The results of our recent study suggest differences in the vascular response to CGRP stimulation between migraine without aura and migraine with aura. Therefore, we hypothesized that the magnitude of the posterior cerebral circulation response in migraine with aura is greater than in migraine without aura and that CGRP stimulation has different effects on the anterior and posterior circulation in migraine with aura and migraine without aura.MethodsBy using transcranial doppler, we studied the hemodynamic effects of CGRP intravenous infusion at a rate of 1.5 mcg/min in 20 min on the mean arterial velocity in the middle cerebral artery and in the posterior cerebral artery in twenty patients with migraine and in a control group of twenty healthy subjects. The same CGRP effects on cerebral hemodynamics were analyzed separately for the group of patients with migraine with aura and the group of patients with migraine without aura. Fifteen patients with migraine (75%) had migraine without aura and 5 patients (25%) had migraine with aura.ResultsWe found that migraine has a significant impact on the vasodilatory response of the anterior (B = 4,249, SE = 1.023, r = 0.363, p < 0.001) and posterior cerebral circulation (B = 3.634, SE = 1.461, r = 0.227, p = 0.014). Migraine with aura was significantly associated with changes in the anterior (B = 2.558, SE = 0.880, r = 0.275, p = 0.005) and posterior cerebral circulation (B = 7.565, SE = 2,368, r = 0.359, p = 0.002), while migraine without aura was only significantly associated with changes in the anterior circulation. In addition, we established a significant impact of migraine with aura on VR PCA (B = 5.901, SE = 2,546, r = 0.291, p = 0.024).ConclusionWe conclude that TVR in the posterior cerebral circulation might be enhanced in MA and that aura might be a consequence of TVR enhancement.

On inlet pressure boundary conditions for CT-based computation of fractional flow reserve: clinical measurement of aortic pressure

Background and objectives A quick calculation approach of steady-state fractional flow reserve (FFRss) based on computed tomography angiography (CTA) images is a reliable non-invasive way of calculate FFR, the assumptions used in the research should be study further to increase forecast accuracy. The effect of inlet and outlet boundary conditions on FFRss was investigated. Methods 15 patients who had been diagnosed with coronary artery disease were enrolled in this study. We investigated the sensitivity of calculating FFR to boundary circumstances, using invasive FFR as a benchmark. There are two types of inlet: (1) aortic pressure based on clinically measured. (2) mean pressure calculated based on physiological formula; we further studied the outlet changes of FFRss under different coronary vasodilation responses (24%, 48%, 72%). Results According to the calculate FFR results of all patients, FFRSST (based on the clinical experiment) and FFRSSM (based on the physiological formula) {r = 0.99, [95% confidence interval (CI):0.0.94 to 1.14] (p < 0.001)}. Although the pressure difference between the two pressure boundary conditions is 15 mmHg, the calculated FFR result does not change significantly. The microcirculation resistance of the outlet gradually rose as the vasodilation state changed, and the computed FFR increased. Conclusions A numerical analysis of the effects of proximal and distal boundary constraints of computational models on computed CT-FFR is presented. The findings revealed that distal boundary circumstances (hyperemic vasodilation response of coronary micro-vessels) have a significant impact on FFR, providing evidence to guide the development and application of a computational model for estimating FFR.

Features of vascular motor endothelial function in hypertensive patients with chronic heart failure

Abstract Funding Acknowledgements Type of funding sources: None. The main part of the pathogenesis of arterial hypertension (AH) is a remodeling of the vascular system and the state of the vascular endothelium, which functions as a trigger mechanism for the implementation vasoregulating mechanisms. Vascular endothelium is involved in the provision of local processes of hemostasis, proliferation, migration of blood cells in the vessel wall, which is also associated with the regulation of vascular tone and the development of atherosclerosis. Purpose To study the vascular motor endothelial function in hypertensive patients with chronic heart failure (CHF). Methods Vascular mechanisms of pathogenesis have been studied in 172 patients aged (54,2±6,2) years, men 84 and women 88, with uncomplicated hypertension II-III degree. Echographic system Acuson Sequoia 512 was used for ultrasound imaging of the heart. The diameter of the brachial artery (D, mm), flow velocity (V, m/s), the ratio D/V, Δd (%) - an increase in the diameter of the artery decompression, ΔdN (%) - an increase in the diameter of the artery sample with nitroglycerin and the ratio Δd/ΔdN were measured. Results Due to the reduction in brachial artery diameter (with AH stage II 55, l% and AH stage III 48.7%, p &amp;lt;0.01) decreased blood flow velocity in the whole group is 36.4%, which is as much depended on the degree of hypertension (p &amp;lt;0.01). Conducting tests for decompression and nitroglycerin showed that reduced growth in brachial artery diameter in the total group on the decompression was compared with the normal 51.1%: in AH II degree - by 48.8% and III level by 52.2% (p&amp;lt;0.01). The test with nitroglycerin: the larger group of hypertensive patients reduced vasodilation was 21.1%, II degree - 13.8%, and III degree - 30.7% (p&amp;lt;0.01). Conducting tests for decompression showed that normal type vasodilatory response occurred only in 27.4% of patients, with II degree in 33.8% and AH III in 22% of patients. Reduced response to decompression was 60% of patients with AH II - in 43.2%, and III degree - at 56.8% and constrictor - in 12.6% of them in AH II degree at - 6.2% and 17 % at AH III. The test with nitroglycerin: normal vascular reaction occurred in 33.5% of patients, decreased in 59.9% and constrictor in 9.6% of patients. Reduced vasoconstrictor response to nitroglycerin was more common in patients with AH II degree. Cunclusion These results define the nature of the preferred choice of antihypertensive drugs such as normalizing pressure and have a positive effect on the endothelium, which is also associated with a reduction of myocardial hypertrophy and vascular wall.

Integrated redox sensitivity of vascular Kv1 channels

Voltage‐gated potassium (Kv) channels in vascular smooth muscle are responsive to a range of molecular signals that link local metabolic demand to blood supply. We recently found that elevation of local lactate:pyruvate, reflective of enhanced tissue oxygen demand and consequent changes in smooth muscle cytosolic NADH:NAD+, potentiate native Kv1 activity in a manner that is dependent on pore complex‐associated Kvβ proteins (i.e., Kvβ1 and Kvβ2). Considering that these proteins also participate in the regulation of Kv function in response to oxidants, we tested the hypothesis that Kv1‐dependent vasoreactivity to the endogenous dilator hydrogen peroxide (H2O2) is modified by intracellular NAD(H) redox status. Application of H2O2 to preconstricted (100 nM U46619, 80 mmHg intravascular pressure) mesenteric arteries from mice resulted in concentration‐dependent vasodilation (0.1‐10 μΜ). However, the magnitude of vasodilation in response to 10 μM H2O2 was significantly enhanced when applied in the presence of 10 mM L‐lactate (10 μΜ Η2Ο2: 16 ± 9%; 10 mM L‐lactate + 10 μM H2O2: 80 ± 15% dilation), but not in the presence of 10 mM pyruvate (~11% H2O2‐induced constriction). Consistent with direct effects of pyridine nucleotide redox state on oxidant‐induced Kv potentiation, increases in single Kv1 channel open probability in excised membrane patches from isolated vascular smooth muscle cells in response to 10 μM H2O2 were significantly enhanced in the presence of 1 mM NADH. Based on these results, we examined the role of the intracellular Kvβ subunit complex in redox‐modified vasodilation in response to H2O2. Arteries isolated from double transgenic mice with doxycycline‐inducible increases in smooth muscle Kvβ1:Kvβ2 protein ratio exhibited a complete loss of L‐lactate‐induced enhancement of vasodilation to H2O2. Together, our results suggest that the Kvβ subunit complex in vascular smooth muscle may integrate multiple redox signals to enable tight control over Kv1 gating, vascular tone, and blood flow upon dynamic changes in metabolic demand.

Estradiol Augments Sensory Nerve‐Mediated Vasodilation in Young Women

Local heating of the skin elicits biphasic vasodilation: a sensory nerve‐mediated initial peak (axon reflex) and an endothelial‐dependent plateau. We have previously shown that sex hormones, particularly estradiol (E2), improves cutaneous endothelial‐dependent vasodilation, but less is known about the effects of E2 on cutaneous sensory nerve‐mediated vasodilation.PURPOSEWe tested the hypothesis that E2 enhances sensory‐nerve mediated vasodilation in response to local heating of the skin in young women.METHODSWe retrospectively analyzed data in 15 healthy young women (age 24 ± 4 yr; MAP 84 ± 6 mmHg; BMI 24 ± 3 kg/m2). Endogenous sex hormone production was suppressed with daily administration of a gonadotropin‐releasing hormone antagonist (GnRHant; Ganirelix) for 10 days. E2 (0.1 mg/day, Vivelle‐Dot) was added back via transdermal patch on days 4–10. Cutaneous vasodilatory responses to local heating were measured on days 4 and 10 using laser Doppler flowmetry during microdialysis perfusions of lactated Ringer’s (control), and in a subset of women (n=6), L‐NAME (non‐selective nitric oxide synthase inhibition). Cutaneous vascular conductance (CVC) was calculated during the initial peak of the axon reflex during local heating (42°C) and normalized to maximal vasodilation achieved by perfusion of sodium nitroprusside (28mM) and heating to 43°C.RESULTSSensory nerve‐mediated vasodilation increased during E2 administration compared to GnRHant (E2: 71 ± 15, GnRHant: 57 ± 18 %CVCmax; p=0.046). Blockade of nitric oxide with L‐NAME did not blunt the axon reflex during GnRHant (control: 57 ± 18 vs. L‐NAME: 50 ± 17 %CVCmax; p=0.401) or E2 administration (control: 71 ± 15 vs. L‐NAME: 52 ± 19 %CVCmax; p=0.152).CONCLUSIONThese preliminary findings suggest that E2 enhances sensory nerve‐mediated vasodilation in young women, and that this may occur through mechanisms independent of nitric oxide. Future mechanistic studies isolating the nitric oxide component and other pathways could help elucidate the role of E2 in sensory nerve‐mediated vasodilation.

Nicotine Alters cAMP Signaling in Arterial Myocytes

Arterial myocytes receive many inputs from sympathetic nerves that regulate vascular reactivity by activating GS protein‐coupled receptors (GSPCRs). The ubiquitous second messenger 3′,5′‐ cyclic adenosine monophosphate (cAMP) is synthesized by adenylyl cyclases (ACs) in response to the activation of various GSPCRs and plays a critical role in integrating the myriad of signals elicited. Importantly, arterial myocyte cAMP signaling has been traditionally linked with relaxation. We found that arterial myocytes from mice exposed to secondhand smoke (SHS) had an attenuated cAMP synthesis in response to the β‐adrenergic receptor (βAR) agonist isoproterenol (ISO) (unpublished data), which could alter vascular reactivity. Tobacco smoke contains over 7,000 chemicals, including nicotine. Nicotine, the primary addictive component in tobacco cigarettes is a strong sympathomimetic with significant vasoactive effects and have been shown to alter cellular cAMP production. However, how nicotine impacts cAMP signaling in the vasculature, particularly in arterial myocytes, and the implications of these changes in vascular reactivity are unclear. We are tackling this knowledge gap by expressing a FRET‐based cAMP biosensor in cultured arterial myocytes to determine cAMP signaling in response to nicotine/cotinine. We also developed a mouse expressing a new cAMP sensor specifically in SM (CamperSM) to investigate cAMP dynamics in native VSM/arteries. We hypothesize that nicotine impairs receptor‐mediated cAMP signaling in arterial myocytes leading to altered vascular reactivity. Our data using unpassaged arterial myocytes expressing a cytosolic FRET‐based cAMP sensor exposed to cotinine (the primary metabolite of nicotine) show a reduction in ISOinduced cAMP production. Similar results were observed in arterial myocytes from CAMPERSM mice infused with nicotine. These results suggest similar effects of cotinine in vitro or nicotine in vivo on βAR‐mediated cAMP synthesis in arterial myocytes. Intriguingly, we found that inhibition of phosphodiesterases rescue cAMP synthesis in CAMPERSM arterial myocytes exposed to nicotine. We correlated the nicotine‐induced reduction in arterial myocytes cAMP synthesis with impaired vasodilation in response to ISO. Our data suggest that nicotine impairs receptormediated cAMP signaling in arterial myocytes leading to altered vascular reactivity. Nicotineinduced cAMP dysregulation leading to altered arterial myocytes excitability could be an underlying factor contributing to cardiovascular disease in individuals exposed to SHS and users of nicotine containing products.

Rho‐related BTB Domain‐containing Protein 1 Reverses Established Arterial Stiffness in Angiotensin II‐treated Mice

Rho‐related BTB domain‐containing protein 1 (RhoBTB1) is associated with blood pressure in GWAS studies, but its function is not well‐understood. The level of RhoBTB1 mRNA was decreased in mice treated with angiotensin II (Ang‐II) concomitant with hypertension, arterial stiffness, and impaired vasodilation. Here, we tested the hypothesis that restoration of RhoBTB1 would reverse hypertension, arterial stiffness, and improve vasodilation response caused by Ang‐II infusion. Genetic complementation of RhoBTB1 in the vasculature was achieved by expressing a smooth muscle‐specific, tamoxifen‐inducible RhoBTB1 transgene (S‐RhoBTB1 mice). Mice expressing smooth muscle‐specific, tamoxifen‐inducible Cre‐recombinase without RhoBTB1 (ISM‐Cre mice) were used as genotype control. ISM‐Cre and S‐RhoBTB1 mice received 6 weeks of Ang‐II infusion, but their transgene was not activated until two weeks after the initiation of Ang‐II treatment. Meanwhile, vehicle‐treated ISM‐Cre and S‐RhoBTB1 mice were used as control. Blood pressure was continuously recorded by radiotelemetry during the entire protocol. Pulse wave velocity (PWV) and aortic compliance assay were used to evaluate arterial stiffness both during and at the end of the protocol. Vasodilation was measured in isolated carotid and mesenteric arteries using wire myography. In vehicle‐treated mice, aortic RhoBTB1 expression was increased in S‐RhoBTB1 mice compared to ISM‐Cre mice. Ang‐II significantly decreased the level of RhoBTB1 in the aorta of ISM‐Cre mice whereas S‐RhoBTB1 mice normalized this effect. At baseline, overexpressing RhoBTB1 in the vasculature had no significant effect on any phenotype. Before transgene activation, Ang‐II increased blood pressure, arterial stiffness, and impaired vasodilation in response to acetylcholine and sodium nitroprusside in both ISM‐Cre and S‐RhoBTB1 mice. Remarkably, in Ang‐II‐treated mice, RhoBTB1 restoration restored arterial stiffness nearly back to baseline without rescuing either hypertension or vasodilation. Since arterial stiffness is often observed to go hand‐in‐hand with hypertension, the current phenotype is particularly interesting as it provides novel insight into the cause‐effect relationship between arterial stiffness and hypertension. Next, we utilized RNA‐seq and biochemical assays to investigate the mechanism by which RhoBTB1 reversed arterial stiffness. Classical contributors of arterial stiffness, for instance, collagen, elastin, and vascular smooth muscle remodeling, were not altered by RhoBTB1 restoration. However, actin polymerization was elevated by Ang‐II and normalized upon RhoBTB1 restoration, as evident by both immunoblotting and fluorescent labeling. In conclusion, we identified a novel function of RhoBTB1 and showed its importance in protecting from and reversing arterial stiffness.

Stratification by Race Reveals Disparate Vascular Toxicity in Response to Anti‐Cancer Therapies

IntroductionCardiovascular toxicity is the primary reason for halting anti‐cancer therapy (CTx) or limiting the dose, and cardiovascular side effects of CTx represent a major clinical problem with implications for survival and quality of life. Ongoing work in our laboratory suggests that CTx induces microvascular endothelial dysfunction, which is a well‐established predictor of future CV events. CV disease in general and CTx‐induced adverse events are significantly more prevalent in Black patients (B) relative to non‐Hispanic White patients (NHW). Despite clinical evidence of these health disparities, little to no data exist to explain the underlying causes. Therefore, we tested the novel hypothesis that CTx‐induced microvascular dysfunction is augmented in B, which may explain the increase in cardiotoxic events in this population compared to NHW.MethodsFresh human tissues were obtained as de‐identified surgical discard specimens. Peripheral adipose arterioles (100–200 µm) from healthy B and NHW were incubated ex vivowith the CTx agents, doxorubicin (Dox), trastuzumab (TZM), or paclitaxel (PTX) for 16–20 h and used for isolated arteriole preparations. Endothelium‐dependent vasodilation in response to flow (Flow‐Mediated Dilation, FMD; 5–100 cmH2O pressure gradient) or acetylcholine (ACh; 10−9 to 10−5 M) and smooth muscle‐dependent dilation to papaverine (100 µM) were evaluated. Data are presented as % max. diameter ± SEM, N. Statistical significance was established with two‐way ANOVA RM.ResultsUnder control conditions, there was no difference in microvascular FMD in healthy B (ACh: 89.8±4.0%, n=5; 85.8±1.8%, n=6) vs. NHW (ACh: 86.2±3.2%, n=6; 78.7±5.9%, n=8). Ex vivo exposure to Dox impaired endothelial function in arterioles from both B and NHW (ACh: 45.0±10.5%, n=4 and 34.2±9.8%, n=7; FMD: Figure 1A). TZM impaired endothelial function to a greater extent in arterioles from B (ACh: 36.1±7.1%, n=3; FMD: Figure 1B) compared to NHW (ACh: 44.4±8.6%, n=8; FMD: Figure 1B). Interestingly, PTX did not affect endothelial function in NHW (ACh: 91.5±1.6%, n=4; FMD: Figure 1C), but impaired endothelial function in arterioles from B (ACh: 54.7±12.9%, n=3; FMD: Figure 1C). Max dilator capacity to the endothelium‐independent dilator papaverine was not changed in either group (not shown).ConclusionOur findings reveal disparate effects of CTx agents on microvascular endothelial function from B and NHW patients. This raises the possibility that microvascular dysfunction may contribute to the known disparities in adverse CV clinical outcomes of cancer patients and potentially other CV pathologies (e.g., disparities in adverse events associated with PTX‐eluting stents.) Future work should differentiate between biological/genetic vs. social/environmental contribution to observed microvascular responses to CTx.

Effect of Short‐Term Estradiol Exposure on Endothelin‐B Receptor‐Mediated Vasodilation in Postmenopausal Women

Endothelin (ET)‐1 contributes to endothelial dysfunction in postmenopausal women (PMW) via the ETB receptor. We have previously shown that the ETB receptor mediates vasodilation in young women, an effect that is lost in PMW. However, it is unclear if ETB receptor‐mediated dilation can be restored in PMW exposed to E2.PURPOSETo test the hypothesis that E2 exposure restores ETB receptor‐mediated vasodilation in PMW.METHODSWe tested 9 PMW (55±2 yrs, 24±3 kg/m2, 91±5 mmHg) who self‐reported natural menopause and were at least 1 year without menses. We measured vasodilation in the cutaneous microcirculation via local heating using laser Doppler flowmetry coupled with intradermal microdialysis during perfusions of lactated Ringer’s (control), ETB receptor antagonist (BQ‐788, 300nM) and ETA receptor antagonist (BQ‐123, 500nM). Cutaneous vascular conductance (CVC) was calculated during the plateau phase of local heating (42°C), and normalized to maximal vasodilation achieved by perfusion of sodium nitroprusside (28mM) and heating to 43°C. All women were tested at baseline (BL) and after 7 days of E2 (Vivelle‐Dot, 0.1mg/day, transdermal patch) administration (+E2). A two‐way repeated measures ANOVA was performed to compare %CVCmax in PMW before and after E2 exposure. Data presented are mean ± standard deviation.RESULTSE2 administration increased serum estradiol (BL: 45±29 vs +E2: 71±42 pg/mL, P&lt;0.01) and reduced follicle stimulating hormone (BL: 71±13 vs +E2: 57±17 pg/mL, P&lt;0.01), but there were no changes in serum progesterone (BL: 0.57±0.93 vs +E2: 0.53±0.88 pg/mL, P=0.14) nor plasma ET‐1 (BL: 1.62±0.66 vs +E2: 1.54±0.54 pg/mL, P=0.42). E2 administration had no effect on cutaneous microvascular responses (Time P=0.41, Drug P=0.17, Interaction P=0.71). At BL, vasodilatory responses to local heating were not different during control and ETB receptor antagonism (control: 83±8 vs. BQ‐788: 88±6 %CVCmax). Similarly, vasodilation was not altered at BL during ETA receptor antagonism in a subset of women (n=6, control: 83±8 vs. BQ‐123: 87±8 %CVCmax). E2 administration did not improve vasodilation in the control site (BL: 83±8 vs. +E2 85±13 %CVCmax). Furthermore, there was no effect of E2 exposure on cutaneous vasodilatory responses during perfusion of ETB (BL: 88±6 vs. +E2: 89±6 %CVCmax) nor ETA (n=6, BL: 87±8 vs +E2: 92±7 %CVCmax) receptor antagonists.CONCLUSIONThese preliminary data suggest that short‐term E2 exposure does not modulate ETB receptor function in PMW. These findings are in contrast to our observations in young women demonstrating that ETB receptors mediate vasodilation in the presence of E2. The role of alternative therapies to modulate ETB receptor function should be considered in future studies.

Calcium‐activated Chloride Channel TMEM16A/ANO1 Does Not Mediate the Regulation of Sweating and Cutaneous Vasodilation in Humans In Vivo

The regulation of human body temperature during a heat stress is dependent on the activation of the heat loss responses of cutaneous vasodilation and eccrine sweating. While significant advancements have been made in our understanding of the mechanisms governing their regulation, new potential modifiers such as transmembrane member 16A (TMEM16A), a Ca2+‐activated Cl‐ channel also known as anoctamin 1 (ANO1), continue to be identified. In vitro and animal studies suggest a role for TMEM16A in the regulation of sweat secretion and vascular tone. For example, Ca2+ induced Cl‐current was abolished in TMEM16A deficit, but not BEST2 deficit, NCL‐SG3 human eccrine sweat gland cell lines. Further, animal models showed the activation of TMEM16A on vascular smooth muscle cells blunts vasodilation. However, direct evidence supporting a role of TMEM16 in mediating cutaneous vasodilation as well as eccrine sweating in humans in vivo remains to be established. Therefore, we evaluated the hypothesis that TMEM16A mediates the regulation of sweating and cutaneous vasodilation during a whole‐body heat stress in young adults.Twelve young (24 ± 2 years) adults (6 females) underwent a whole‐body heat stress induced by a water‐perfused suit to raise core temperature 1.1±0.1ºC above baseline resting levels. Sweat rate (ventilated capsule) and cutaneous vascular conductance (assessed as skin blood flow/mean arterial pressure and normalised to maximal conductance achieved via simultaneous local skin heating to 44°C and administration of sodium nitroprusside) were assessed at four skin sites on the forearm treated with 1) lactated Ringers (control), 2) 5 % dimethyl sulfoxide serving as a vehicle control, 3) 1 mM T16Ainh‐A01 (a TMEM16A blocker) or 4) 2 mM benzbromarone (another TMEM16A blocker). All drugs were administered continuously via intradermal microdialysis.Sweat rate increased as heating continued at all skin sites (P &lt; 0.001 for a main effect of heating stage). Neither the main effect of treatment site nor the interaction between treatment site and heating stage was significant (both P ≥ 0.318). Cutaneous vascular conductance increased in parallel to elevations in core temperature (P &lt; 0.001 for a main effect of heating stage). Although no main effect of treatment site was observed (P = 0.109), a significant interaction between heating stage and treatment site was measured (P &lt; 0.001). Post‐hoc analysis revealed that cutaneous vascular conductance at the DMSO‐treated site was higher than the control site at baseline (P = 0.047). By contrast, cutaneous vascular conductance at moderate elevations in core temperature (i.e., ≥0.4‐0.8 °C) was greater at the control site relative to the DMSO‐treated site (all P ≤ 0.013). Importantly, no differences were measured in cutaneous vascular conductance between the DMSO‐treated site and TMEM16A blocker‐treated sites (all P ≥ 0.073).Taken together, our findings demonstrate for the first time that TMEM16A does not contribute to the regulation of sweating and cutaneous vasodilation during whole‐body heating in young adults in vivo.

Renal Vascular Resistance Does Not Differ Between Younger Men and Women

PurposeThe incidence of acute kidney injury is nearly two‐fold higher risk in men compared to women. A common cause of acute kidney injury is supply‐demand mismatch that is characterized by the presence of a comparatively low renal oxygen supply (i.e., blood flow) relative to tissue oxygen demand. Thus, it is possible that the control of renal blood flow differs between men and women. For instance, the vasodilator actions of estrogen may protect against excessive renal vasoconstriction or more readily promote renal vasodilation in pre‐menopausal women compared to men. However, whether renal vascular control, a function of both vasoconstrictor and vasodilator capacities, differs between men and women is unknown. Therefore, this study tested the hypotheses that (1) renal vasoconstriction during a sympathetic stimulus (i.e., the cold pressor test, CPT) will be greater in young healthy men compared to young healthy women, and (2) renal vasodilation following an oral protein load will be greater in women compared to men.MethodsHealthy, men (n=13, age 26 ± 3 years) and eumenorrheic women (n=12, age 23 ± 3 years, 4 ± 1 day of menstrual cycle) completed a 2‐min CPT after one hour of supine rest to assess renal vasoconstriction. A whey protein shake (1.2 g of protein/kg of body weight) was ingested 1‐h post‐CPT to assess renal vasodilation. Blood pressure (Penaz method) was measured continually. Renal segmental artery blood velocity (BV, Doppler ultrasound) was measured before and during the CPT (1‐ and 2‐min), and pre‐protein and 75‐ and 150‐min post‐protein. Segmental artery vascular resistance (VR) was calculated as mean arterial pressure (MAP) divided by segmental BV. Data are presented as mean ± SD.ResultsMAP increased in both men and women from pre‐CPT to 2 min (p&lt;0.001), and the magnitude of increase did not differ between men (peak: +25 ± 13 mmHg) and women (peak: +23 ± 9 mmHg, p=0.667). Peak reductions in segmental BV (men: ‐1.3 ± 3.2 cm/s, women: ‐3.7 ± 2.9 cm/s, p=0.065) and increases in segmental VR (men: +1.5 ± 1.0 mmHg/cm/s, women: +2.1 ± 1.0 mmHg/cm/s, p=0.155) during the CPT did not differ between men and women. MAP did not differ between men and women pre‐protein or at 75‐min post‐protein (p&gt;0.586) but was higher in men at 150‐min post‐protein (86 ± 7 mmHg vs. 78 ± 8 mmHg, p=0.042). Peak increases in segmental BV following protein ingestion were greater in women (+5.3 ± 2.2 cm/s) compared to men (+2.7 ± 2.4 cm/s, p=0.009). Similarly, peak reductions in segmental VR were greater in women (‐1.1 ± 0.6 mmHg/cm/s) compared to men (‐0.6 ± 0.6 mmHg/cm/s, p=0.030) following protein ingestion.ConclusionRenal vasoconstrictor responses to sympathetic stimulation do not appear to differ between healthy men and women. However, women displayed greater renal vasodilatory responses to an oral protein load. Therefore, renal vascular control differs between healthy men and women tested in the early follicular phase of the menstrual cycle. These findings may provide mechanistic insights in support of observations that acute kidney injury incidence differs between men and women.

Role of Potassium Channel Function and Long‐term Exercise Training in Soleus Resistance Arterioles

IntroductionNuclear factor (erythroid‐derived 2)‐like 2 (Nrf2) expression has been linked to potassium (K+) channel function. We hypothesized that myogenic responses are enhanced by exercise training and also modulated by K+ channel function but would be impaired in arterioles from skeletal muscle of trained rats in which Nrf2 (Nrf2 KO) is deleted.MethodsExercise training was performed in male wildtype (WT) Sprague‐Dawley rats, and their littermates lacking the Nrf2 gene (Nrf2 KO). Rats were exercise trained using a mixed moderate and high‐intensity interval treadmill program for 10‐12 weeks. A WT cage‐confined control group was also studied to compare the impact of exercise training. After exercise training or sedentary cage‐confinement, soleus resistance arterioles were isolated and cannulated for the assessment of vascular reactivity. Vasodilatory responses to low concentrations of KCl [10 and 20mM} were assessed. Vasoconstrictor responses to higher concentrations of KCl [30–100mM] and myogenic responses to increasing intraluminal pressure (pressure 0cm H2O – 130cm H2O) were also assessed. The contribution of inward rectifying K+ channels (Kir) to vasodilation during low concentrations of KCl was assessed by treatment with barium chloride (BaCl2, 100mM).ResultsSoleus resistance arteriole myogenic responsiveness was reduced in trained Nrf2 KO as compared to WT (p&lt;0.01). Trained WT rats produced a significantly higher myogenic responsiveness than sedentary WT controls (p&lt;0.05). Vasodilation to low concentrations of KCl was reduced in soleus arterioles from trained Nrf2 KO rats as compared to arterioles from trained WT rats (p&lt;0.01). Exercise training did not alter vasodilation to low KCl concentrations in WT rats (p&gt;0.05). Constriction to higher concentrations of KCl was similar in arterioles from trained WT and Nrf2 KO rats. Inhibiting Kir channels with BaCl2 eliminated the genotype‐ and training‐related differences in myogenic responses and vasodilation to KCl.ConclusionOur data indicate that Nrf2 may be a critical regulator of vascular adaptation that occurs in skeletal muscle arterioles in response to long‐term exercise training. Elimination of differences in myogenic responsiveness by BaCl2 suggest that Nrf2‐mediated adaptation of the myogenic responsiveness may involve Kir channels.

Endothelial KIR channel dysfunction in aged cerebral parenchymal arterioles

Aging is associated with cognitive decline via mechanisms that are not fully understood. Endothelial dysfunction is known to occur in parenchymal arterioles, the bottlenecks of the cerebral microcirculation. Dilation of these arterioles is highly dependent on changes in extracellular K+ ([K+]E), caused by activity of inwardly rectifying K+ channels (KIR2) or by opening of Ca2+‐activated small‐ and intermediate‐conductance K+ channels (KCa2.3 / KCa3.1, respectively). The effects of aging on activity of these channels in the endothelium of cerebral parenchymal arterioles remain undetermined, as well as their impact on endothelium‐dependent dilation. Thus, we hypothesized that advanced age will impair both KIR2 and KCa2.3 / KCa3.1 function in parenchymal arterioles of Aged mice (&gt;24 months) when compared to Young (4‐6 months). Data are means ± SEM, Young vs. Aged males and females. No sex differences were observed, thus the data were combined. Using electrophysiology, we observed that freshly‐isolated endothelial cell tubes from Aged mice showed a similar hyperpolarization to the purinergic receptor agonist 2‐methyl‐S‐ATP (1 µM), which purportedly activates the downstream effectors KIR2 and KCa2.3 / KCa3.1, than those from Young mice (Δmembrane potential (Vm): ‐21.0 ± 1.7 vs ‐23.9 ± 5.1 mV, n = 5‐7), although vasodilatory responses of pressurized parenchymal arterioles were smaller (%Vasodilation: 22.4 ± 3.8 vs. 12.7 ± 3.2%, p&lt;0.05, Student’s t‐test, n = 7‐6). Activity of KCa2.3 / KCa3.1 after activation by NS‐309 (1 µM) was unchanged by aging, as evidenced by similar hyperpolarization (ΔVm: ‐36.1 ± 2.4 vs. ‐32.4 ± 2.2 mV, n = 13‐14) and vasodilation (%Vasodilation: 16.5 ± 2.0 vs. 21.3 ± 3.2%, n = 8‐6). Activation of KIR2 by 15 mM [K+]E induced a smaller hyperpolarization in Aged than Young (ΔVm: ‐11.0 ± 1.1 vs ‐6.9 ± 0.7 mV, p&lt;0.05, Student’s t‐test, n = 13‐13), with a paradoxical larger vasodilation that was unchanged by endothelium removal (%Vasodilation: 10.7 ± 2.3 vs. 25.0 ± 3.6% vs. 27.3 ± 4.5, one‐way ANOVA, Young endothelium intact vs. Aged endothelium intact vs Aged endothelium removed, n = 7‐13‐6), suggesting a compensation by smooth muscle KIR2. Lastly, spontaneous myogenic tone was higher in parenchymal arterioles from Aged mice, and endothelium removal did not further enhance myogenic tone (%myogenic tone: 25.4 ± 1.7 vs. 34.3 ± 2.3 vs 36.4 ± 3.2%, p&lt;0.05, one‐way ANOVA, Young endothelium intact vs. Aged endothelium intact vs Aged endothelium removed, n = 22‐23‐6). In conclusion, aging impairs endothelial KIR2 channels and increases resting myogenic tone, which may underlie vascular dysfunction and exacerbate cognitive decline.