bluuebunny/crossref_metadata_2025 huggingface dataset:
Microtubules modulate cardiomyocyte β-adrenergic response in cardiac hypertrophy
Authors: Palmer Bradley M. , Valent Scott , Holder Emma L. , Weinberger Howard D. , Bies Roger D.
Published: December 2017

The role of microtubules in modulating cardiomyocyte β-adrenergic response was investigated in rats with cardiac hypertrophy. Male Sprague-Dawley rats underwent stenosis of the abdominal aorta (hypertensive, HT) or sham operation (normotensive, NT). Echocardiography and isolated left ventricular cardiomyocyte dimensions demonstrated cardiac hypertrophy in the HT rats after 30 wk. Cardiomyocyte microtubule fraction was assayed by high-speed centrifugation and Western blot. In contrast to previous reports of increased microtubules after acute pressure overload, microtubule fraction for HT was significantly lower than that for NT. Cardiomyocytes were exposed to either 1 μM colchicine, 10 μM taxol, or equivalent volume of vehicle. Colchicine decreased microtubules, and taxol increased microtubules in both groups. Cardiomyocyte cytosolic calcium ([Ca2+]c) and shortening/relaxation dynamics were assessed during exposure to increasing isoproterenol concentrations. The β-adrenergic response for these variables in the HT group was blunted compared with NT. However, increased microtubule assembly by taxol partially recovered the normal β-adrenergic response for time to peak [Ca2+]c, time to peak shortening, and mechanical relaxation variables. Microtubule assembly may play a significant role in determining cardiomyocyte β-adrenergic response in chronic cardiac hypertrophy.
Phospholipase A2is not responsible for lysophosphatidylcholine-induced damage in cardiomyocytes
Authors: Chen Min , Xiao Chun-Yang , Hashizume Hiroko , Abiko Yasushi
Published: December 2017

Lysophosphatidylcholine (LPC) is known to increase the intracellular concentration of Ca2+([Ca2+]i), leading to cell damage. In the present study we examined whether LPC activates phospholipase A2(PLA2) and whether the activation of PLA2is responsible for the LPC-induced cell damage in isolated rat cardiomyocytes. LPC (15 μM) produced an increase in [Ca2+]i, a change in cell shape from rod to round, and the release of creatine kinase (CK) accompanied by a significant elevation of the cellular level of nonesterified fatty acids (NEFA), especially arachidonic acid. Three PLA2inhibitors, 7,7-dimethyl-(5 Z,8 Z)-eicosadienoic acid (DEDA), 3-(4-octadecylbenzoyl)acrylic acid (OBAA), and manoalide, attenuated the LPC-induced accumulation of unsaturated NEFA to a similar degree. Nevertheless, whereas both DEDA and OBAA attenuated the LPC-induced increase in [Ca2+]i, change in cell shape, and release of CK, manoalide attenuated none of them. In the Ca2+-free solution, LPC did not increase [Ca2+]iwith significantly less accumulation of NEFA, but it changed the cell shape from rod to round and increased the release of CK. These results suggest that exogenous LPC increases the PLA2activity, which, however, may not be responsible for the LPC-induced damage in cardiomyocytes.
Effect of ANG II on pHi, [Ca2+]i, and contraction in rabbit ventricular myocytes from infarcted hearts
Authors: Skolnick Rita L. , Litwin Sheldon E. , Barry William H. , Spitzer Kenneth W.
Published: December 2017

In this study we examined Na+/H+exchange activity, Ca2+transients, and contractility in rabbit ventricular myocytes isolated from normal and chronically (8–12 wk) infarcted left ventricles. Myocytes from infarcted hearts (post-MI myocytes) were isolated from the peri-infarcted region of the left ventricle. Intracellular pH (pHi) and Ca2+ concentration ([Ca2+]i) were measured with the fluorescent pH indicators seminaphthorhodafluor 1 and fluo 3, respectively, and contractility was assessed from changes in cell shortening during field stimulation. Experiments were performed at extracellular pH 7.4 in the presence and absence (HEPES buffer) of CO2 and[Formula: see text]. Our findings demonstrate that 1) myocytes after myocardial infarction (post-MI) were significantly larger than normal, 2) post-MI hypertrophy was not accompanied by changes in non-CO2intracellular buffering power, 3) post-MI hypertrophy did not significantly affect the ability of Na+/H+exchange to mediate pHi recovery from intracellular acidosis, 4) the stimulatory effect of ANG II (100 nM) on Na+/H+exchange was significantly reduced in post-MI myocytes, 5) in[Formula: see text]-buffered solutions, ANG II did not significantly stimulate pHirecovery from acidosis in post-MI myocytes, 6) the angiotensin AT1 receptor mediates the stimulatory action of ANG II on Na+/H+exchange in normal and post-MI myocytes, and 7) the stimulatory effect of ANG II on the Ca2+ transient and contraction was blunted in post-MI myocytes bathed in HEPES-buffered solution. A suppressed ventricular responsiveness to ANG II may be beneficial in the intact myocardium by attenuating ATP consumption and by reducing intracellular Na+accumulation during ischemia-reperfusion.
Heart and lung VEGF mRNA expression in rats with monocrotaline- or hypoxia-induced pulmonary hypertension
Authors: Partovian Chohreh , Adnot Serge , Eddahibi Saadia , Teiger Emmanuel , Levame Micheline , Dreyfus Patrick , Raffestin Bernadette , Frelin Christian
Published: December 2017

Vascular endothelial growth factor (VEGF) is an endothelial cell-specific mitogen that is upregulated during exposure to hypoxia. In this study, we analyzed heart and lung VEGF mRNA expression and examined pulmonary vascular remodeling as well as myocardial capillary density in two rat models of pulmonary hypertension involving exposure to chronic hypoxia (CH) and treatment with monocrotaline (MCT), respectively. The rats were studied after 0.5, 1, 3, 15, and 30 days of exposure to 10% O2 or 1, 6, and 30 days after a subcutaneous MCT injection (60 mg/kg). Both CH and MCT induced pulmonary hypertension and hypertrophy of the right ventricle (RV) with increased RV weight and atrial natriuretic peptide mRNA expression. VEGF mRNA expression as assessed by Northern blot analysis was potently induced after 12 h of hypoxia in both the right and left ventricles. After prolonged exposure to hypoxia, VEGF mRNA returned to baseline in the left ventricle (LV) but remained increased in the RV, where it peaked after 30 days. In MCT rats, VEGF mRNA was unchanged in the LV but decreased by 50% in the RV and by 90% in the lungs after 30 days. VEGF mRNA remained unchanged in the lungs from CH rats. Pulmonary vascular remodeling was more pronounced in MCT than in CH rats. The number of capillaries per RV myocyte was increased in rats exposed to 30 days of hypoxia, whereas it remained unchanged in MCT rats despite a similar degree of RV hypertrophy. Our results suggest that the sustained increase in VEGF expression in the hypertrophied RV during CH may account for the increased number of capillaries per myocyte. In contrast, reduced VEGF expression in the lungs and RV of MCT rats may aggravate pulmonary vascular remodeling and compromise RV myocardial perfusion.
Rapid resetting of carotid baroreceptor reflex by afferent input from skeletal muscle receptors
Authors: Potts Jeffrey T. , Mitchell Jere H.
Published: December 2017

Resetting of the arterial baroreflex is mediated by central (central command) or peripheral (exercise pressor reflex) mechanisms. The purpose of this study was to determine the effect of somatosensory input from skeletal muscle receptors on resetting of the carotid baroreceptor reflex. Resetting of the baroreflex was determined by measuring carotid sinus threshold pressure (Pth) during a ramp protocol that consisted of a linear increase in sinus pressure from 50 to 250 mmHg at ∼3 mmHg/s. Experiments were performed in seven α-chloralose-anesthetized and vagotomized dogs. To determine the effect of skeletal muscle afferent input on resetting, electrically induced muscle contraction was used to activate mechanically and metabolically senstive afferent fibers, whereas passive stretch of the hindlimb was used to activate predominantly mechanically sensitive afferent fibers. Pth for heart rate (HR) and arterial blood pressure (BP) during the control ramp protocol was 110 ± 4 and 118 ± 7 mmHg, respectively. Electrically induced muscle contraction increased hindlimb tension (5.7 ± 0.4 kg) and significantly increased Pth-HR and Pth-BP above control (135 ± 6 and 141 ± 5 mmHg, respectively; P< 0.05). Muscle paralysis prevented the increase in Pth-HR and Pth-BP during ventral root stimulation (104 ± 7 and 116 ± 5 mmHg, respectively; P = not significant). Passive muscle stretch ( n = 3) increased hindlimb tension (5.9 ± 0.9 kg) and significantly increased Pth-BP (125 ± 21 vs. 159 ± 16 mmHg, control vs. contraction; P< 0.05). There was no difference in the magnitude of Pth resetting between muscle contraction or stretch. The present study demonstrates that activation of skeletal muscle afferent fibers, by either muscle contraction or stretch, increases Pth of the carotid baroreflex. Therefore, neural input from skeletal muscle receptors resets the carotid baroreflex in a manner similar to that ascribed by central command.
Pretreatment with tyrosine kinase inhibitors partially attenuates ischemic preconditioning in rat hearts
Authors: Fryer Ryan M. , Schultz Jo El J. , Hsu Anna K. , Gross Garrett J.
Published: December 2017

Ischemic preconditioning (IPC) confers cardioprotection against a prolonged ischemic insult. Tyrosine kinase (TK) inhibitors have been shown to attenuate IPC; however, it is unclear whether TK is involved in the initiation of and/or the maintenance of this phenomenon. Thus the hypothesis that TK acts primarily during the initiation of IPC was examined in a rat model of myocardial infarction. Hearts were subjected to 30 min of coronary artery occlusion and 2 h of reperfusion. IPC was elicited by three 5-min occlusions interspersed with 5 min of reperfusion before the prolonged occlusion period. Genistein, a nonspecific TK inhibitor, was administered before or during the final 2 min of the first or third occlusion period of IPC. Daidzein, an inactive structural analog of genistein, and lavendustin A, a more specific TK inhibitor, were also tested in this model. IPC markedly reduced infarct size expressed as a percentage of the area at risk compared with control (56.3 ± 2.8 to 7.1 ± 2.0%). This cardioprotection was attenuated by genistein pretreatment (5 mg/kg: 34.7 ± 2.2%, 10 mg/kg: 33.5 ± 5.9%). However, genistein administered during the first or third occlusion period of IPC did not significantly attenuate cardioprotection (10.3 ± 2.9% and 6.4 ± 2.0%). Lavendustin A (1.0 mg/kg) pretreatment also attenuated IPC (30.1 ± 2.2%), whereas daidzein (5 mg/kg) had no effect (7.9 ± 2.4%). These results suggest that activation of a TK is involved in the initiation but not the maintenance of IPC in the rat myocardium.
Temperature dependence of early and late currents in human cardiac wild-type and long Q-T ΔKPQ Na+ channels
Authors: Nagatomo Toshihisa , Fan Zheng , Ye Bin , Tonkovich Gayle S. , January Craig T. , Kyle John W. , Makielski Jonathan C.
Published: December 2017

Na+current ( I Na) through wild-type human heart Na+channels (hH1) is important for normal cardiac excitability and conduction, and it participates in the control of repolarization and refractoriness. I Na kinetics depend strongly on temperature, but I Na for hH1 has been studied previously only at room temperature. We characterized early I Na (the peak and initial decay) and late I Na of the wild-type hH1 channel and a mutant channel (ΔKPQ) associated with congenital long Q-T syndrome. Channels were stably transfected in HEK-293 cells and studied at 23 and 33°C using whole cell patch clamp. Activation and inactivation kinetics for early I Na were twofold faster at higher temperature for both channels and shifted activation and steady-state inactivation in the positive direction, especially for ΔKPQ. For early I Na (<24 ms), ΔKPQ decayed faster than the wild type for voltages negative to −20 mV but slower for more positive voltages, suggesting a reduced voltage dependence of fast inactivation. Late I Na at 240 ms was significantly greater for ΔKPQ than for the wild type at both temperatures. The majority of late I Na for ΔKPQ was not persistent; rather, it decayed slowly, and this late component exhibited slower recovery from inactivation compared with peak I Na. Additional kinetic changes for early and peak I Na for ΔKPQ compared with the wild type at both temperatures were 1) reduced voltage dependence of steady-state inactivation with no difference in midpoint, 2) positive shift for activation kinetics, and 3) more rapid recovery from inactivation. This study represents the first description of human Na+ channel kinetics near physiological temperature and also demonstrates complex gating changes in the ΔKPQ that are present at 33°C and that may underlie the electrophysiological and clinical phenotype of congenital long Q-T Na+ channel syndromes.
Thermodynamic limitation for Ca2+handling contributes to decreased contractile reserve in rat hearts
Authors: Tian Rong , Halow Jessica M. , Meyer Markus , Dillmann Wolfgang H. , Figueredo Vincent M. , Ingwall Joanne S. , Camacho S. Albert
Published: December 2017

The free energy release from ATP hydrolysis (‖ΔG∼p‖) is decreased by inhibiting the creatine kinase (CK) reaction, which may limit the thermodynamic driving force for the sarcoplasmic reticulum (SR) Ca2+pumps and thereby cause a decrease in contractile reserve. To determine whether a decrease in ‖ΔG∼p‖ results in decreased contractile reserve by impairing Ca2+handling, we measured left ventricular pressure and cytosolic Ca2+concentration ([Ca2+]c; by indo 1 fluorescence) in isolated perfused rat hearts, with >95% inhibition of CK with 90 μmol iodoacetamide. Iodoacetamide did not directly alter SR Ca2+-ATPase activity, baseline left ventricular developed pressure, or baseline [Ca2+]c. When perfusate Ca2+concentration was increased from 1.2 to 3.3 mM, LV developed pressure increased from 67 ± 6 to 119 ± 8 mmHg in control hearts ( P < 0.05) but did not significantly increase in CK-inhibited hearts. Similarly, the amplitude of the [Ca2+]ctransient increased from 548 ± 54 to 852 ± 140 nM in control hearts ( P < 0.05) but did not significantly increase in CK-inhibited hearts. We conclude that decreased ‖ΔG∼p‖ limits intracellular Ca2+handling and thereby limits contractile reserve.
Early and delayed tolerance to simulated ischemia in heat-preconditioned endothelial cells: a role for HSP27
Authors: Loktionova Svetlana A. , Ilyinskaya Olga P. , Kabakov Alexander E.
Published: December 2017

An ischemia-mimicking metabolic stress in cultured endothelial cells from the human aorta or umbilical vein caused ATP depletion, a rise in cytosolic free Ca2+, fragmentation and aggregation of actin microfilaments, retraction of the cytoplasm, and disintegration of cell monolayer. Simultaneously, the constitutive heat shock protein 27 (HSP27) underwent dephosphorylation and formed granules inside cell nuclei. Prior heat shock (45°C, 10 min) in confluent cultures conferred two phases (early and delayed) of tolerance to simulated ischemia. Although heat preconditioning did not retard the ATP drop and the free Ca2+overload within ischemia-stressed cells, each phase of the tolerance was manifested in longer preservation of normal cell morphology during the stress. Cells exhibiting the early tolerance within 3 h after heating altered the F-actin response to ischemic stress; no microfilament debris but, instead, translocation of F-actin to the tight submembranous layer was observed. In contrast, the delayed cytoprotection preserved the preexisting F-actin bundles under simulated ischemia; this happened only after 12- to 14-h post-heat shock recovery, elevating the intracellular HSP content, and was sensitive to blockers of HSP synthesis, cycloheximide and quercetin. The dephosphorylation and intranuclear granulation of HSP27 were markedly suppressed in both phases of the heat-induced tolerance. Without heat pretreatment, similar attenuation of the HSP27 dephosphorylation/granulation and the actin cytoskeleton stability during simulated ischemia were achieved by treating cells with the protein phosphatase inhibitors cantharidin or sodium orthovanadate. We suggest that prior heat shock ameliorates the F-actin response to ischemic stress by suppressing the HSP27 dephosphorylation/granulation; this prolongs a sojourn in the cytosol of phosphorylated HSP27, which protects microfilaments from the disruption and aggregation.
Arterial heparin deposition: role of diffusion, convection, and extravascular space
Authors: Lovich Mark A. , Philbrook Mike , Sawyer Sean , Weselcouch Ed , Edelman Elazer R.
Published: December 2017

Transvascular transport has been studied with atherogenic, tracer, and inert compounds such as low-density lipoprotein, horseradish peroxidase, and albumin, respectively. Few studies used vasoactive compounds, and virtually all studies examined entry from the lumen and not from the perivascular space. We compared several mechanisms that govern arterial heparin deposition after administration to the perivascular and endovascular aspects of the calf carotid artery in vitro and the rabbit iliac artery in vivo. In the absence of transmural hydrostatic pressure gradients, heparin deposition following endovascular administration was unaffected by deendothelialization and was indistinguishable from perivascular delivery. Deposition in the former was enhanced by the addition of a pressure gradient and to a greater extent in denuded arteries, indicating that convection influences transport but is dampened by the endothelium. Neither the endothelium nor the adventitia pose significant resistances to heparin. Deposition in vivo was greater following endovascular hydrogel release than perivascular application from similar devices to native or denuded arteries. The loss of drug to extra-arterial microvessels exceeded the loss of drug to the lumen flow. These findings are essential for describing vascular pharmacokinetics and for implementing local pharmacotherapies.
Myocardial ischemia-reperfusion injury in CD18- and ICAM-1-deficient mice
Authors: Palazzo Anthony J. , Jones Steven P. , Girod Wesley G. , Anderson Donald C. , Granger D. Neil , Lefer David J.
Published: December 2017

Previous studies have demonstrated that circulating neutrophils (PMNs) contribute to the pathophysiology of myocardial ischemia-reperfusion (MI/R) injury. PMN-endothelial cell interactions are highly regulated by adhesive interactions between PMN CD11/CD18 and coronary endothelial cell intercellular adhesion molecule-1 (ICAM-1). We investigated the effects of MI/R in wild-type, CD18-, and ICAM-1-deficient (−/−) mice. Wild-type ( n = 6), CD18 −/− ( n = 6), and ICAM-1 −/− ( n = 6) mice were subjected to 30 min of myocardial ischemia and 120 min of reperfusion to determine the extent of PMN infiltration and myocardial cell necrosis. Myocardial infarction (% of the area at risk) was 45.1 ± 5.9 in wild-type mouse hearts. In contrast, the extent of myocardial infarction was significantly ( P < 0.05) reduced in the CD18 (19.3 ± 5.1%)- and ICAM-1 (17.9 ± 3.2%)-deficient mice. Similarly, PMN infiltration into the ischemic-reperfused myocardium was attenuated by 54% in the CD18 −/− mice and by 32% in ICAM-1 −/− mice compared with wild-type hearts. Deficiency in either CD18 or ICAM-1 expression results in a marked reduction in PMN accumulation and myocardial necrosis after acute MI/R.
Reduced NO enhances the central gain of cardiac sympathetic afferent reflex in dogs with heart failure
Authors: Ma Rong , Zucker Irving H. , Wang Wei
Published: December 2017

The aim of the present study was to test the hypothesis that a decrease in central nitric oxide (NO) is involved in the enhancement of the central gain of the cardiac “sympathetic afferent” reflex (CSAR) in dogs with congestive heart failure (CHF). Thirteen dogs with pacing-induced CHF and sixteen sham dogs were anesthetized with α-chloralose and were baroreceptor denervated and vagotomized. The CSAR was evoked by stimulation of the left ventral ansa. A lateral cerebroventricular cannula was inserted to deliver sodium nitroprusside (SNP) and N G-nitro-l-arginine methyl ester (l-NAME). Arterial pressure, heart rate, and renal sympathetic nerve activity (RSNA) were recorded at baseline and during elicitation of the CSAR. We found that 1) the responses of RSNA to stimulation were augmented in dogs with CHF, 2) SNP depressed the increase in RSNA induced by the CSAR in CHF dogs but had no effect in sham dogs, and 3)l-NAME potentiated the CSAR-induced increase in RSNA in sham dogs but not in dogs with CHF. We conclude that reduced central NO is involved in the enhanced central gain of the CSAR in CHF dogs.
Cardiac electrical restitution properties and stability of reentrant spiral waves: a simulation study
Authors: Qu Zhilin , Weiss James N. , Garfinkel Alan
Published: December 2017

Spiral wave breakup is a proposed mechanism underlying the transition from ventricular tachycardia to fibrillation. We examined the importance of the restitution of action potential duration (APD) and of conduction velocity (CV) to the stability of spiral wave reentry in a two-dimensional sheet of simulated cardiac tissue. The Luo-Rudy ventricular action potential model was modified to eliminate its restitution properties, which are caused by deactivation or recovery from inactivation of K+, Ca2+, and Na+ currents ( I K, I Ca, and I Na, respectively). In this model, we find that 1) restitution of I Ca and I Na are the main determinants of the steepness of APD restitution; 2) for promoting spiral breakup, the range of diastolic intervals over which the APD restitution slope is steep is more important than the maximum steepness; 3) CV restitution promotes spiral wave breakup independently of APD restitution; and 4) “defibrillation” of multiple spiral wave reentry is most effectively achieved by combining an antifibrillatory intervention based on altering restitution with an antitachycardia intervention. These findings suggest a novel paradigm for developing effective antiarrhythmic drugs.
Differential depression of myocardial function and metabolism by lactate and H+
Authors: Samaja Michele , Allibardi Sonia , Milano Giuseppina , Neri Gabriella , Grassi Bruno , Gladden L. Bruce , Hogan Michael C.
Published: December 2017

The effects of both high blood H+ concentration ([H+]) and high blood lactate concentration ([lactate]) under ischemia-reperfusion conditions are receiving attention, but little is known about their effects in nonischemic hearts. Isolated rat hearts were Langendorff perfused at constant flow with media at two pH values (7.4 and 7.0) and two [lactate] (0 and 20 mM) in various sequences ( n = 6/group). Coronary flow and arterial O2content were kept constant at levels that allowed hearts to function without O2 supply limitation. We measured contractility, O2 uptake, diastolic pressure, and at the end of the protocol, tissue [lactate] and pH. Perfusion with high [lactate] raised tissue [lactate] from 5.5 ± 0.1 to 17.5 ± 2.6 μmol/heart ( P < 0.0001), whereas decreasing the pH of the medium decreased tissue pH from 6.94 ± 0.02 to 6.81 ± 0.06 ( P = 0.002). Heart rate was not affected by high [lactate] but was reversibly depressed by high [H+] ( P = 0.004). Developed pressure declined by 20% in response to high [lactate], high [H+], and high [lactate] + high [H+] ( P = 0.002). After the high-[lactate] challenge was withdrawn, pressure continued to decline. In contrast, withdrawing the high [H+] challenge allowed partial recovery. The behavior of diastolic pressure mirrored that of developed pressure. Although unaffected by high [lactate], the O2 uptake was reversibly depressed by high [H+]. This suggests higher O2 cost per contraction in the presence of high [lactate]. We conclude that for similar acute contractility depression, high [lactate] induces irreversible damage, likely at some point in the pathway of O2 utilization. In contrast, the effect of high [H+] appears reversible. These differential behaviors may have implications for heart function during heavy exercise and ischemia-reperfusion events.
P2 purinoceptor-mediated dilations in the rat middle cerebral artery after ischemia-reperfusion
Authors: Marrelli Sean P. , Khorovets Andrei , Johnson T. David , Childres William F. , Bryan Robert M.
Published: December 2017

Endothelial-mediated dilations to selective P2Y1 and P2Y2 purinoceptor agonists [2-methylthioadenosine triphosphate (2MeS-ATP) and uridine 5′-triphosphate (UTP), respectively] were evaluated in middle cerebral arteries (MCAs) of rats after 2 h of ischemia followed by 24 h of reperfusion (I/R). MCAs were harvested, pressurized to 85 mmHg, and luminally perfused. 2MeS-ATP, which dilates by the synthesis and release of nitric oxide (NO), had significantly reduced maximum dilations following I/R. Reduced smooth muscle sensitivity to NO may explain the reduced dilation to 2MeS-ATP. In contrast, the dilations elicited by UTP were potentiated in that the concentration of agonist necessary to produce one-half of the maximum dilation was reduced by 75%. The potentiated dilation to UTP was the result of an endothelial factor having all the characteristics of the endothelium-derived hyperpolarizing factor (EDHF). That is, it was neither NO nor a cyclooxygenase metabolite, and its actions involved calcium-activated potassium channels and smooth muscle hyperpolarization. We conclude that the effect of I/R on endothelial-mediated dilations depends on the receptor system and the mechanism of dilation. Dilations elicited by 2MeS-ATP were attenuated, while dilations UTP were potentiated due to the upregulation of the EDHF mechanism.
Superoxide anion scavengers restore NO-mediated pulmonary vasodilation after lung transplantation
Authors: Seki Sumihiko , Flavahan Nicholas A. , Smedira Nicholas G. , Murray Paul A.
Published: December 2017

Left lung autotransplantation (LLA) results in a chronic attenuation in endothelium-dependent, nitric oxide (NO)-mediated pulmonary vasodilation. We tested the hypothesis that this abnormality involves a decrease in the effective concentration of NO due to inactivation by superoxide anion. Size- and position-matched pulmonary arterial rings were isolated from the right (control) and left (LLA) lungs of seven dogs 1–5 mo post-LLA. The rings were suspended for isometric tension recording and contracted with phenylephrine, and cumulative dose-response curves for ACh or calcium ionophore (A-23187) were generated. Endothelium-dependent relaxation to ACh was inhibited post-LLA, with the maximum vasorelaxation response reduced from 88 ± 5 to 63 ± 5% ( P < 0.01) post-LLA. In contrast, after pretreatment with the superoxide anion scavengers tiron or superoxide dismutase (SOD), the dose-response relationships for ACh were similar in control and LLA rings. Oxypurinol, which inhibits superoxide anion production by endothelial xanthine oxidase, also restored the vasorelaxation response to ACh in LLA rings. The pulmonary vasorelaxant response to A-23187 was also attenuated ( P < 0.01) post-LLA, and this effect was entirely reversed by pretreatment with tiron, SOD, or oxypurinol. These results indicate that the attenuated responses to these pulmonary vasorelaxants post-LLA involve inactivation of NO by superoxide anion generated by endothelial xanthine oxidase.
An adenosine agonist and preconditioning shift the distribution of myocardial blood flow in conscious pigs
Authors: Huang Cheng-Hsiung , Kim Song-Jung , Ghaleh Bijan , Kudej Raymond K. , Shen You-Tang , Bishop Sanford P. , Vatner Stephen F.
Published: December 2017

The goal of this study was to determine whether the cardioprotective effects of an A1-receptor agonist and ischemic preconditioning (IPC) involve a shift in the pre-coronary artery occlusion (CAO) spatial distribution of myocardial blood flow, which might shed light on the mechanism of IPC and explain its heterogeneous effects. Accordingly, 60 min of CAO followed by 72 h of coronary artery reperfusion (CAR) was examined in three groups of conscious pigs 10–14 days after instrumentation with aortic and left atrial catheters and coronary artery occluders. Myocardial infarct size, expressed as a fraction of the area at risk (AAR), was reduced significantly ( P < 0.05) by infusion of the A1 agonist (27.1 ± 6.6%) and to a greater extent ( P < 0.05) by IPC (11.6 ± 5.1%) compared with infarct size in vehicle-treated animals (55.1 ± 2.9%). Transmural myocardial blood flow (radioactive microspheres) in the AAR shifted toward lower levels after infusion of the A1 agonist (1.27 ± 0.19 vs. 0.74 ± 0.10 ml ⋅ min−1 ⋅ g−1) or IPC (1.27 ± 0.11 vs. 0.96 ± 0.09 ml ⋅ min−1 ⋅ g−1) but not after infusion of the vehicle (1.20 ± 0.10 vs. 1.23 ± 0.09 ml ⋅ min−1 ⋅ g−1). This study demonstrated that both pretreatment with an adenosine A1 agonist and also IPC altered the spatial distribution of pre-CAO myocardial blood flow, which might reflect a downregulation of metabolic state and thus play a role in the cardioprotective effects of IPC.
Endothelium-independent vascular relaxation mediating ETBreceptor in rabbit mesenteric arteries
Authors: Iwasaki Takanori , Notoya Mitsuru , Hayasaki-Kajiwara Yoko , Shimamura Toshitake , Naya Noriyuki , Ninomiya Mitsuyoshi , Nakajima Masatoshi
Published: December 2017

Vascular response mediating endothelin (ET)Breceptor was studied using isolated rabbit mesenteric arteries. ET-1 (0.1–30 nM) caused a concentration-dependent contraction, whereas ET-3 >100 nM caused only weak contraction. Up to 1 μM of sarafotoxin S6c showed no contraction. In arteries precontracted with phenylephrine, ET-3 (0.03–1 nM) caused a concentration-dependent relaxation, which was not affected by endothelium denudation. The ET-3-induced relaxation was antagonized by BQ-788 and PD-142893 but not by BQ-123 in the endothelium-denuded arteries. Treatment with indomethacin but not with NG-nitro-l-arginine methyl ester completely inhibited the relaxation. ET-3 stimulated the release of 6-keto-PGF1αand PGE2from the endothelium-denuded arteries. ET-3 also significantly increased cAMP content but not cGMP content in the arteries. Radioligand-binding studies using serial sections of the artery revealed the expression of not only ETAbut also ETBreceptors in the smooth muscle layer of the arteries. These results suggest that ET-3 activates ETBreceptor in smooth muscle cells of rabbit mesenteric artery, producing vasodilator prostaglandins from arachidonic acid probably via a catalysis of cyclooxygenase, which accumulates cAMP in subendothelial tissues and produces relaxations.
Endocardial endothelium mediates luminal ACh-NO signaling in isolated frog heart
Authors: Gattuso Alfonsina , Mazza Rosa , Pellegrino Daniela , Tota Bruno
Published: December 2017

ACh exerted a biphasic effect in the in vitro working heart of Rana esculenta. High concentrations (10−7M) of ACh depressed stroke volume (SV) and stroke work (SW) by ∼30% with a shorter systolic phase and reduced peak pressure. Doses from 10−10M induced a positive response peaking at 10−8M (SV: +8.6%; SW: +6.5%) and a prolonged systolic phase without affecting peak pressure. Atropine and pirenzepine blocked both the positive and the negative effects of ACh. Pretreatment with Triton X-100 (0.1 ml, 0.05%) or with nitric oxide (NO)-cGMP pathway antagonists ( NG-nitro-l-arginine, NG-nitro-l-arginine methyl ester, NG-monomethyl-l-arginine, and 1 H-[1,2,4]oxadiazolo-[4,3- a]quinoxalin-1-one) abolished the positive and negative cholinergic effects. Infusion of 8-bromoguanosine 3′,5′-cyclic monophosphate reverted the positive effect of ACh to a negative effect. Milrinone blocked the positive inotropism but did not change the negative cholinergic response. The NO donor 3-morpholinosydnonimine generated a biphasic dose-response curve with a maximum positive effect at 10−8M (SV: +8%; SW: +5.6%; systolic phase: +28 ms) and a negative effect at 5 × 10−8M (SV and SW: about −12%; systolic phase: −70 ms; peak pressure: −1.50 mm). We conclude that in the avascular frog heart the endocardial endothelium mediates the inotropic effect of luminal cholinergic stimuli via a NO-cGMP pathway.
Modulation of arterial Na+-K+-ATPase-induced [Ca2+]ireduction and relaxation by norepinephrine, ET-1, and PMA
Authors: Pérez-Vizcaíno Francisco , Cogolludo Angel , Tamargo Juan
Published: December 2017

Na+-K+-ATPase plays a major role in regulating membrane potential and vascular tone. We analyzed the modulation by norepinephrine (NE), endothelin-1 (ET-1), and phorbol 12-myristate 13-acetate (PMA) of Na+-K+-ATPase-induced cytoplasmic free Ca2+concentration ([Ca2+]i) reduction and relaxation in isolated endothelium-denuded piglet mesenteric arteries. KCl (0.2–8.8 mM)-induced [Ca2+]ireduction and relaxation in arteries incubated in K+-free solution were used as functional indicators of Na+-K+-ATPase activity. KCl-induced relaxations after exposure to K+-free solution were associated with a reduction in [Ca2+]i, as measured by fura 2 fluorescence. However, KCl reduced [Ca2+]ibelow resting values, whereas force was reduced to near resting values. NE, ET-1, and PMA inhibited the relaxant effects of KCl, and this effect was attenuated by the protein kinase C inhibitor staurosporine but not by the phospholipase A2inhibitor quinacrine. However, ET-1 and PMA potentiated the [Ca2+]i-reducing effect of KCl. In conclusion, ET-1, PMA, and NE are functional inhibitors of Na+-K+-ATPase activity in endothelium-denuded piglet mesenteric arteries, even when the direct effect on the enzyme activity may be stimulatory rather than inhibitory. This can be explained because ET-1, PMA, and NE induce Ca2+sensitization for smooth muscle contraction, and therefore relaxations do not parallel the reductions in [Ca2+]iafter the activation of Na+-K+-ATPase.
Nonstressed rat model of acute endotoxemia that unmasks the endotoxin-induced TNF-α response
Authors: Beno David W. A. , Kimura Robert E.
Published: December 2017

Previous investigators have demonstrated that the tumor necrosis factor-α (TNF-α) response to endotoxin is inhibited by exogenous corticosterone or catecholamines both in vitro and in vivo, whereas others have reported that surgical and nonsurgical stress increase the endogenous concentrations of these stress-induced hormones. We hypothesized that elevated endogenous stress hormones resultant from experimental protocols attenuated the endotoxin-induced TNF-α response. We used a chronically catheterized rat model to demonstrate that the endotoxin-induced TNF-α response is 10- to 50-fold greater in nonstressed (NS) rats compared with either surgical-stressed (SS, laparotomy) or nonsurgical-stressed (NSS, tail vein injection) models. Compared with the NS group, the SS and NSS groups demonstrated significantly lower mean peak TNF-α responses at 2 mg/kg and 6 μg/kg endotoxin [NS 111.8 ± 6.5 ng/ml and 64.3 ± 5.9 ng/ml, respectively, vs. SS 3.9 ± 1.1 ng/ml ( P < 0.01) and 1.3 ± 0.5 ng/ml ( P < 0.01) or NSS 5.2 ± 3.2 ng/ml ( P < 0.01) at 6 μg/kg]. Similarly, baseline concentrations of corticosterone and catecholamines were significantly lower in the NSS group [84.5 ± 16.5 ng/ml and 199.8 ± 26.2 pg/ml, respectively, vs. SS group 257.2 ± 35.7 ng/ml ( P< 0.01) and 467.5 ± 52.2 pg/ml ( P < 0.01) or NS group 168.6 ± 14.4 ng/ml ( P < 0.01) and 1,109.9 ± 140.7 pg/ml ( P < 0.01)]. These findings suggest that the surgical and nonsurgical stress inherent in experimental protocols increases baseline stress hormones, masking the endotoxin-induced TNF-α response. Subsequent studies of endotoxic shock should control for the effects of protocol-induced stress and should measure and report baseline concentrations of corticosterone and catecholamines.
Maintained upregulation of pulmonary eNOS gene and protein expression during recovery from chronic hypoxia
Authors: Resta Thomas C. , Chicoine Louis G. , Omdahl John L. , Walker Benjimen R.
Published: December 2017

We previously demonstrated augmented endothelium-derived nitric oxide (EDNO)-dependent pulmonary arterial dilation and increased arterial endothelial nitric oxide synthase (eNOS) levels in chronic hypoxic (CH) and monocrotaline (nonhypoxic) models of pulmonary arterial hypertension. Therefore, we hypothesized that the long-term elevation of arterial eNOS levels associated with CH is related to pulmonary hypertension or some factor(s) associated with hypertension and not directly to hypoxia. To test this hypothesis, we examined responses to the EDNO-dependent dilator ionomycin in U-46619-constricted, isolated, saline-perfused lungs from control rats, CH (4 wk at 380 mmHg) rats, and rats previously exposed to CH but returned to normoxia for 4 days or 2 wk. Microvascular pressure was assessed by double-occlusion technique, allowing calculation of segmental resistances. In addition, vascular eNOS immunoreactivity was assessed by quantitative immunohistochemistry, and eNOS mRNA abundance was determined by RT-PCR assays. Our findings indicate that 4-day and 2-wk posthypoxic rats exhibit persistent pulmonary hypertension, likely due to maintained arterial remodeling and polycythemia associated with prior exposure to CH. Furthermore, arterial dilation to ionomycin was augmented in lungs from each experimental group compared with controls. Finally, arterial eNOS immunoreactivity and whole lung eNOS mRNA levels remained elevated in posthypoxic animals. These findings suggest that altered vascular mechanical forces or vascular remodeling contributes to enhanced EDNO-dependent arterial dilation and upregulation of arterial eNOS in various models of established pulmonary hypertension.
Central Fos expression in fetal and adult sheep after intraperitoneal hypertonic saline
Authors: Caston-Balderrama A. , Nijland M. J. M. , McDonald T. J. , Ross M. G.
Published: December 2017

We hypothesized that neural structures, involved in sensing extracellular body fluid composition in adult animals during an osmotic challenge, would show similar patterns of activation in fetal sheep. Eight adult sheep [4 hypertonic saline-treated adults (HYP-A), 4 isotonic saline-treated adults] and six near-term fetal sheep [3 hypertonic saline-treated fetuses (HYP-F), 3 isotonic saline-treated fetuses; 130 days gestation] were prepared with vascular and intraperitoneal catheters. Seventy-five minutes before tissue collection, hypertonic (1.5 M) or isotonic saline was infused via an intraperitoneal catheter to adult (18 ml/kg) or fetal sheep (6 ml/kg). Brains were examined for patterns of neuronal activation (demonstrated by Fos protein expression). HYP-A and HYP-F demonstrated similar acute increases in plasma osmolality (∼10 mosmol/kgH2O) and comparable patterns of Fos expression within the organum vasculosum of the lamina terminalis (HYP-A, 67 ± 2 vs. HYP-F, 63 ± 6; means ± SE) and hypothalamic supraoptic (SON; HYP-A, 107 ± 8 vs. HYP-F, 102 ± 7) and paraventricular nuclei (PVN; HYP-A, 71 ± 18 vs. HYP-F, 124 ± 19). Fewer activated neurons were detected in HYP-A vs. HYP-F within the subfornical organ (HYP-A, 33 ± 8 vs. HYP-F, 91 ± 17) and median preoptic nucleus (HYP-A, 33 ± 5 vs. HYP-F, 70 ± 6). In adults and fetuses, counterstaining for arginine vasopressin revealed that neurons within the SON and PVN respond to osmotic challenge. These findings demonstrate that central osmoregulatory centers in adult and near-term fetal sheep are similarly activated by osmotic challenge.
Effects of β2-adrenergic stimulation on single-channel gating of rat cardiac L-type Ca2+channels
Authors: Schröder Frank , Herzig Stefan
Published: December 2017

Cardiac L-type Ca2+channels can be stimulated by activation of β2-adrenoceptors. We intended to determine how the gating behavior at the single-channel level (cell-attached configuration) is affected after selective stimulation of β2-adrenoceptors. Rat cardiomyocytes were exposed to zinterol, a β2-agonist ( n = 7), isoproterenol ( n = 6), a nonselective agonist, 8-bromo-cAMP ( n = 6), and a combination of isoproterenol and ICI-118551 ( n = 8), a selective β2-receptor antagonist, or isoproterenol and CGP-20712A, a β1-selective antagonist ( n = 7). In all groups the ensemble-average current and the availability of the channels to open on depolarization were increased in a similar fashion. In addition, the open probability ( Po) within active sweeps was elevated. However, zinterol exerted this effect in a unique manner. It elevated Ponot by shortening closed times but solely by reducing active sweeps with very low Poand a short burst duration. All zinterol effects were abolished by ICI-118551 ( n = 5) and mimicked by isoproterenol plus CGP-20712A ( n = 7). We conclude that β2-adrenoceptor activation of L-type channels differs qualitatively from the classical cAMP-dependent mechanism.
Effects of AT1 receptor blockade after myocardial infarct on myocardial fibrosis, stiffness, and contractility
Authors: Thai Hoang M. , Van Hohai T. , Gaballa Mohamed A. , Goldman Steven , Raya Thomas E.
Published: December 2017

Angiotensin II type 1 (AT1) receptor blockade attenuates myocardial fibrosis after myocardial infarction (MI). However, whether inhibition of fibrosis by AT1 receptor blockade influences myocardial stiffness and contractility is unknown. We measured left ventricular (LV) hemodynamics, papillary muscle function, and myocardial stiffness and fibrosis in rats randomized to losartan or placebo 1 day after MI and treated subsequently for 8 wk. Losartan decreased LV and right ventricular weights as well as mean aortic and LV systolic pressures in sham and MI rats. LV end-diastolic pressure increased after MI and was decreased with losartan. Maximal developed tension and peak rate of tension rise and decline were decreased in MI vs. sham rats. Interstitial fibrosis developed after MI and was prevented in losartan-treated MI rats. The development of abnormal myocardial stiffness after MI was prevented by losartan. After MI, AT1 receptor blockade prevents an abnormal increase in myocardial collagen content. This effect was associated with a normalization of passive myocardial stiffness.
Betamethasone-mediated vascular dysfunction and changes in hematological profile in the ovine fetus
Authors: Anwar M. A. , Schwab M. , Poston L. , Nathanielsz P. W.
Published: December 2017

Glucocorticoid administration to fetal sheep induces a sustained systemic blood pressure rise and an associated increase in femoral vascular resistance. We utilized a small vessel myograph to compare isometric vascular responses of small femoral arterial branches from fetal sheep infused intravenously with either betamethasone or vehicle in vivo from 128 days gestation. Changes in hematological parameters were also determined. Betamethasone was infused for 48 h to produce fetal plasma betamethasone concentrations similar to those observed in human fetuses after maternal treatment with betamethasone to accelerate fetal lung maturation. When compared with vessels removed from vehicle-infused fetuses, vessels obtained from betamethasone-treated fetuses exhibited 1) enhanced sensitivity to depolarizing potassium solutions; 2) no differences in response to the thromboxane mimetic U-46619 or norepinephrine; and 3) differential responses to vasodilators, enhanced sensitivity to ACh, but decreased response to bradykinin and forskolin. In addition, erythrocyte and leukocyte counts were increased in betamethasone-infused fetuses. These observations indicate that multiple mechanisms operate to increase fetal vascular resistance during antenatal betamethasone exposure.
Distinct scavenger receptor expression and function in the human CD14+/CD16+monocyte subset
Authors: Draude Georg , von Hundelshausen Philipp , Frankenberger Marion , Ziegler-Heitbrock H. W. Löms , Weber Christian
Published: December 2017

The CD14+/CD16+subset of human blood monocytes, which expresses low levels of the lipopolysaccharide receptor CD14 and high levels of the Fc receptor CD16 and exhibits features of mature tissue macrophages, is expanded in certain inflammatory conditions and may be relevant in atherosclerosis. Scavenger receptors (ScR) are important for lipid accumulation into macrophage-derived foam cells in atherogenesis and for the clearance of pathogens. Hence, we compared the function and expression of ScR in CD33lowCD16+and CD33highCD14++monocyte subsets. Double immunofluorescence analysis of isolated monocytes revealed that the CD33lowsubset showed lower specific, ScR-mediated binding of DiI-labeled modified low-density lipoproteins (LDL) than CD33highcells. Differences in modified LDL binding between subsets were accompanied by changes in mRNA expression. RT-PCR in sorted cells indicated lower ScR class A type I/II (ScR-AI/II) mRNA levels in CD14+/CD16+than in CD14++cells, whereas CD36 transcripts were unaltered. This was paralleled by findings in mostly CD16+monocyte-derived macrophages showing a marked reduction in ScR-mediated binding of acetylated LDL, but not in the binding of oxidized LDL, and lower expression of ScR-AI/II mRNA, but not CD36 transcripts, after exposure to tumor necrosis factor-α for 48 h in vitro. Thus the subset of CD14+/CD16+monocytes shows distinct ScR function and expression, possibly reflecting a preactivation by cytokines with a predilection for specific inflammatory or vascular conditions, e.g., atherogenesis.
On the validity of blood flow measurement using colored microspheres
Authors: Hodeige D. , de Pauw M. , Eechaute W. , Weyne J. , Heyndrickx G. R.
Published: December 2017

The aim of this study was 1) to investigate the validity of repeated estimations of blood flow using colored microspheres (CMS) and 2) to develop and validate a method that permits four consecutive estimations in the same animal using nonradiolabeled microspheres (NRMS). Several mixtures of different types of microspheres were injected in dogs, with each mixture containing the radiolabeled microspheres (RMS; labeled with113Sn) with either three CMS, four CMS, or three CMS and one type of fluorescent (crimson labeled) microsphere (FMS). The blood flows estimated with the use of any of the injected microspheres were compared with those measured using the RMS as the “gold standard.” The results were analyzed by 1) regression analysis, 2) variance analysis (ANOVA I), and 3) estimation of the limits of agreement between RMS and NRMS flow rates. The results indicate that simultaneous estimations of blood flow obtained with the use of more than three CMS lack accuracy and reliability. A combination of three types of CMS with crimson-labeled FMS, however, offers the possibility to estimate consecutively four different flow rates in the same animal in an accurate way and with relatively high precision.