bluuebunny/crossref_metadata_2025 huggingface dataset:
Microvascular oxygenation, oxidative stress, NO suppression and superoxide dismutase during postischemic reperfusion
Authors: Bertuglia S. , Giusti A.
Published: March 2015

Increased formation of reactive oxygen species (ROS) on reperfusion after ischemia underlies ischemia-reperfusion (I/R) damage. We measured, in real time, oxygen tension in both microvessels and tissue and oxidant stress during postischemic reperfusion in the hamster cheek pouch microcirculation. We measured Po2 by using phosphorescence quenching microscopy and ROS production in the systemic blood. We evaluated the effects of a nitric oxide synthase inhibitor ( NG-monomethyl-l-arginine, l-NMMA) and SOD on the oxidative stress during reperfusion. Microvascular injury was assessed by measuring diameter change, the perfused capillary length (PCL), and leukocyte adhesion. During early reperfusion, arteriolar Po2 was significantly lower than baseline, whereas capillary Po2 varied between 7 and 0 mmHg. Arterial blood flow did not regain baseline values, whereas Po2 returned to baseline in arterioles and tissue after 30 min of reperfusion. During 5 and 15 min of reperfusion, ROS increased by 72 and 89% versus baseline, respectively, and declined to baseline after 30 min of reperfusion. Pretreatment with SOD maintained ROS at normal levels, increased arteriolar diameter, blood flow, and PCL, and decreased leukocyte adhesion ( P < 0.05). l-NMMA decreased ROS only within 5 min of reperfusion, which increased significantly by 72% later during reperfusion. l-NMMA worsened leukocyte adhesion ( P < 0.05). In conclusion, our results show that the early reperfusion is characterized by low Po2 linked to increased production of ROS. At early reperfusion both SOD and l-NMMA decreased ROS production, whereas only SOD reduced it during later reperfusion. We suggest that low-flow hypoxia profoundly affects vascular endothelial damage during reperfusion through changes in ROS and nitric oxide production.
Stress upregulates arterial matrix metalloproteinase expression and activity via endothelin A receptor activation
Authors: Ergul Adviye , Portik-Dobos Vera , Giulumian Ararat D. , Molero Mariela M. , Fuchs Leslie C.
Published: March 2015

Degradation of the extracellular matrix proteins by matrix metalloproteinases (MMP) is an important regulatory step in the vascular remodeling process. Recent studies demonstrated that ETA receptors regulate cardiac MMP activity and fibrosis in DOCA-salt hypertension. However, little is known about endothelin (ET)-1 regulation of vascular MMP activity in hypertension. Thus early changes in ET-1-mediated regulation of MMP activity were measured in borderline hypertensive rats that develop impaired vasorelaxation and hypertension with chronic exposure to stress. Experiments were performed after 10 days of exposure to the behavioral stressor, air-jet stress, but before the onset of stress-induced hypertension. Study groups were 1) control ( n = 8); 2) air-jet stress for 10 days ( n = 8); 3) control plus ETA antagonist ABT-627 ( n = 4), and 4) air-jet stress plus ETA antagonist ( n = 4). MMP activity in the thoracic aorta was assessed by gelatin zymography. MMP protein and tissue ET-1 levels were evaluated by immunohistochemistry, and ET receptor density was determined by immunoblotting. Exposure to stress caused a twofold increase in plasma ET-1 levels ( P < 0.05), and there was increased ET-1 staining at the tissue level. Total MMP activity and expression of MMP-2 and MMP-9 were increased in the stress group. ETA receptor antagonism prevented the increase in MMP expression and activation in the stress group. These results provide evidence that the MMP system is activated before the development of hypertension and ET-1 mediates these early events in vascular remodeling.
Ischemic cardiomyopathy in pigs with two-vessel occlusion and viable, chronically dysfunctional myocardium
Authors: Fallavollita James A. , Canty John M.
Published: March 2015

A chronic left anterior descending coronary artery (LAD) stenosis leads to the development of hibernating myocardium with severe regional hypokinesis but normal global ventricular function after 3 mo. We hypothesized that two-vessel occlusion would accelerate the progression to hibernating myocardium and lead to global left ventricular (LV) dysfunction and heart failure. Pigs were instrumented with a fixed 1.5-mm constrictor on the proximal LAD and circumflex arteries. After 2 mo, there were no overt signs of right-heart failure and triphenyl tetrazolium chloride infarction was trivial (1.4 ± 0.1% of the LV). Compared with shams, regional function [myocardial systolic excursion (ΔWT); 2.1 ± 0.3 vs. 4.6 ± 0.4 mm, P < 0.05] and resting perfusion (0.90 ± 0.13 vs. 1.32 ± 0.09 ml · min−1· g−1, P < 0.05) were reduced, consistent with hibernating myocardium. Pulmonary systolic (45.9 ± 3.3 vs. 36.5 ± 2.2 mmHg, P < 0.05) and wedge pressures (19.1 ± 1.6 vs. 11.2 ± 0.9 mmHg, P < 0.05) were increased with global ventricular dysfunction (ejection fraction 43 ± 2 vs. 50 ± 2%, P < 0.05). Early LV remodeling was present with increased cavity size and mass. Reductions in sarcoplasmic reticulum Ca2+-ATPase and phospholamban were confined to the dysfunctional LAD region with no change in calsequestrin. Thus combined stenoses of the LAD and circumflex arteries accelerate the development of hibernating myocardium and result in compensated heart failure.
Carotid baroreflex function during and following voluntary apnea in humans
Authors: Muenter Swift N. , Cutler M. J. , Fadel P. J. , Wasmund W. L. , Ogoh S. , Keller D. M. , Raven P. B. , Smith M. L.
Published: March 2015

Muscle sympathetic nerve activity (MSNA) and arterial pressure increase concomitantly during apnea, suggesting a possible overriding of arterial baroreflex inhibitory input to sympathoregulatory centers by apnea-induced excitatory mechanisms. Apnea termination is accompanied by strong sympathoinhibition while arterial pressure remains elevated. Therefore, we hypothesized that the sensitivity of carotid baroreflex control of MSNA would decrease during apnea and return upon apnea termination. MSNA and heart rate responses to –60-Torr neck suction (NS) were evaluated during baseline and throughout apnea. Responses to +30-Torr neck pressure (NP) were evaluated during baseline and throughout 1 min postapnea. Apnea did not affect the sympathoinhibitory or bradycardic response to NS ( P > 0.05); however, whereas the cardiac response to NP was maintained postapnea, the sympathoexcitatory response was reduced for 50 s ( P < 0.05). These data demonstrate that the sensitivity of carotid baroreflex control of MSNA is not attenuated during apnea. We propose a transient rightward and upward resetting of the carotid baroreflex-MSNA function curve during apnea and that return of the function curve to, or more likely beyond, baseline (i.e., a downward and leftward shift) upon apnea termination may importantly contribute to the reduced sympathoexcitatory response to NP.
Mechanism of RhoA/Rho kinase activation in endothelin-1- induced contraction in rabbit basilar artery
Authors: Miao Liyan , Dai Yun , Zhang John
Published: March 2015

This study was undertaken to demonstrate the role of the RhoA/Rho kinase pathway in endothelin-1 (ET-1)-induced contraction of the rabbit basilar artery. Isometric tension and Western blot were used to examine ET-1-induced contraction and RhoA activation. The upstream effect on ET-1-induced RhoA activity was determined by using ETAand ETBreceptor antagonists, protein kinase C (PKC), tyrosine kinase, and phosphatidylinositol-3 kinase inhibitors. The downstream effect of ET-1-induced contraction and RhoA activity was studied in the presence of the Rho kinase inhibitor Y-27632. The effect of Rho kinase inhibitor on ET-1-induced myosin light chain (MLC) phosphorylation was investigated by using urea-glycerol-PAGE immunoblotting. We found 1) ET-1 increased RhoA activity (membrane binding RhoA) in a concentration-dependent manner; 2) ETA, but not ETB, receptor antagonist abolished the effect of ET-1 on RhoA activation; 3) phosphodylinositol-3 kinase inhibitor, but not PKC and tyrosine kinase inhibitors, reduced ET-1-induced RhoA activation; 4) Rho kinase inhibitor Y-27632 (10 μM) inhibited ET-1-induced contraction; and 5) ET-1 increased the level of MLC phosphorylation. Rho kinase inhibitor Y-27632 reduced the effect of ET-1 on MLC phosphorylation. This study demonstrated that RhoA/Rho kinase activation is involved in ET-1-induced contraction in the rabbit basilar artery. Phosphodylinositol-3 kinase and MLC might be the upstream and downstream factors of RhoA activation.
Increased expression of protein kinase C isoforms in heart failure due to myocardial infarction
Authors: Wang Jingwei , Liu Xueliang , Sentex Emmanuelle , Takeda Nobuakira , Dhalla Naranjan S.
Published: March 2015

The activities of cardiac protein kinase C (PKC) were examined in hemodynamically assessed rats subsequent to myocardial infarction (MI). Both Ca2+-dependent and Ca2+-independent PKC activities increased significantly in left ventricular (LV) and right ventricular (RV) homogenates at 1, 2, 4, and 8 wk after MI was induced. PKC activities were also increased in both LV and RV cytosolic and particulate fractions from 8-wk infarcted rats. The relative protein contents of PKC-α, -β, -ε, and -ζ isozymes were significantly increased in LV homogenate, cytosolic (except PKC-α), and particulate fractions from the failing rats. On the other hand, the protein contents of PKC-α, -β, and -ε isozymes, unlike the PKC-ζ isozyme, were increased in RV homogenate and cytosolic fractions, whereas the RV particulate fraction showed an increase in the PKC-α isozyme only. These changes in the LV and RV PKC activities and protein contents in the 8-wk infarcted animals were partially corrected by treatment with the angiotensin-converting enzyme inhibitor imidapril. No changes in protein kinase A activity and its protein content were seen in the 8-wk infarcted hearts. The results suggest that the increased PKC activity in cardiac dysfunction due to MI may be associated with an increase in the expression of PKC-α, -β, and -ε isozymes, and the improvement of heart function in the infarcted animals by imidapril may be due to partial prevention of changes in PKC activity and isozyme contents.
Antioxidant MCI-186 inhibits mitochondrial permeability transition pore and upregulates Bcl-2 expression
Authors: Rajesh Katare Gopalrao , Sasaguri Shiro , Suzuki Ryoko , Maeda Hironori
Published: March 2015

Reperfusion after a period of ischemia is associated with the formation of reactive oxygen species (ROS) and Ca2+overload resulting in the opening of a nonspecific pore in the inner membrane of the mitochondria, called the mitochondrial permeability transition pore (PTP), leading to cell damage. Although endogenous antioxidants are activated because of oxidative stress following ischemia, their levels are not high enough to prevent reperfusion injury. Hence there is always a need for exogenous supplement of antioxidants, especially after acute ischemia. Here we demonstrated the effects of the antioxidant 3-methyl-1-phenyl-2-pyrazolin-5-one (MCI-186) in preventing reperfusion injury of the heart by inhibition of PTP opening. Ischemia (30 min) by left coronary artery (LCA) occlusion and reperfusion (120 min) in Wistar rats after pretreatment with MCI-186 (10 mg/kg iv) infusion starting from 30 min before LCA occlusion resulted in 1) less area of myocardial infarction (19.2% vs. 61.6%), 2) well-maintained myocardial ATP content ( P < 0.03 vs. control), 3) decreased mitochondrial swelling and reduced cytochrome c release, 4) increased expression of BCl-2, 5) lower prevalence of apoptotic cells (14.3% vs. 2.9%), and 6) reduced DNA fragmentation in the MCI-186-treated group. These cytoprotective effects of MCI-186 were inhibited on opening PTP before MCI-186 treatment with the PTP activators lonidamine (10 mg/kg iv) or atractyloside (5 mg/kg iv) but failed to inhibit the protective effects exerted by another antioxidant, allopurinol, suggesting that the PTP inhibiting property is specific for MCI-186. These results demonstrate that the radical scavenger MCI-186, by inhibiting the opening of the PTP, prevents necrosis and cytochrome c release and hence pathological apoptosis.
Atrial contraction and mitral annular dynamics during acute left atrial and ventricular ischemia in sheep
Authors: Timek Tomasz A. , Lai David T. , Tibayan Frederick , Daughters George T. , Liang David , Dagum Paul , Lo Sidney , Miller D. Craig , Ingels Neil B.
Published: March 2015

In six sheep, radiopaque markers were placed on the left ventricle (LV), the mitral annulus, the left atrium (LA), and the central edge of both mitral leaflets to investigate the effects of acute LV ischemia on atrial contraction, mitral annular area (MAA), and mitral regurgitation (MR). Animals were studied with biplane videofluoroscopy and transesophageal echocardiography before and during balloon occlusion of the left anterior descending (LAD), distal circumflex (dLCX), and proximal circumflex (pLCX) coronary arteries. MAA and LA area were calculated from the corresponding markers. LAD occlusion did not alter LA area reduction or presystolic MAA reduction, whereas dLCX occlusion resulted in a mild decrease in the former with no change in the latter. Neither occlusion resulted in MR. pLCX occlusion, however, significantly decreased LA area and presystolic MAA reduction and resulted in increased end-diastolic MAA, delayed valve closure from end diastole, and MR. Decreased atrial contractile function, as observed during acute posterolateral ischemia, is linked to diminished presystolic mitral annular reduction, a larger mitral annular size at end diastole, and MR.
Nucleoside reverse transcriptase inhibitors impair endothelium-dependent relaxation by increasing superoxide
Authors: Sutliff Roy L. , Dikalov Sergey , Weiss Daiana , Parker Jeremy , Raidel Scott , Racine Andrea K. , Russ Rodney , Haase Chad P. , Taylor W. Robert , Lewis William
Published: March 2015

Nucleoside reverse transcriptase inhibitors (NRTIs) have been used successfully to reduce acquired immunodeficiency syndrome mortality. However, the use of these compounds is associated with numerous tissue toxicities, including cardiomyopathy. These studies address the effects of NRTIs on vascular function. Functional assays of contraction and relaxation were performed on isolated mouse aorta segments obtained from FVB/n mice exposed to zidovudine (AZT), stavudine, or water for 35 days. AZT and stavudine treatment impaired sensitivity to endothelium-dependent relaxation by acetylcholine. Dihydroethidium staining revealed that AZT treatment was associated with an increase in superoxide levels. Pretreatment of AZT-treated vessels with tiron (1 mM), a free radical scavenger, restored endothelium-dependent relaxation in mice. In cellular preparations, electron spin resonance measurements revealed elevated superoxide in cultured endothelial cells exposed to AZT; elevation was dependent on the length of exposure. These results indicate that NRTIs impair endothelium-dependent relaxation by increasing superoxide levels and suggest that NRTI therapy contributes to cardiovascular complications in acquired immunodeficiency syndrome.
Normal contractions triggered by I Ca,L in ventricular myocytes from rats with postinfarction CHF
Authors: Sjaastad Ivar , Bøkenes Janny , Swift Fredrik , Wasserstrom J. Andrew , Sejersted Ole M.
Published: March 2015

Attenuated L-type Ca2+ current ( I Ca,L), or current-contraction gain have been proposed to explain impaired cardiac contractility in congestive heart failure (CHF). Six weeks after coronary artery ligation, which induced CHF, left ventricular myocytes from isoflurane-anesthetized rats were current or voltage clamped from −70 mV. In both cases, contraction and contractility were attenuated in CHF cells compared with cells from sham-operated rats when cells were only minimally dialyzed using high-resistance microelectrodes. With patch pipettes, cell dialysis caused attenuation of contractions in sham cells, but not CHF cells. Stepping from −50 mV, the following variables were not different between sham and CHF, respectively: peak I Ca,L (4.5 ± 0.3 vs. 3.8 ± 0.3 pApF−1 at 23°C and 9.4 ± 0.5 vs. 8.4 ± 0.5 pApF−1 at 37°C), the bell-shaped voltage-contraction relationship in Cs+ solutions (fractional shortening, 15.2 ± 1.0% vs. 14.3 ± 0.7%, respectively, at 23°C and 7.5 ± 0.4% vs. 6.7 ± 0.5% at 37°C) and the sigmoidal voltage-contraction relationship in K+ solutions. Caffeine-induced Ca2+ release and sarcoplasmic reticulum Ca2+-ATPase-to-phospholamban ratio were not different. Thus CHF contractions triggered by I Ca,L were normal, and the contractile deficit was only seen in undialyzed cardiomyocytes stimulated from −70 mV.
Baicalein attenuates oxidant stress in cardiomyocytes
Authors: Shao Zuo-Hui , Vanden Hoek Terry L. , Qin Yimin , Becker Lance B. , Schumacker Paul T. , Li Chang-Qing , Dey Lucy , Barth Eugene , Halpern Howard , Rosen Gerald M. , Yuan Chun-Su
Published: March 2015

Flavonoids within Scutellaria baicalensis may be potent antioxidants on the basis of our studies of S. baicalensis extract. To further this work, we studied the antioxidative effects of baicalein, a flavonoid component of S. baicalensis, in a chick cardiomyocyte model of reactive oxygen species (ROS) generation during hypoxia, simulated ischemia-reperfusion, or mitochondrial complex III inhibition with antimycin A. Oxidant stress was measured by oxidation of the intracellular probes 2′,7′-dichlorofluorescin diacetate and dihydroethidium. Viability was assessed by propidium iodide uptake. Baicalein attenuated oxidant stress during all conditions studied and acted within minutes of treatment. For example, baicalein given only at reperfusion dose dependently attenuated the ROS burst at 5 min after 1 h of simulated ischemia. It also decreased subsequent cell death at 3 h of reperfusion from 52.3 ± 2.5% in untreated cells to 29.4 ± 3.0% (with return of contractions; P < 0.001). In vitro studies using electron paramagnetic resonance spectroscopy with the spin trap 5-methoxycarbonyl-5-methyl-1-pyrroline- N-oxide revealed that baicalein scavenges superoxide but does not mimic the effects of superoxide dismutase. We conclude that baicalein can scavenge ROS generation in cardiomyocytes and that it protects against cell death in an ischemia-reperfusion model when given only at reperfusion.
17β-Estradiol inhibits angiotensin II activation of area postrema neurons
Authors: Pamidimukkala Jaya , Hay Meredith
Published: March 2015

It is well established that the area postrema, as a circumventricular organ, is susceptible to modulation by circulating hormones and peptides. Furthermore, activation of the area postrema has been shown to modulate central neurons involved in the regulation of cardiovascular function and blood pressure. In particular, the vasoactive peptide angiotensin II (ANG II) has been shown to inhibit baroreflex regulation of heart rate and increase sympathetic outflow and blood pressure via activation of area postrema neurons. Estrogen is thought to protect against hypertension in both humans and animal models and has been shown in a number of systems to alter the effects of ANG II. The purpose of the present study was to determine the effects of estrogen on ANG II activation of area postrema neurons. In this study, the effects of ANG II and KCl on fura 2-measured cytosolic Ca2+concentration ([Ca2+]i) responses in cultured area postrema neurons in the presence and absence of 12-h exposure to 100 nM 17β-estradiol (E2) were evaluated. In neurons incubated in control vehicle media, 50 nM ANG II increased [Ca2+]iby 92 ± 12%. In neurons preincubated with 100 nM E2, ANG II increased [Ca2+]iby only 68 ± 11%, for a total inhibition of the ANG II-evoked response of 24%. Coapplication of the estrogen receptor antagonist ICI-182,780 did not inhibit the effects of E2. In the same cells in which the effects of E2on ANG II-evoked responses were tested, the effects of incubation in E on the depolarization-induced increased [Ca2+2]idue to 60 mM KCl were also tested. Incubation of the cells with 100 nM E increased the KCl-evoked [Ca2+2]iresponse, and this response was blocked by ICI-182,780. These results suggest that in the area postrema, estrogen may utilize multiple pathways to modulate neural activity and responses to ANG II.
Microvascular oxygen distribution in awake hamster window chamber model during hyperoxia
Authors: Tsai Amy G. , Cabrales Pedro , Winslow Robert M. , Intaglietta Marcos
Published: March 2015

The microvascular effects and hemodynamic events following exposure to normobaric hyperoxia (because of inspiration of 100% O2) were studied in the awake hamster window chamber model and compared with normoxia. Hyperoxia increased arterial blood Po2to 477.9 ± 19.9 from 60.0 ± 1.2 mmHg ( P < 0.05). Heart rate and blood pressure were unaltered, whereas cardiac index was reduced from 196 ± 13 to 144 ± 31 ml · min–1· kg–1( P < 0.05) in hyperoxia. Direct measurements in the microcirculation showed there was arteriolar vasoconstriction, reduction of microvascular flow (83% of control, P < 0.05), and functional capillary density (FCD, 74 ± 16% of control), the latter change being significant ( P < 0.05). Calculations of oxygen delivery and oxygen consumption based on the measured changes in microvascular blood flow velocity and diameter and estimates of oxygen saturation corrected for the Bohr effect due to the lowered pH and increased Pco2showed that oxygen transport in the microvascular network did not change between normal and hyperoxic condition. The congruence of systemic and microvascular hemodynamics events found with hyperoxia suggests that the microvascular findings are common to most tissues in the organism, and that hyperoxia, due to vasoconstriction and the decrease of FCD, causes a maldistribution of perfusion in the microcirculation.
Endothelin-induced modulation of neuropeptide Y and norepinephrine release from the rat mesenteric bed
Authors: Hoang Dan , Macarthur Heather , Gardner Alice , Westfall Thomas C.
Published: March 2015

The effect of three endothelin (ET) agonists [ET-1, ET-3, and sarafotoxin (STX6C)] on the nerve stimulation-induced release of norepinephrine (NE) and neuropeptide Y-immunoreactive compounds (NPY-ir) from the perfused mesenteric arterial bed of the rat as well as the effect on perfusion pressure were examined. ET-1, ET-3, and STX6C all produced a significant, concentration-dependent decrease in the evoked release of NPY-ir but had no effect on the release of NE. In contrast, all three ETs potentiated the nerve stimulation-induced increase in perfusion pressure. The inhibition of nerve stimulation-induced NPY-ir release by ET-1 was significantly blocked by the ETA/ETBantagonist PD-142893 and the ETB antagonist RES-701-1 but not by the ETA antagonist BQ-123. The potentiation of the nerve stimulation-induced increase in perfusion pressure by ET-1 was significantly blocked by PD-142893 and BQ-123 and attenuated by RES-701-1. Prior exposure of the preparation to indomethacin or meclofenamate failed to alter the attenuation of the evoked release of NPY-ir or the potentiation of the increase in perfusion pressure produced by ET-1 or ET-3. These results are consistent with the idea that sympathetic cotransmitters can be preferentially modulated by paracrine mediators at the vascular neuroeffector junction.
Effects of Na+/Ca2+ exchanger downregulation on contractility and [Ca2+]i transients in adult rat myocytes
Authors: Tadros George M. , Zhang Xue-Qian , Song Jianliang , Carl Lois L. , Rothblum Lawrence I. , Tian Qiang , Dunn Jeremy , Lytton Jonathan , Cheung Joseph Y.
Published: March 2015

Postmyocardial infarction (MI) rat myocytes demonstrated depressed Na+/Ca2+exchange (NCX1) activity, altered contractility, and intracellular Ca2+ concentration ([Ca2+]i) transients. We investigated whether NCX1 downregulation in normal myocytes resulted in contractility changes observed in MI myocytes. Myocytes infected with adenovirus expressing antisense (AS) oligonucleotides to NCX1 had 30% less NCX1 at 3 days and 66% less NCX1 at 6 days. The half-time of relaxation from caffeine-induced contracture was twice as long in ASNCX1 myocytes. Sarcoplasmic reticulum (SR) Ca2+-ATPase abundance, SR Ca2+uptake, resting membrane potential, action potential amplitude and duration, L-type Ca2+ current density and cell size were not affected by ASNCX1 treatment. At extracellular Ca2+ concentration ([Ca2+]o) of 5 mM, ASNCX1 myocytes had significantly lower contraction and [Ca2+]i transient amplitudes and SR Ca2+ contents than control myocytes. At 0.6 mM [Ca2+]o, contraction and [Ca2+]i transient amplitudes and SR Ca2+ contents were significantly higher in ASNCX1 myocytes. At 1.8 mM [Ca2+]o, contraction and [Ca2+]i transient amplitudes were not different between control and ASNCX1 myocytes. This pattern of contractile and [Ca2+]i transient abnormalities in ASNCX1 myocytes mimics that observed in rat MI myocytes. We conclude that downregulation of NCX1 in adult rat myocytes resulted in decreases in both Ca2+ influx and efflux during a twitch. We suggest that depressed NCX1 activity may partly account for the contractile abnormalities after MI.
Downregulation of profilin with antisense oligodeoxynucleotides inhibits force development during stimulation of smooth muscle
Authors: Tang Dale D. , Tan Jian
Published: March 2015

The actin-regulatory protein profilin has been shown to regulate the actin cytoskeleton and the motility of nonmuscle cells. To test the hypothesis that profilin plays a role in regulating smooth muscle contraction, profilin antisense or sense oligodeoxynucleotides were introduced into the canine carotid smooth muscle by a method of reversible permeabilization, and these strips were incubated for 2 days for protein downregulation. The treatment of smooth muscle strips with profilin antisense oligodeoxynucleotides inhibited the expression of profilin; it did not influence the expression of actin, myosin heavy chain, and metavinculin/vinculin. Profilin sense did not affect the expression of these proteins in smooth muscle tissues. Force generation in response to stimulation with norepinephrine or KCl was significantly lower in profilin antisense-treated muscle strips than in profilin sense-treated strips or in muscle strips not treated with oligodeoxynucleotides. The depletion of profilin did not attenuate increases in phosphorylation of the 20-kDa regulatory light chain of myosin (MLC20) in response to stimulation with norepinephrine or KCl. The increase in F-actin/G-actin ratio during contractile stimulation was significantly inhibited in profilin-deficient smooth muscle strips. These results suggest that profilin is a necessary molecule of signaling cascades that regulate carotid smooth muscle contraction, but that it does not modulate MLC20phosphorylation during contractile stimulation. Profilin may play a role in the regulation of actin polymerization or organization in response to contractile stimulation of smooth muscle.
A highly potent peptide analgesic that protects against ischemia-reperfusion-induced myocardial stunning
Authors: Wu Dunli , Soong Yi , Zhao Guo-Min , Szeto Hazel H.
Published: March 2015

We recently discovered an opioid peptide analgesic, 2′,6′-dimethyltyrosine (Dmt)-d-Arg-Phe-Lys-NH2([Dmt1]DALDA), that can protect against ischemia-induced myocardial stunning. In buffer-perfused hearts, 30-min global ischemia followed by reperfusion resulted in a significant increase in norepinephrine (NE) overflow immediately upon reperfusion and significant decline in contractile force (45%). Pretreatment with [Dmt1]DALDA before ischemia completely abolished myocardial stunning and significantly reduced NE overflow (68%). In contrast, pretreatment with morphine before ischemia only provided brief protection against myocardial stunning and no reduction in NE overflow. [Dmt1]DALDA inhibited [3H]NE uptake into cardiac synaptosomes in vitro (IC50 = 3.9 μM), whereas morphine had no effect. Surprisingly, protection against myocardial stunning was apparent even when hearts were perfused with [Dmt1]DALDA only upon reperfusion, whereas reperfusion with morphine had no effect. Binding studies with [3H][Dmt1]DALDA revealed no high-affinity specific binding in cardiac membranes, suggesting that the cardioprotective actions of [Dmt1]DALDA are not mediated via opioid receptors. These findings suggest that [Dmt1]DALDA is a potent analgesic that may be useful for myocardial stunning resulting from cardiac interventions or myocardial ischemia.
H2O2-induced Ca2+overload in NRVM involves ERK1/2 MAP kinases: role for an NHE-1-dependent pathway
Authors: Rothstein Emily C. , Byron Kenneth L. , Reed Ryan E. , Fliegel Larry , Lucchesi Pamela A.
Published: March 2015

Generation of reactive oxygen species (ROS) and intracellular Ca2+overload are key mechanisms involved in ischemia-reperfusion (I/R)-induced myocardial injury. The relationship between I/R injury and Ca2+overload has not been fully characterized. The increase in Na+/H+exchanger (NHE-1) activity observed during I/R injury is an attractive candidate to link increased ROS production with Ca2+overload. We have shown that low doses of H2O2increase NHE-1 activity in an extracellular signal-regulated kinase (ERK)-dependent manner. In this study, we examined the effect of low doses of H2O2on intracellular Ca2+in fura 2-loaded, spontaneously contracting neonatal rat ventricular myocytes. H2O2induced a time- and concentration-dependent increase in diastolic intracellular Ca2+concentration that was blocked by inhibition of ERK1/2 activation with 5 μM U-0126 (88%) or inhibition of NHE-1 with 5 μM HOE-642 (50%). Increased NHE activity was associated with phosphorylation of the NHE-1 carboxyl tail that was blocked by U-0126. These results suggest that H2O2induced Ca2+overload is partially mediated by NHE-1 activation secondary to phosphorylation of NHE-1 by the ERK1/2 MAP kinase pathway.
Targeted deletion of MMP-2 attenuates early LV rupture and late remodeling after experimental myocardial infarction
Authors: Hayashidani Shunji , Tsutsui Hiroyuki , Ikeuchi Masaki , Shiomi Tetsuya , Matsusaka Hidenori , Kubota Toru , Imanaka-Yoshida Kyoko , Itoh Takeshi , Takeshita Akira
Published: March 2015

Matrix metalloproteinase-2 (MMP-2) is prominently overexpressed both after myocardial infarction (MI) and in heart failure. However, its pathophysiological significance in these conditions is still unclear. We thus examined the effects of targeted deletion of MMP-2 on post-MI left ventricular (LV) remodeling and failure. Anterior MI was produced in 10- to 12-wk-old male MMP-2 knockout (KO) and sibling wild-type (WT) mice by ligating the left coronary artery. By day 28, MI resulted in a significant increase in mortality in association with LV cavity dilatation and dysfunction. The MMP-2 KO mice had a significantly better survival rate than WT mice (56% vs. 85%, P < 0.05), despite a comparable infarct size (50 ± 3% vs. 51 ± 3%, P = not significant), heart rate, and arterial blood pressure. The KO mice had a significantly lower incidence of LV rupture (10% vs. 39%, P < 0.05), which occurred within 7 days of MI. The KO mice exerted less LV cavity dilatation and improved fractional shortening after MI by echocardiography. The LV zymographic MMP-2 level significantly increased in WT mice after coronary artery ligation; however, this was completely prevented in KO mice. In contrast, the increase in the LV zymographic MMP-9 level after MI was similar between KO and WT mice. MMP-2 activation is therefore considered to contribute to an early cardiac rupture as well as late LV remodeling after MI. The inhibition of MMP-2 activation may therefore be a potentially useful therapeutic strategy to manage post-MI hearts.
Shear stress stimulates phosphorylation of eNOS at Ser635by a protein kinase A-dependent mechanism
Authors: Boo Yong Chool , Hwang Jinah , Sykes Michelle , Michell Belinda J. , Kemp Bruce E. , Lum Hazel , Jo Hanjoong
Published: March 2015

Shear stress stimulates nitric oxide (NO) production by phosphorylating endothelial NO synthase (eNOS) at Ser1179in a phosphoinositide-3-kinase (PI3K)- and protein kinase A (PKA)-dependent manner. The eNOS has additional potential phosphorylation sites, including Ser116, Thr497, and Ser635. Here, we studied these potential phosphorylation sites in response to shear, vascular endothelial growth factor (VEGF), and 8-bromocAMP (8-BRcAMP) in bovine aortic endothelial cells (BAEC). All three stimuli induced phosphorylation of eNOS at Ser635, which was consistently slower than that at Ser1179. Thr497was rapidly dephosphorylated by 8-BRcAMP but not by shear and VEGF. None of the stimuli phosphorylated Ser116. Whereas shear-stimulated Ser635phosphorylation was not affected by phosphoinositide-3-kinase inhibitors wortmannin and LY-294002, it was blocked by either treating the cells with a PKA inhibitor H89 or infecting them with a recombinant adenovirus-expressing PKA inhibitor. These results suggest that shear stress stimulates eNOS by two different mechanisms: 1) PKA- and PI3K-dependent and 2) PKA-dependent but PI3K-independent pathways. Phosphorylation of Ser635may play an important role in chronic regulation of eNOS in response to mechanical and humoral stimuli.
Renal vascular responses to static handgrip: role of muscle mechanoreflex
Authors: Momen Afsana , Leuenberger Urs A. , Ray Chester A. , Cha Susan , Handly Brian , Sinoway Lawrence I.
Published: March 2015

During exercise, the sympathetic nervous system is activated, which causes vasoconstriction. The autonomic mechanisms responsible for this vasoconstriction vary based on the particular tissue being studied. Attempts to examine reflex control of the human renal circulation have been difficult because of technical limitations. In this report, the Doppler technique was used to examine renal flow velocity during four muscle contraction paradigms in conscious humans. Flow velocity was divided by mean arterial blood pressure to yield an index of renal vascular resistance (RVR). Fatiguing static handgrip (40% of maximal voluntary contraction) increased RVR by 76%. During posthandgrip circulatory arrest, RVR remained above baseline (2.1 ± 0.2 vs. 2.8 ± 0.2 arbitrary units; P < 0.017) but was only 40% of the end-grip RVR value. Voluntary biceps contraction increased RVR within 10 s of initiation of contraction. This effect was not associated with an increase in blood pressure. Finally, involuntary biceps contraction also raised RVR. We conclude that muscle contraction evokes renal vasoconstriction in conscious humans. The characteristic of this response is consistent with a primary role for mechanically sensitive afferents. This statement is based on the small posthandgrip circulatory arrest response and the vasoconstriction that was observed with involuntary biceps contraction.
Comparison of generalized and gender-specific transfer functions for the derivation of aortic waveforms
Authors: Hope Sarah A. , Tay David B. , Meredith Ian T. , Cameron James D.
Published: March 2015

Arterial transfer functions have been promoted for the derivation of central aortic waveform characteristics not usually accessible noninvasively, but possibly of prognostic significance. The utility of generalized rather than gender-specific transfer functions has not been assessed. Invasive central aortic and noninvasive radial (Millar Mikro-tip tonometer) blood pressure waveforms were recorded simultaneously in 78 subjects (61 male and 17 female). Average transfer functions were obtained for the whole group and for each gender by two methods. Reverse transformation was performed with the use of each transfer function. Measured aortic waveform parameters were compared with those derived using average, gender-appropriate, and gender-inappropriate transfer functions. Differences in central waveform characteristics were demonstrated between men and women. Derived waveform parameters were significantly different from measured values [e.g., subendocardial viability index and augmentation index ( P < 0.001)]. A gender-appropriate transfer function significantly improved the derivation of some parameters, including systolic pressure and systolic and diastolic pressure time integrals ( P < 0.05). Generalized arterial transfer functions may not be universally applicable across all waveform parameters of potential interest, and gender-specific transfer functions may be more appropriate.
Elevated plasma levels of human urotensin-II immunoreactivity in congestive heart failure
Authors: Russell Fraser D. , Meyers Deborah , Galbraith Andrew J. , Bett Nick , Toth Istvan , Kearns Philip , Molenaar Peter
Published: March 2015

Human urotensin-II (hU-II) is the most potent endogenous cardiostimulant identified to date. We therefore determined whether hU-II has a possible pathological role by investigating its levels in patients with congestive heart failure (CHF). Blood samples were obtained from the aortic root, femoral artery, femoral vein, and pulmonary artery from CHF patients undergoing cardiac catheterization and the aortic root from patients undergoing investigative angiography for chest pain who were not in heart failure. Immunoreactive hU-II (hU-II-ir) levels were determined with radioimmunoassay. hU-II-ir was elevated in the aortic root of CHF patients (230.9 ± 68.7 pg/ml, n = 21; P < 0.001) vs. patients with nonfailing hearts (22.7 ± 6.1 pg/ml, n = 18). This increase was attributed to cardiopulmonary production of hU-II-ir because levels were lower in the pulmonary artery (38.2 ± 6.1 pg/ml, n = 21; P < 0.001) than in the aortic root. hU-II-ir was elevated in the aortic root of CHF patients with nonischemic cardiomyopathy (142.1 ± 51.5 pg/ml, n = 10; P < 0.05) vs. patients with nonfailing hearts without coronary artery disease (27.3 ± 12.4 pg/ml, n = 7) and CHF patients with ischemic cardiomyopathy (311.6 ± 120.4 pg/ml, n = 11; P < 0.001) vs. patients with nonfailing hearts and coronary artery disease (19.8 ± 6.6 pg/ml, n = 11). hU-II-ir was significantly higher in the aortic root than in the pulmonary artery and femoral vein, with a nonsignificant trend for higher levels in the aortic root than in the femoral artery. The findings indicated that hU-II-ir is elevated in the aortic root of CHF patients and that hU-II-ir is cleared at least in part from the microcirculation.
α1-Adrenoceptor stimulation directly induces growth of vascular wall in vivo
Authors: Erami Cauveh , Zhang Hua , Ho Jason G. , French David M. , Faber James E.
Published: March 2015

Previous studies suggesting that norepinephrine is directly trophic for the vascular wall have been confounded by concomitant hemodynamic disturbances. Herein, a microcatheter connected to an osmotic minipump was implanted adjacent to the rat carotid for 2-wk perivascular suffusion of agents at systemic levels ∼1,000 times below the threshold for altering arterial pressure. Norepinephrine decreased lumen and adventitial areas and circumference by 10, 14, and 5%, respectively (all P < 0.05); a nonsubtype-specific α1-adrenoceptor (AR) antagonist had no effect. When begun at the time of balloon injury, 2-wk norepinephrine increased lumen loss by 45%, increased neointimal area by 64% and collagen content by 33%, and reduced vessel circumference by 5% (all P < 0.05). α1-AR antagonists decreased neointimal area by 33% (all P < 0.05). α1A-AR antagonist reduced lumen loss by 70%, neointimal area by 54%, circumference decline by 84%, and adventitial thickening by 87% (all P < 0.05), whereas α1B-, α1D-, α2- and β-AR antagonists were without effect. These are the first in vivo studies demonstrating that norepinephrine is directly trophic for the vascular wall and augments injury-induced intimal lesion growth.
Longitudinal strain quantitates regional right ventricular contractile function
Authors: Jamal Fadi , Bergerot Cyrille , Argaud Laurent , Loufouat Joseph , Ovize Michel
Published: March 2015

The assessment of contractile function of the right ventricle (RV) is an important clinical issue, but this remains difficult because of its complex anatomy and structure. We thought to investigate whether new Doppler-derived myocardial deformation indexes may quantify regional contractile RV function during varying loading conditions. In nine pigs, ultrasonic crystals were inserted longitudinally in the RV inflow and outflow tracts to assess regional contractile function. The same RV segments and the interventricular septum were imaged using apical echocardiographic views. Regional function was assessed using two parameters: 1) systolic strain (SS), representing the relative magnitude of segmental systolic shortening; and 2) its temporal derivative, peak systolic strain rate (SR), i.e., the maximal velocity of segmental shortening. Data were acquired at baseline and during partial pulmonary artery constriction (PAC) and inferior vena cava occlusion (IVCO). SS decreased significantly after PAC and IVCO in both the inflow and outflow tracts but only during IVCO in the septum. SR was less sensitive to loading variations in all segments. A significant correlation was found between SS values derived from sonomicrometry and myocardial Doppler in RV segments ( r = 0.84, P < 0.001). Thus regional strain and SR provide complementary information on the heterogeneous RV contractile function and can be accurately and noninvasively quantified using Doppler myocardial imaging.
Morphometry and strain distribution of the C57BL/6 mouse aorta
Authors: Guo X. , Kono Y. , Mattrey R. , Kassab G. S.
Published: March 2015

The goal of the present study was to obtain a systematic set of data along the length of the mouse aorta to study variations of morphometry (diameter, wall thickness, and curvature), strain, and stress of the mouse aorta. Five mice were imaged with a 13-MHz ultrasound probe to determine the in vivo diameter along the aorta. A cast was made of these aortas to validate the ultrasonic diameter measurements. The root mean squared and systematic errors for these measurements were 12.6% and 6.4% of the mean ultrasound diameter, respectively. The longitudinal variations of geometry, stress, and strain from the aortic valve to the common iliac bifurcation were documented. Our results show that the residual circumferential strain leads to a uniformity of transmural strain of the aorta in the loaded state along the entire length of the aorta. Furthermore, we validated the incompressibility condition along the length of the aorta. These data of normal mice will serve as a reference state for the study of disease in future knockout models.
Purkinje involvement in arrhythmias after coronary artery reperfusion
Authors: Arnar David O. , Martins James B.
Published: March 2015

Previous studies have indicated that the endocardium may be responsible for a large portion of ventricular tachycardia (VT) seen with reperfusion of ischemic myocardium. To evaluate the role of the Purkinje system in nonreentrant VT arising from the endocardium after reperfusion, the anterior descending coronary artery was occluded for 20 min and then reperfused in 23 dogs after instrumentation of the risk zone with 21 multipolar plunge needles. VT with focal Purkinje origin was defined as a focal endocardial VT with Purkinje potentials recorded before the earliest endocardial myopotential. A total of 19 VTs (mean cycle length 214 ± 2 ms) were observed with 11 (58%) having focal Purkinje origin. Fifty-eight percent of the VTs degenerated to ventricular fibrillation, with occurrences of two or more independent foci per complex (seen in 7 of 11 compared with 1 of 8 nonsustained VTs). In conclusion, these data show that Purkinje tissue may be important in the genesis of reperfusion VT.
Downregulation of ANG II receptor is associated with compensated pressure-overload hypertrophy in the young dog
Authors: Schultz David , Su Xuefeng , Wei Chih-Chang , Bishop Sanford P. , Powell Pamela , Hankes Gerald H. , Dillon A. Ray , Rynders Patricia , Spinale Francis G. , Walcott Gregory , Ideker Raymond , Dell'Italia Louis J.
Published: March 2015

We studied the gradual onset of pressure overload (PO) induced by a mildly constricting aortic band in 8-wk-old puppies ( n = 8) that increased to 98 ± 11 mmHg at 9 mo. Left ventricular (LV) weight/body weight was increased in PO versus sham-operated littermate controls [8.11 ± 0.60 (SE) vs. 4.46 ± 0.38 g/kg, P < 0.001]. LV end-diastolic diameter, diastolic pressure, and fractional shortening did not differ in PO versus control dogs. There were no inducible arrhythmias in response to an aggressive electrophysiological stimulation protocol in PO dogs. Furthermore, isolated cardiomyocyte function did not differ between control and PO dogs. LV angiotensin II (ANG II) levels were increased (68 ± 12 vs. 20 ± 5 pg/g, P < 0.01) as steady-state ANG II type 1 (AT1) receptor mRNA was decreased 40% and endothelial nitric oxide synthase mRNA levels were increased 2.5-fold in PO versus control dogs ( P < 0.05). Total ANG II receptor binding sites of freshly prepared cardiac membranes demonstrated no difference in the dissociation constant, but there was a 60% decrease in maximum binding (Bmax) in PO versus control dogs ( P< 0.01). LV ANG II levels correlated negatively with AT1receptor mRNA levels ( r = −0.75, P < 0.01) and total AT1 receptor Bmax( r = −0.77, P < 0.02). These results suggest that LV ANG II negatively regulates AT1 receptor expression and that this is an adaptive response to chronic PO before the onset of myocardial failure in the young dog.
Hypovolemia and MSNA discharge patterns: assessing and interpreting sympathetic responses
Authors: Kimmerly D. S. , Shoemaker J. K.
Published: March 2015

We previously demonstrated that diuretic-induced hypovolemia resulted in an enhanced baroreflex-mediated increase in integrated muscle sympathetic nerve activity (MSNA) and vasomotor tone during lower body negative pressure (LBNP) ( Am J Physiol Heart Circ Physiol 282: H645–H655, 2002). The purpose of this study was to perform a retrospective analysis of these data and examine the ability of relative MSNA burst amplitude distributions to highlight differences in baseline sympathetic nerve discharge patterns. An additional purpose was to determine whether differential responses in MSNA burst frequency and burst amplitude affect conclusions regarding sympathetic reflex control. MSNA, stroke volume (SV, Doppler), and estimated central venous pressure (CVP, dependent arm technique) were measured during LBNP within the placebo (Normo) and diuretic (Hypo; 100 mg/day spironolactone for 3 days) conditions ( n = 8). Compared with Normo, MSNA burst frequency at rest was elevated, and there was a rightward shift in the median of the relative burst amplitude distribution ( P < 0.05) in Hypo. During LBNP, the larger rise in total MSNA during Hypo versus Normo was due to greater increases in relative burst amplitude with no difference in the burst frequency response. The MSNA burst frequency response to LBNP was shifted to a higher position on the same MSNA-CVP curve during Hypo compared with Normo. In contrast, the Hypo burst amplitude response was shifted to a new curve with a slope that was similar to the Normo relationship. These data support the use of probability distribution analysis to examine intraindividual differences in baseline and reflex-mediated increases in MSNA burst amplitude. Furthermore, the differential effect of hypovolemia on the responses of burst frequency and amplitude during graded LBNP suggests that burst frequency data alone may not adequately represent reflex control of sympathetic outflow.