bluuebunny/crossref_metadata_2025 huggingface dataset:
Estradiol reduces F2α-isoprostane production in cultured human endothelial cells
Authors: Hermenegildo Carlos , Garcı́a-Martı́nez Marı́a Cinta , Tarı́n Juan J. , Cano Antonio
Published: March 2015

Free radical-generated F2α-isoprostanes are a group of compounds with vasoconstrictor properties. To investigate whether estradiol exerts antioxidant actions modifying F2α-isoprostane production, cultured human umbilical vein endothelial cells were exposed to estradiol and other compounds and F2α-isoprostanes were measured in culture medium. Exposure to 1 and 10 nM estradiol for 24 h reduced F2α-isoprostane production by 36 and 49%, respectively ( P < 0.001 vs. control). Exposure to antiestrogens alone (ICI-182780 or EM-652) slightly reduced F2α-isoprostanes ( P < 0.05 vs. control), but much less than exposure to estradiol ( P < 0.05). ICI-182780 reversed the estradiol-induced reduction of F2α-isoprostane concentration ( P < 0.05). Along with time-course analysis, these results suggest that estradiol effects were mediated through estrogen receptor-dependent and -independent mechanisms. Progestogens alone (progesterone or medroxyprogesterone acetate) did not modify F2α-isoprostane production at any of the tested concentrations (1, 10, and 100 nM). Progesterone completely reversed estradiol-induced reduction of F2α-isoprostane production ( P < 0.05 vs. control and estradiol), but medroxyprogesterone acetate did not ( P < 0.05 vs. control).
Behavior of ectopic surface: effects of β-adrenergic stimulation and uncoupling
Authors: Arutunyan Ara , Pumir Alain , Krinsky Valentin , Swift Luther , Sarvazyan Narine
Published: March 2015

By using both experimental and theoretical means, we have addressed the progression of ectopic activity from individual cardiac cells to a multicellular two-dimensional network. Experimental conditions that favor ectopic activity have been created by local perfusion of a small area of cardiomyocyte network (I-zone) with an isoproterenol-heptanol containing solution. The application of this solution initially slowed down and then fully blocked wave propagation inside the I-zone. After a brief lag period, ectopically active cells appeared in the I-zone, followed by evolution of the ectopic clusters into slowly propagating waves. The changing pattern of colliding and expanding ectopic waves confined to the I-zone persisted for as long as the isoproterenol-heptanol environment was present. On restoration of the control environment, the ectopic waves from the I-zone broke out into the surrounding network causing arrhythmias. The observed sequence of events was also modeled by FitzHugh-Nagumo equations and included a cell's arrangement of two adjacent square regions of 20 × 20 cells. The control zone consisted of well-connected, excitable cells, and the I-zone was made of weakly coupled cells (heptanol effect), which became spontaneously active as time evolved (isoproterenol effect). The dynamic events in the system have been studied numerically with the use of a finite difference method. Together, our experimental and computational data have revealed that the combination of low coupling, increased excitability, and spatial heterogeneity can lead to the development of ectopic waves confined to the injured network. This transient condition appears to serve as an essential step for the ectopic activity to “mature” before escaping into the surrounding control network.
Potassium (BKCa) currents are reduced in microvascular smooth muscle cells from insulin-resistant rats
Authors: Dimitropoulou Christiana , Han Guichun , Miller Allison W. , Molero Mariela , Fuchs Leslie C. , White Richard E. , Carrier Gerald O.
Published: March 2015

Insulin resistance (IR) syndrome is associated with impaired vascular relaxation; however, the underlying pathophysiology is unknown. Potassium channel activation causes vascular smooth muscle hyperpolarization and relaxation. The present study determined whether a reduction in large conductance calcium- and voltage-activated potassium (BKCa) channel activity contributes to impaired vascular relaxation in IR rats. BKCachannels were characterized in mesenteric microvessels from IR and control rats. Macroscopic current density was reduced in myocytes from IR animals compared with controls. In addition, inhibition of BKCachannels with tetraethylammonium (1 mM) or iberiotoxin (100 nM) was greater in myocytes from control (70%) compared with IR animals (∼20%). Furthermore, activation of BKCachannels with NS-1619 was three times more effective at increasing outward current in cells from control versus IR animals. Single channel and Western blot analysis of BKCachannels revealed similar conductance, amplitude, voltage sensitivity, Ca2+sensitivity, and expression density between the two groups. These data provide the first direct evidence that microvascular potassium currents are reduced in IR and suggest a molecular mechanism that could account for impaired vascular relaxation in IR.
Relationship between lymph and tissue hyaluronan in skin and skeletal muscle
Authors: Armstrong Shayn E. , Bell Donald R.
Published: March 2015

The size of hyaluronan was compared between tissue and lymph using a combination of agarose gel electrophoresis and radiometric assay. Prenodal lymph was collected from heel skin and the gastrocnemius muscle in anesthetized rabbits. The major fraction of hyaluronan in both tissues had a molecular weight >4 million. Lymph contained primarily low-molecular-weight hyaluronan (<0.79 × 106), which was absent from tissue. Volume loading produced a preferential increase in the flux of low-molecular-weight hyaluronan, indicating that tissue contains a small quantity of mobile, low-molecular-weight hyaluronan. The maximum daily removal of hyaluronan by lymph was <1% of the tissue content. The amount of lysosomal hyaluronidase activity in tissue was more than enough to account for a rapid turnover of hyaluronan. The data support the conclusion that lymph drainage is not significant in the normal catabolism of hyaluronan and may represent a small amount that becomes detached from the pericellular and extracellular matrixes.
Comparison of time- and voltage-dependent K+ currents in myocytes from left and right atria of adult mice
Authors: Lomax Alan E. , Kondo Colleen S. , Giles Wayne R.
Published: March 2015

Consistent differences in K+ currents in left and right atria of adult mouse hearts have been identified by the application of current- and voltage-clamp protocols to isolated single myocytes. Left atrial myocytes had a significantly ( P < 0.05) larger peak outward K+ current density than myocytes from the right atrium. Detailed analysis revealed that this difference was due to the rapidly activating sustained K+ current, which is inhibited by 100 μM 4-aminopyridine (4-AP); this current was almost three times larger in the left atrium than in the right atrium. Accordingly, 100 μM 4-AP caused a significantly ( P < 0.05) larger increase in action potential duration in left than in right atrial myocytes. Inward rectifier K+ current density was also significantly ( P < 0.05) larger in left atrial myocytes. There was no difference in the voltage-dependent L-type Ca2+ current between left and right atria. As expected from this voltage-clamp data, the duration of action potentials recorded from single myocytes was significantly ( P < 0.05) shorter in myocytes from left atria, and left atrial tissue was found to have a significantly ( P < 0.05) shorter effective refractory period than right atrial tissue. These results reveal similarities between mice and other mammalian species where the left atrium repolarizes more quickly than the right, and provide new insight into cellular electrophysiological mechanisms responsible for this difference. These findings, and previous results, suggest that the atria of adult mice may be a suitable model for detailed studies of atrial electrophysiology and pharmacology under control conditions and in the context of induced atrial rhythm disturbances.
ATP stimulates chemically sensitive and sensitizes mechanically sensitive afferents
Authors: Li Jianhua , Sinoway Lawrence I.
Published: March 2015

We examined whether ATP stimulation of P2X purinoceptors would raise blood pressure in decerebrate cats. Femoral arterial injection of the P2X receptor agonist α,β-methylene ATP into the blood supply of the triceps surae muscle induced a dose-dependent increase in arterial blood pressure. The maximal increase in mean arterial pressure (MAP) evoked by 0.1, 0.2, and 0.5 mM α,β-methylene ATP (0.5 ml/min injection rate) was 6.2 ± 2.5, 22.5 ± 4.4, and 35.2 ± 3.9 mmHg, respectively. The P2X receptor antagonist pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid (2 mM ia) attenuated the increase in MAP elicited by intra-arterial α,β-methylene ATP (0.5 mM), whereas the P2Y receptor antagonist reactive blue 2 (2 mM ia) did not affect the MAP response to α,β-methylene ATP. In a second group of experiments, we tested the hypothesis that ATP acting through P2X receptors would sensitize muscle afferents and, thereby, augment the blood pressure response to muscle stretch. Two kilograms of muscle stretch evoked a 26.5 ± 4.3 mmHg increase in MAP. This MAP response was enhanced when 2 mM ATP or 0.1 mM α,β-methylene ATP (0.5 ml/min) was arterially infused 10 min before muscle stretch. Furthermore, this effect of ATP on the pressor response to stretch was attenuated by 2 mM pyridoxal phosphate-6-azophenyl-2′,4′-disulfonic acid ( P < 0.05) but not by the P1 purinoceptor antagonist 8-( p-sulfophenyl)-theophylline (2 mM). These data indicate that activation of ATP-sensitive P2X receptors evokes a skeletal muscle afferent-mediated pressor response and that ATP at relatively low doses enhances the muscle pressor response to stretch via engagement of P2X receptors.
Impact of coronary vasa vasorum functional structure on coronary vessel wall perfusion distribution
Authors: Gössl M. , Malyar N. M. , Rosol M. , Beighley P. E. , Ritman E. L.
Published: March 2015

Noncoronary vasa vasorum have been described as networks of microvessels in the wall of arteries and veins. However, we have shown, using microcomputerized tomography (micro-CT) imaging methods, that porcine coronary vasa vasorum have a tree-like branching structure similar to the vasculature in general. In this study, we elucidate functional aspects of coronary vasa vasorum perfusion territories. Three pig hearts were injected with radiopaque Microfil via the coronary sinus to fill the left anterior descending coronary arteries (LADs) retrogradely at atmospheric pressure. In three other hearts, LADs were injected antegradely at 100-mmHg pressure via the left main carotid artery. Additionally, six LADs were injected in vivo with a suspension of 100- or 300-μm-diameter microspheres before harvesting of the hearts and injection of the LADs with Microfil. All harvested LADs were scanned intact with micro-CT (20 μm cubic voxels). The spatial density of vasa vasorum (no. of vasa/mm2) was measured in 20-μm-thick cross sections (at 0.4-mm intervals). Retrogradely injected LADs showed high and uniformly distributed vasa vasorum densities in the adventitia (means ± SE; 5.38 ± 0.09 vs. 3.58 ± 0.1 vasa/mm2 in antegradely prepared LADs; P < 0.001). Antegradely prepared LADs showed patchy distributed, low-vasa-vasorum-density territories especially on the myocardial side of the coronary artery wall (epicardial density: 4.29 ± 0.13 vasa/mm2 vs. myocardial density: 2.80 ± 0.1 vasa/mm2, P < 0.001). Microembolization reduced vasa vasorum densities significantly (100-μm-diameter microspheres: 3.26 ± 0.07 vasa/mm2, P < 0.05; 300-μm-diameter microspheres: 2.66 ± 0.07 vasa/mm2, P < 0.001 vs. antegrade controls) and increased the size of low-vasa-vasorum-density territories. We conclude that coronary vasa vasorum are functional endarteries not connected via a plexus. This characteristic may have a significant impact on the spatial distribution of perfusion and drainage of the coronary vessel wall.
Trauma induced by nontraumatic coronary devices and its impact on vascular reactivity and morphology
Authors: Strotmann Jörg M. , Bauersachs Johann , Fraccarollo Daniela , Kirchengast Michael , Schnabel Philipp A. , Sykora Jaromir , Ertl Georg , Voelker Wolfram
Published: March 2015

This study evaluated the impact of low-pressure balloon devices on coronary morphology and function. An active coronary perfusion catheter (2.5-mm balloon diameter, inflation with 1 bar for 30 min) was placed in the left anterior descending coronary artery of 12 German landrace pigs under general anesthesia. After 3 mo, coronary segments with balloon contact were compared with control segments taken from the right coronary artery as to histology, vascular reactivity, and expression of endothelial nitric oxide synthase. Thirty-three balloon treated segments were analyzed. Twenty of these segments (61%) showed neointima formation. In these segments endothelium-independent relaxation induced by sodium nitroprusside was preserved. However, endothelium-dependent bradykinin-induced relaxation was significantly attenuated compared with both the control segments and the balloon-treated segments without neointima formation. In >60% of the ballooned arterial segments examined, low-pressure balloon devices induced neointima formation accompanied by reduced endothelium-dependent relaxation. Thus interventions with so-called nontraumatic coronary devices can induce relevant vascular injury, with potential adverse clinical consequences.
Oral antioxidant therapy improves endothelial function in Type 1 but not Type 2 diabetes mellitus
Authors: Beckman Joshua A. , Goldfine Allison B. , Gordon Mary Beth , Garrett Leslie A. , Keaney John F. , Creager Mark A.
Published: March 2015

Oxidative stress decreases the bioavailability of endothelium-derived nitric oxide in diabetic patients. We investigated whether impaired endothelium-dependent vasodilation (EDV) in diabetes can be improved by long-term administration of oral antioxidants. Forty-nine diabetic subjects [26 Type 1 (T1) and 23 Type 2 (T2)] and 45 matched healthy control subjects were randomized to receive oral vitamin C (1,000 mg) and vitamin E (800 IU) daily or matching placebo for 6 mo. Vascular ultrasonography was used to determine brachial artery EDV and endothelium-independent vasodilation (EIV). EDV was decreased in both T1 (4.9 ± 0.9%, P = 0.015) and T2 (4.1 ± 1.0%, P < 0.01) subjects compared with control subjects (7.7 ± 0.7%). EIV was decreased in T2 (15.0 ± 1.2%, P < 0.01) but not T1 subjects (18.5 ± 2.3%, P = 0.3) compared with controls (21.8 ± 1.8%). Administration of antioxidant vitamins increased EDV in T1 (by 3.4 ± 1.4%, P = 0.023) but not T2 subjects (by 0.5. ± 0.4%, P = 0.3). Antioxidant therapy had no signigicant affect on EIV. Oral antioxidant therapy improves EDV in T1 but not T2 diabetes. These results are consistent with the lack of clinical benefit in studies that have included primarily T2 diabetic patients.
Poincaré plot interpretation using a physiological model of HRV based on a network of oscillators
Authors: Brennan Michael , Palaniswami Marimuthu , Kamen Peter
Published: March 2015

In this paper, we develop a physiological oscillator model of which the output mimics the shape of the R-R interval Poincaré plot. To validate the model, simulations of various nervous conditions are compared with heart rate variability (HRV) data obtained from subjects under each prescribed condition. For a variety of sympathovagal balances, our model generates Poincaréplots that undergo alterations strongly resembling those of actual R-R intervals. By exploiting the oscillator basis of our model, we detail the way that low- and high-frequency modulation of the sinus node translates into R-R interval Poincaré plot shape by way of simulations and analytic results. With the use of our model, we establish that the length and width of a Poincaré plot are a weighted combination of low- and high-frequency power. This provides a theoretical link between frequency-domain spectral analysis techniques and time-domain Poincaré plot analysis. We ascertain the degree to which these principles apply to real R-R intervals by testing the mathematical relationships on a set of data and establish that the principles are clearly evident in actual HRV records.
CYP4A metabolites of arachidonic acid and VEGF are mediators of skeletal muscle angiogenesis
Authors: Amaral Sandra L. , Maier Kristopher G. , Schippers Daniela N. , Roman Richard J. , Greene Andrew S.
Published: March 2015

Vascular endothelial growth factor (VEGF) has been implicated in angiogenesis induced by electrical stimulation in skeletal muscle. Less is known about the role of arachidonic acid metabolites in the control of growth of blood vessels in vivo. The present study examined the role of 20-hydroxyeicosatetraenoic acid (20-HETE) on the angiogenesis induced by electrical stimulation in skeletal muscle. The tibialis anterior and extensor digitorum longus muscles of rats were stimulated for 7 days. Electrical stimulation significantly increased the 20-HETE formation and angiogenesis in the muscles, which was blocked by chronic treatment with N-hydroxy- N′-(4-butyl-2-methylphenol)formamidine (HET0016) or 1-aminobenzotriazole (ABT). Chronic treatment with either HET0016 or ABT did not block the increases in VEGF protein expression in both muscles. To analyze the role of VEGF on 20-HETE formation, additional rats were treated with VEGF-neutralizing antibody (VEGF Ab). VEGF Ab blocked the increases of 20-HETE formation induced by stimulation. These results place 20-HETE in the downstream signaling pathway for angiogenesis and show that both VEGF and 20-HETE are involved in the angiogenesis induced by electrical stimulation in skeletal muscle.
Apoptotic cascade initiated by angiotensin II in neonatal cardiomyocytes: role of DNA damage
Authors: Grishko Valentina , Pastukh Viktor , Solodushko Viktoriya , Gillespie Mark , Azuma Junichi , Schaffer Stephen
Published: March 2015

Angiotensin II contributes to ventricular remodeling by promoting both cardiac hypertrophy and apoptosis; however, the mechanism underlying the latter phenomenon is poorly understood. One possibility that has been advanced is that angiotensin II activates NADPH oxidase, generating free radicals that trigger apoptosis. In apparent support of this notion, it was found that angiotensin II-mediated apoptosis in the cardiomyocyte is blocked by the NADPH oxidase inhibitor diphenylene iodonium. However, three lines of evidence suggest that peroxynitrite, rather than superoxide, is responsible for angiotensin II-mediated DNA damage and apoptosis. First, the inducible nitric oxide inhibitor aminoguanidine prevents angiotensin II-induced DNA damage and apoptosis. Second, based on ligation-mediated PCR, the pattern of angiotensin II-induced DNA damage resembles peroxynitritemediated damage rather than damage caused by either superoxide or nitric oxide. Third, angiotensin II activates p53 through the phosphorylation of Ser15 and Ser20, residues that are commonly phosphorylated in response to DNA damage. It is proposed that angiotensin II promotes the oxidation of DNA, which in turn activates p53 to mediate apoptosis.
Inhibitory effect of recombinant iNOS gene expression on vasomotor function of canine basilar artery
Authors: Eguchi Daihiko , d'Uscio Livius V. , Wambi Chris , Weiler Deborah , Kovesdi Imre , O'Brien Timothy , Katusic Zvonimir S.
Published: March 2015

The present study was designed to determine the effect of recombinant inducible nitric oxide (NO) synthase (iNOS) gene expression on vasomotor function in cerebral arteries. Isolated canine basilar arteries were exposed ex vivo (30 min at 37°C) to an adenoviral vector [107, 108, or 109plaque-forming units (pfu)/ml] encoding either the iNOS gene or the β-galactosidase reporter gene. Twenty-four hours after transduction, Western blot analysis demonstrated expression of iNOS protein only in iNOS (109pfu/ml)-transduced arteries. Immunohistochemical analysis localized iNOS expression predominantly in adventitia. Vascular reactivity of isolated basilar arteries was studied by isometric force recording. Concentration-response curves to UTP (10−9–10−3M) and diethylaminodiazen-1-ium-1,2-dioate (10−10–10−5M) were significantly shifted to the right in iNOS gene (109pfu/ml)-transduced rings compared with control and β-galactosidase-transduced rings ( P < 0.05, n = 5–6). Endothelium-dependent relaxation to bradykinin was significantly attenuated in iNOS-transduced rings ( P < 0.001, n = 8). The basal level of cGMP and superoxide anion (O[Formula: see text]·) production were elevated in iNOS-transduced rings ( P < 0.05, n = 7 for cGMP; P < 0.01, n = 6–9 for O[Formula: see text]· production). Our results suggest that expression of recombinant iNOS in cerebral arteries reduces vasomotor reactivity to both vasoconstrictor and vasodilator agonists. Attenuation of contractions is most likely due to functional antagonism between UTP and cGMP. Reduction of endothelium-dependent relaxation to bradykinin appears to be mediated in part by reduced reactivity of smooth muscle cells to NO.
β-Adrenergic receptor blockade impairs NO-dependent dilation of large coronary arteries during exercise
Authors: Okajima Masaki , Takamura Masayuki , Véquaud Philippe , Parent Robert , Lavallée Michel
Published: March 2015

Shear stress-dependent nitric oxide (NO) formation prevents immoderate vascular constriction. We examined whether shear stress-dependent NO formation limits exercise-induced coronary artery constriction after β-adrenergic receptor blockade in dogs. Control exercise led to increases ( P < 0.01) in coronary blood flow (CBF) by 38 ± 5 ml/min from 41 ± 5 ml/min and in the external diameter of epicardial coronary arteries (CD) by 0.24 ± 0.03 mm from 3.33 ± 0.20 mm. CD and shear stress were linearly related. After propranolol, CD fell ( P < 0.01) during exercise (0.08 ± 0.03 from 3.23 ± 0.19 mm), and the slope of the relationship between CD and shear stress was reduced ( P< 0.01). This slope was not further altered by the additional blockade of NO formation. In propranolol-treated resting dogs, flow-dependent effects of intracoronary adenosine to mimic exercise-induced increases in shear stress (after propranolol) led to increases ( P< 0.01) in CD (0.09 ± 0.02 from 3.68 ± 0.27 mm). Thus both shear stress-dependent NO formation and β-adrenergic receptor activation are required to cause CD dilation during exercise. Suppression of β-adrenergic receptor activation leads to impaired shear stress-dependent NO formation and allows α-adrenergic constriction to become dominant.
Aldosterone stimulates proliferation of cardiac fibroblasts by activating Ki-RasA and MAPK1/2 signaling
Authors: Stockand James D. , Meszaros J. Gary
Published: March 2015

Aldosterone plays a pathological role in cardiac fibrosis by directly affecting cardiac fibroblasts. Understanding of the cellular mechanisms of aldosterone action in cardiac fibroblasts, however, is rudimentary. One possibility is that aldosterone promotes proliferation of cardiac fibroblasts by activating specific cellular signaling cascades. The current study tests whether aldosterone stimulates proliferation of isolated adult rat cardiac myofibroblasts (RCF) by activating Kirsten Ras (Ki-RasA) and its effector, the MAPK1/2 cascade. Aldosterone (10 nM) significantly increased RCF proliferation. This action was sensitive to the mineralocorticoid receptor (MR) antagonist spironolactone. Expression of MR in RCF and the whole rat heart was confirmed by immunoblotting. Aldosterone significantly increased absolute and active (GTP bound) Ki-RasA levels in RCF. Aldosterone, in addition, significantly increased phospho-c-Raf and phospho-MAPK1/2. The effects of aldosterone on Ki-RasA and phospho-c-Raf proteins were inhibited by spironolactone but not RU-486, suggesting that aldosterone acts via MR. Inhibitors of MEK1/2 and c-Raf prevented aldosterone-induced activation of MAPK1/2 and proliferation. These results show that aldosterone directly increases RCF proliferation through MR-dependent activation of Ki-RasA and its effector, the MAPK1/2 cascade. Activation of cardiac fibroblasts through such a cascade may play a role in the pathological actions exerted by aldosterone on the heart.
Calcium dynamics in the failing heart: restoration by β-adrenergic receptor blockade
Authors: Plank David M. , Yatani Atsuko , Ritsu Honda , Witt Sandra , Glascock Betty , Lalli M. Jane , Periasamy Muthu , Fiset Celine , Benkusky Nancy , Valdivia Héctor H. , Sussman Mark A.
Published: March 2015

Changes in calcium (Ca2+) regulation contribute to loss of contractile function in dilated cardiomyopathy. Clinical treatment using β-adrenergic receptor antagonists (β-blockers) slows deterioration of cardiac function in end-stage heart failure patients; however, the effects of β-blocker treatment on Ca2+ dynamics in the failing heart are unknown. To address this issue, tropomodulin-overexpressing transgenic (TOT) mice, which suffer from dilated cardiomyopathy, were treated with a nonselective β-receptor blocker (5 mg · kg-1 · day-1 propranolol) for 2 wk. Ca2+ dynamics in isolated cardiomyocytes of TOT mice significantly improved after treatment compared with untreated TOT mice. Frequency-dependent diastolic and Ca2+ transient amplitudes were returned to normal in propranolol-treated TOT mice and but not in untreated TOT mice. Ca2+ kinetic measurements of time to peak and time decay of the caffeine-induced Ca2+ transient to 50% relaxation were also normalized. Immunoblot analysis of untreated TOT heart samples showed a 3.6-fold reduction of sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), whereas Na+/Ca2+ exchanger (NCX) concentrations were increased 2.6-fold relative to nontransgenic samples. Propranolol treatment of TOT mice reversed the alterations in SERCA and NCX protein levels but not potassium channels. Although restoration of Ca2+ dynamics occurred within 2 wk of β-blockade treatment, evidence of functional improvement in cardiac contractility assessed by echocardiography took 10 wk to materialize. These results demonstrate that β-adrenergic blockade restores Ca2+ dynamics and normalizes expression of Ca2+-handling proteins, eventually leading to improved hemodynamic function in cardiomyopathic hearts.
Thyroid hormone regulation of cardiac bioenergetics: role of intracellular creatine
Authors: Queiroz Marcia Silva , Shao Yvonne , Berkich Deborah A. , Lanoue Kathryn F. , Ismail-Beigi Faramarz
Published: March 2015

The effect of thyroid hormone (T3) on the content of myocardial creatine (Cr), Cr phosphate (CrP), and high-energy adenine nucleotides and on cardiac function was examined. In the hearts of control and T3-treated rats perfused in vitro, while “low” and “high” contractile work was performed, T3 treatment resulted in a ∼50% reduction in CrP, Cr, total Cr content (Cr + CrP), and in the CrP-to-Cr ratio. In addition, there was a slight fall in myocardial content of ATP and a large rise in calculated free ADP (fADP), resulting in a significant decrease in the ATP-to-fADP ratio in the hearts of hyperthyroid compared with euthyroid rats. Moreover, there was a substantial decrease in the level of ATP in hearts of T3-treated rats under high work conditions. Importantly, the ratio of cardiac work to oxygen consumption was not altered by thyroid status. Treatment with T3 also resulted in an almost threefold reduction in the content of Na+/Cr transporter mRNA in the ventricular myocardium and skeletal muscle but not in the brain. We conclude with the following: 1) changes in the expression of the Na+/Cr transporter mRNA correlate with Cr + CrP in the myocardium; 2) hearts of hyperthyroid rats contain lower levels of ATP and higher levels of fADP under both low and high work conditions but no reduction in efficiency of work output; and 3) the reduction in Cr and ATP in hearts of hyperthyroid rats may be the basis for the reduced maximal work capacity of the hyperthyroid heart.
Simultaneous in situ monitoring of intracellular Ca2+and NO in endothelium of coronary arteries
Authors: Yi Fu-Xian , Zhang Andrew Y. , Campbell William B. , Zou Ai-Ping , van Breemen Cornelis , Li Pin-Lan
Published: March 2015

We developed an in situ assay system to simultaneously monitor intracellular Ca2+concentration ([Ca2+]i, fura 2 as indicator) and nitric oxide (NO) levels [4,5-diaminofluorescein as probe] in the intact endothelium of small bovine coronary arteries by using a fluorescent microscopic imaging technique with high-speed wavelength switching. Bradykinin (BK; 1 μM) stimulated a rapid increase in [Ca2+]ifollowed by an increase in NO production in the endothelial cells. The protein tyrosine phosphatase inhibitor phenylarsine oxide (PAO; 10 μM) induced a gradual, small increase in [Ca2+]iand a slow increase in intracellular NO levels. Removal of extracellular Ca2+and depletion of Ca2+stores completely blocked BK-induced increase in NO production but had no effect on PAO-induced NO production. However, a further reduction of [Ca2+]iby application of BAPTA-AM or EGTA with ionomycin abolished the PAO-induced NO increase. These results indicate that a simultaneous monitoring of [Ca2+]iand intracellular NO production in the intact endothelium is a powerful tool to study Ca2+-dependent regulation of endothelial nitric oxide synthase, which provides the first direct evidence for a permissive role of Ca2+in tyrosine phosphorylation-induced NO production.
Local vascular responses affecting blood flow in postural tachycardia syndrome
Authors: Stewart Julian M. , Medow Marvin S. , Montgomery Leslie D.
Published: March 2015

Postural tachycardia syndrome (POTS) is defined by orthostatic intolerance associated with abnormal upright tachycardia. Some patients have defective peripheral vasoconstriction and increased calf blood flow. Others have increased peripheral arterial resistance and decreased blood flow. In 14 POTS patients (13–19 yr) evenly subdivided among low-flow POTS (LFP) and high-flow POTS (HFP) we tested the hypothesis that myogenic, venoarteriolar, and reactive hyperemic responses are abnormal. We used venous occlusion plethysmography to measure calf venous pressure and blood flow in the supine position and when the calf was lowered by 40 cm to evoke myogenic and venoarteriolar responses and during venous hypertension by 40-mmHg occlusion to evoke the venoarteriolar response. We measured calf reactive hyperemia with plethysmography and cutaneous laser-Doppler flowmetry. Baseline blood flow in LFP was reduced compared with HFP and control subjects (0.8 ± 0.2 vs. 4.4 ± 0.5 and 2.7 ± 0.4 ml · min–1· 100 ml–1) but increased during leg lowering (1.2 ± 0.5 ml · min–1· 100 ml–1) while decreasing in the others. Baseline peripheral arterial resistance was increased in LFP and decreased in HFP compared with control subjects (39 ± 13 vs. 15 ± 3 and 22 ± 5 mmHg · ml–1· 100 ml · min) but decreased to 29 ± 13 mmHg · ml–1· 100 ml · min in LFP during venous hypertension. Resistance increased in the other groups. Maximum calf hyperemic flow and cutaneous flow were similar in all subjects. The duration of hyperemic blood flow was curtailed in LFP compared with either control or HFP subjects (plethysmographic time constant = 20 ± 2 vs. 29 ± 4 and 28 ± 4 s; cutaneous time constant = 60 ± 25 vs. 149 ± 53 s in controls). Local blood flow regulation in low-flow POTS is impaired.
Macrophage migration inhibitory factor is a cardiac-derived myocardial depressant factor
Authors: Garner Leslie B. , Willis Monte S. , Carlson Deborah L. , DiMaio J. Michael , White Michael D. , White D. Jean , Adams Glenn A. , Horton Jureta W. , Giroir Brett P.
Published: March 2015

Macrophage migration inhibitory factor (MIF) is a pluripotent proinflammatory cytokine that is ubiquitously expressed in organs, including the heart. However, no specific role for MIF in modulating cardiac performance has yet been described. Therefore, we examined cardiac MIF expression in mice after LPS challenge (4 mg/kg) and tested the hypothesis that MIF is a mediator of LPS-induced cardiac dysfunction. Western blots of whole heart lysates, as well as immunohistochemistry, documented constitutive MIF protein expression in the heart. Cardiac MIF protein levels significantly decreased after LPS challenge, reaching a nadir at 12 h, and then returned to baseline by 24 h. This pattern was consistent with MIF release from cytoplasmic stores after endotoxin challenge. After release of protein, MIF mRNA levels increased 24–48 h postchallenge. To determine the functional consequences of MIF release, we treated LPS-challenged mice with anti-MIF neutralizing antibodies or isotype control antibodies. Anti-MIF-treated animals had significantly improved cardiac function, as evidenced by a significant improvement in left ventricular (LV) fractional shortening percentage at 8, 12, 24, and 48 h after endotoxin challenge. In support of these findings, perfusion of isolated beating mouse hearts (Langendorff preparation) with recombinant MIF (20 ng/ml) led to a significant decrease in both systolic and diastolic performance [LV pressure (LVP), positive and negative first derivative of LVP with respect to time, and rate of LVP rise at developed pressure of 40 mmHg]. This study demonstrates that MIF mediates LPS-induced cardiac dysfunction and suggests that MIF should be considered a pharmacological target for the treatment of cardiac dysfunction in sepsis and potentially other cardiac diseases.
Endothelial vasodilator production by uterine and systemic arteries. IX. eNOS gradients in cycling and pregnant ewes
Authors: Joyce Jeffrey M. , Phernetton Terrance M. , Shaw Cynthia E. , Modrick Mary L. , Magness Ronald R.
Published: March 2015

10.1152/ajpheart.00438.2001.—The follicular phase (FOL) and pregnancy exhibit increases in uterine blood flow (UBF), estrogen levels, and uterine artery (UA) endothelial nitric oxide synthase (eNOS) expression. UA branching within the mesometrium increases the total vascular cross-sectional area, which reduces the vascular perfusion pressure gradient, thus locally decreasing the blood flow velocity. Shear stress (SS) activates eNOS and may be associated with UBF elevations during FOL and pregnancy. We hypothesized that regional differences in eNOS responses are observed with both decreases in vessel diameter and during the ovarian cycle and pregnancy. Endothelial isolated proteins were collected from renal (RA) and internal iliac arteries (II) as well as from primary (UA 1°), secondary (UA 2°), and tertiary (UA 3°) UA branches of nonpregnant luteal phase (LUT; n = 6) and FOL ( n = 6) as well as midpregnant (MP; 82 ± 1 days gestation, n = 6) and late pregnant (LP; 127 ± 3 days gestation, n= 6) ewes (term = 145 ± 3 days gestation) for Western blot analysis. LUT RA, II, and UA 1° eNOS levels were similar. There was a 60.7 ± 9.8% reduction in eNOS expression in UA 2° and UA 3°. A similar decreasing eNOS regional expression gradient was observed in LP ewes. No eNOS regional expression gradient was observed in FOL or MP ewes because eNOS increased in UA 2° and UA 3°. In UA 2° and UA 3°, MP > LP = FOL > LUT. Thus, with increasing UBF, FOL and pregnancy rises in SS may regulate eNOS protein expression in smaller diameter UAs. A decrease in LUT and LP UA 2° and UA 3° endothelial eNOS suggest a possible negative feedback mechanism due to downregulation of eNOS if SS is normalized.
Cardiac SR-coupled PP1 activity and expression are increased and inhibitor 1 protein expression is decreased in failing hearts
Authors: Gupta Ramesh C. , Mishra Sudhish , Rastogi Sharad , Imai Makoto , Habib Omar , Sabbah Hani N.
Published: March 2015

Type 1 protein phosphatase (PP1) is a negative regulator of cardiac function. However, studies on the status and regulation of sarcoplasmic reticulum (SR)-associated PP1 activity in failing hearts are limited. We studied PP1 activity and protein and mRNA expression of the catalytic subunit of PP1 (PP1C) and protein levels of PP1-specific inhibitors [inhibitor 1 (Inh-1) and inhibitor 2 (Inh-2)] in the left ventricular (LV) myocardium of 6 dogs with heart failure (HF; LV ejection fraction, 23 ± 2%) and 6 normal dogs. In failing LV tissue, PP1 activity values (expressed as pmol32P · min–1· mg of noncollagen protein–1) in the homogenate, crude membranes, cytosol, and purified SR were increased by 52, 54, 55, and 72%, respectively. Trypsin treatment released PP1 but not type 2A protein phosphatase from the SR. In the supernatant of trypsin-treated SR, PP1 activity was ∼24% higher in failing hearts than in normal control hearts. A similar increase in protein expression of PP1C was observed in the nontrypsinized SR. Heat-denatured phosphorylated SR inhibited PP1 activity by 30%, which suggests the presence of Inh-1 or -2 or both in the SR. With the use of a specific antibody, both Inh-1 and -2 proteins were found in the SR; the former was decreased by 56% in the failing SR, whereas the latter did not change. These results suggest that protein phosphatase activity bound to the SR is increased and is predominantly type 1. Increased SR-associated PP1 activity in failing hearts appears to be due partly to increased expression of PP1C and partly to reduced levels of Inh-1 but not Inh-2 protein. Thus inhibition of PP1 activity in the SR appears to be a potential therapeutic target for improving LV function in failing hearts, because it may lead to increased SR Ca2+uptake, which is impaired in failing hearts.
Cardiovascular mechanisms activated by microinjection of baclofen into NTS of conscious rats
Authors: Landulpho Carlos Daniel Almeida Pitanga , Dias Ana Carolina Rodrigues , Colombari Eduardo
Published: March 2015

The peripheral mechanisms responsible for pressor response produced by microinjections of baclofen (GABAB agonist) into the nucleus tractus solitarii (NTS) of conscious rats were studied. Bilateral microinjections of baclofen (10–1,000 pmol/100 nl) produced a dose-related increase in mean arterial pressure (MAP) and heart rate. The maximal response was observed after 15 min. Intravenous injection of prazosin decreased MAP to control levels. Subsequent treatment with Manning compound (vasopressin receptor antagonist; iv) produced an additional decrease in MAP. In a different group of rats, vasopressin antagonist was injected first and MAP was significantly decreased; however, it remained elevated compared with prebaclofen injection levels. Subsequent treatment with prazosin abolished the baclofen-induced pressor response. Reductions in baclofen-induced pressor response with prazosin treatment were followed by a reflex tachycardia in animals that received a 100 pmol/100 nl dose of baclofen. The tachycardia was not observed with a dose of 1,000 pmol/100 nl. The pressor response induced by microinjection of baclofen into the NTS of conscious rats may be produced by both increases in sympathetic tonus and vasopressin release.
Human coronary endothelial cells convert 14,15-EET to a biologically active chain-shortened epoxide
Authors: Fang Xiang , Weintraub Neal L. , Oltman Christine L. , Stoll Lynn L. , Kaduce Terry L. , Harmon Shawn , Dellsperger Kevin C. , Morisseau Christophe , Hammock Bruce D. , Spector Arthur A.
Published: March 2015

Cytochrome P-450 epoxygenase-derived epoxyeicosatrienoic acids (EETs) play an important role in the regulation of vascular reactivity and function. Conversion to the corresponding dihydroxyeicosatrienoic acids (DHETs) by soluble epoxide hydrolases is thought to be the major pathway of EET metabolism in mammalian vascular cells. However, when human coronary artery endothelial cells (HCEC) were incubated with3H-labeled 14,15-EET, chain-shortened epoxy fatty acids, rather than DHET, were the most abundant metabolites. After 4 h of incubation, 23% of the total radioactivity remaining in the medium was converted to 10,11-epoxy-hexadecadienoic acid (16:2), a product formed from 14,15-EET by two cycles of β-oxidation, whereas only 15% was present as 14,15-DHET. Although abundantly present in the medium, 10,11-epoxy-16:2 was not detected in the cell lipids. Exogenously applied3H-labeled 10,11-epoxy-16:2 was neither metabolized nor retained in the cells, suggesting that 10,11-epoxy-16:2 is a major product of 14,15-EET metabolism in HCEC. 10,11-Epoxy-16:2 produced potent dilation in coronary microvessels. 10,11-Epoxy-16:2 also potently inhibited tumor necrosis factor-α-induced production of IL-8, a proinflammatory cytokine, by HCEC. These findings implicate β-oxidation as a major pathway of 14,15-EET metabolism in HCEC and provide the first evidence that EET-derived chain-shortened epoxy fatty acids are biologically active.
Reactive oxygen species mediate arachidonic acid-induced dilation in porcine coronary microvessels
Authors: Oltman Christine L. , Kane Neal L. , Miller Francis J. , Spector Arthur A. , Weintraub Neal L. , Dellsperger Kevin C.
Published: March 2015

Reactive oxygen species (ROS) have been proposed to mediate vasodilation in the microcirculation. We investigated the role of ROS in arachidonic acid (AA)-induced coronary microvascular dilation. Porcine epicardial coronary arterioles (110 ± 4 μm diameter) were mounted onto pipettes in oxygenated Krebs buffer. Vessels were incubated with vehicle or 1 mM Tiron (a nonselective ROS scavenger), 250 U/ml polyethylene-glycolated (PEG)-superoxide dismutase (SOD; an [Formula: see text] scavenger), 250 U/ml PEG-catalase (a H2O2 scavenger), or the cyclooxygenase (COX) inhibitors indomethacin (10 μM) or diclofenac (10 μM) for 30 min. After endothelin constriction (30–60% of resting diameter), cumulative concentrations of AA (10–10–10–5 M) were added and internal diameters measured by video microscopy. AA (10–7 M) produced 37 ± 6% dilation, which was eliminated by the administration of indomethacin (4 ± 7%, P < 0.05) or diclofenac (–8 ± 8%, P < 0.05), as well as by Tiron (–4 ± 5%, P < 0.05), PEG-SOD (–10 ± 6%, P < 0.05), or PEG-catalase (1 ± 4%, P < 0.05). Incubation of small coronary arteries with [3H]AA resulted in the formation of prostaglandins, which was blocked by indomethacin. In separate studies in microvessels, AA induced concentration-dependent increases in fluorescence of the oxidant-sensitive probe dichlorodihydrofluorescein diacetate, which was inhibited by pretreatment with indomethacin or by SOD + catalase. We conclude that in porcine coronary microvessels, COX-derived ROS contribute to AA-induced vasodilation.
Angiotensin II attenuates functional hyperemia in the mouse somatosensory cortex
Authors: Kazama Ken , Wang Gang , Frys Kelly , Anrather Josef , Iadecola Costantino
Published: March 2015

We investigated whether angiotensin II (ANG II), a peptide that plays a central role in the genesis of hypertension, alters the coupling between synaptic activity and cerebral blood flow (CBF), a critical homeostatic mechanism that assures adequate cerebral perfusion to active brain regions. The somatosensory cortex was activated by stroking the facial whiskers in anesthetized C57BL/6J mice while local CBF was recorded by laser-Doppler flowmetry. Intravenous ANG II infusion (0.25 μg·kg–1·min–1) increased mean arterial pressure (MAP) from 82 ± 2 to 102 ± 3 mmHg ( P < 0.05) without affecting resting CBF ( P > 0.05). ANG II attenuated the CBF increase produced by whisker stimulation by 65% ( P < 0.05) but did not affect the response to hypercapnia or to neocortical application of the nitric oxide donor S-nitroso- N-acetyl penicillamine ( P > 0.05). The effect of ANG II on functional hyperemia persisted if the elevation in MAP was offset by controlled hemorrhage or prevented by topical application of the peptide to the activated cortex. ANG II did not reduce the amplitude of the P1 wave of the field potentials evoked by whisker stimulation ( P > 0.05). Infusion of phenylephrine increased MAP ( P > 0.05 from ANG II) but did not alter the functional hyperemic response ( P > 0.05). The data suggest that ANG II alters the coupling between CBF and neural activity. The mechanisms of the effect are not related to the elevation in MAP and/or to inhibition of the synaptic activity evoked by whisker stimulation. The imbalance between CBF and neural activity induced by ANG II may alter the homeostasis of the neuronal microenvironment and contribute to brain dysfunction during ANG II-induced hypertension.
Differential vasoconstrictions induced by angiotensin II: role of AT1 and AT2 receptors in isolated C57BL/6J mouse blood vessels
Authors: Zhou Yingbi , Dirksen Wessel P. , Babu Gopal J. , Periasamy Muthu
Published: March 2015

Genetically altered mice are increasingly used as experimental models. However, ANG II responses in mouse blood vessels have not been well defined. Therefore, the aim of this study was to determine the role of ANG II in regulating major blood vessels in C57/BL6J mice with isometric force measurements. Our results showed that in mouse abdominal aorta ANG II induced a concentration-dependent contraction (EC504.6 nM) with a maximum contraction of 75.1 ± 4.9% at 100 nM compared with that of 60 mM K+. Similarly, femoral artery also exhibited a contractile response of 76.0 ± 3.4% to the maximum concentration of ANG II (100 nM). In contrast, ANG II (100 nM)-induced contraction was significantly less in carotid artery (24.5 ± 6.6%) and only minimal (3.5 ± 0.31%) in thoracic aorta. The nitric oxide synthase inhibitor Nω-nitro-l-arginine methyl ester and the AT2 antagonist PD-123319 failed to enhance ANG II-induced contractions. However, an AT1 antagonist, losartan (10 μM), completely inhibited ANG II (100 nM) response in abdominal aorta and carotid artery. An AT1 agonist, [Sar1]-ANG II (100 nM), behaved similarly to ANG II (100 nM) in abdominal aorta and carotid artery. RT-PCR analyses showed that mouse thoracic aorta has a significantly lower AT1 mRNA level than abdominal aorta. These results demonstrate that major mouse vessels exhibit differential contractions to ANG II, possibly because of varied AT1 receptor levels.
Intraluminal pressure stimulates MAPK phosphorylation in arterioles: temporal dissociation from myogenic contractile response
Authors: Spurrell Brian E. , Murphy Timothy V. , Hill Michael A.
Published: March 2015

Members of the MAPK family of enzymes, p42/44 and p38, have been implicated in both the regulation of contractile function and growth responses in vascular smooth muscle. We determined whether such kinases are activated during the arteriolar myogenic response after increases in intraluminal pressure. Particular emphasis was placed on temporal aspects of activation to determine whether such phosphorylation events parallel the known time course for myogenic contraction. Experiments used single cannulated arterioles isolated from the cremaster muscle of rats with some vessels loaded with the fluorescent Ca2+-sensitive dye fura 2 (2 μM). The p42/44 inhibitor PD-98059 (50 μM) caused vasodilation but did not prevent pressure-induced myogenic constriction. The vasodilator response was accompanied by decreased smooth muscle intracellular Ca2+. Western blotting revealed a significant increase in the level of phosphorylation of p42/44 15 min after the application of a 30- to 100-mmHg pressure step. Phosphorylation of p42/44 was a late event that appeared to be temporally dissociated from contraction, which was complete within 1–5 min. EGF (80 nM) caused marked phosphorylation of p42/44 but only acted as a weak vasoconstrictor. The p38 inhibitor SB-203580 (10 μM) did not alter baseline diameter, nor did it prevent myogenic vasoconstriction. Consistent with these observations, SB-203580 did not cause a measurable change in intracellular Ca2+. The results demonstrate activation of the p42/44 class of MAPK resulting from increased transmural pressure. Such activation is, however, dissociated from the acute pressure-induced vasoconstrictor response in terms of time course and may represent the activation of compensatory, but parallel, pathways, including those related to growth and remodeling.