bluuebunny/crossref_metadata_2025 huggingface dataset:
Phospholamban: a major determinant of the cardiac force-frequency relationship
Authors: Bluhm Wolfgang F. , Kranias Evangelia G. , Dillmann Wolfgang H. , Meyer Markus
Published: December 2017

The cardiac force-frequency relationship has been known for over a century, yet its mechanisms have eluded thorough understanding. We investigated the hypothesis that phospholamban, a potent regulator of the sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), determines the cardiac force-frequency relationship. Isolated left ventricular papillary muscles from wild-type (WT) and phospholamban knockout (KO) mice were stimulated at 2 to 6 Hz. The force-frequency relationship was positive in WT but negative in KO muscles, i.e., it was inverted by ablation of phospholamban ( P < 0.01, n = 6 mice). From 2 to 6 Hz, relaxation accelerated considerably (by 10 ms) in WT muscles but only minimally (by 2 ms) in KO muscles (WT vs. KO: P < 0.0001, n = 6). To show that the lack of frequency potentiation in KO muscles was not explained by the almost maximal basal contractility, twitch duration was prolonged in six KO muscles with the SERCA inhibitor cyclopiazonic acid to WT values. Relaxation still failed to accelerate with increased frequency. In conclusion, our results clearly identify phospholamban as a major determinant of the cardiac force-frequency relationship.
Inhibition by calcium antagonism of circulating and renal endothelin in experimental congestive heart failure
Authors: Wei Chiming , Burnett John C.
Published: December 2017

Endothelin (ET) is a potent vasoconstrictor and sodium-regulating peptide whose tissue and plasma concentrations are increased in congestive heart failure (CHF). ET may mediate its vasoconstrictor and sodium-regulatory actions secondary to an increase in intracellular calcium. Calcium influx may augment ET synthesis. Although felodipine, a dihydropyridine calcium-channel antagonist, is effective in reducing vascular resistance in generalized vasoconstriction, its actions in CHF on circulating and local tissue ET remain undefined. The current studies were designed to determine the modulating actions of felodipine (oral, 40 mg/day for 7 days; n= 6) in an experimental canine model of CHF produced by chronic thoracic inferior vena caval constriction (TIVCC) compared with normal ( n = 7) and TIVCC-alone ( n = 7) dogs. We hypothesized that felodipine would decrease circulating and renal ET. Plasma ET was significantly increased in TIVCC compared with normal dogs (26 ± 0.5 vs. 12 ± 0.7 pg/ml, P < 0.05) and was markedly decreased by felodipine compared with TIVCC alone (14 ± 3 vs. 26 ± 0.5 pg/ml, P < 0.05). Renal ET immunohistochemical staining demonstrated the presence of ET in normal kidney, which was markedly increased in renal cortex and medulla in TIVCC dogs. Renal cortical and medullary ET staining densities were markedly decreased with felodipine compared with those with TIVCC alone. In the TIVCC + felodipine group, cardiovascular hemodynamics also was markedly improved compared with the TIVCC-alone group [systemic vascular resistance: 27 ± 2 vs. 44 ± 3 resistance units (RU), P < 0.05; pulmonary vascular resistance: 3.3 ± 0.1 vs. 5.7 ± 0.4 RU, P < 0.05; cardiac output: 2.9 ± 0.2 vs. 1.7 ± 0.1 l/min, P < 0.05]. This study demonstrates important modulating inhibitory actions of felodipine on renal and plasma ET in an experimental model of CHF.
Estrogen status affects sensitivity to focal cerebral ischemia in stroke-prone spontaneously hypertensive rats
Authors: Carswell H. V. O. , Dominiczak A. F. , Macrae I. M.
Published: December 2017

Estrogen treatment has been shown to reduce ischemic brain damage. Because endogenous estrogen levels fluctuate markedly during the estrous cycle, we investigated the effect of stage of estrous cycle on ischemic brain damage. Halothane anesthetized 3- to 5-mo-old female Wistar-Kyoto rats (WKY) and stroke-prone spontaneously hypertensive rats (SHRSP) in proestrus (high estradiol levels) or metestrus (low estradiol levels) underwent permanent middle cerebral artery occlusion. In SHRSP, infarct volume at 24 h postocclusion was 24% smaller in proestrus compared with metestrus [208.6 ± 9.5 mm3 ( n = 7) vs. 272.7 ± 23.8 mm3 ( n = 7), respectively, means ± SE; P = 0.0278, unpaired t-test]. In WKY, infarct volumes were similar in proestrus and metestrus [157.0 ± 5.4 mm3 ( n= 5) and 131.5 ± 16.5 mm3 ( n = 8), respectively; P = not significant (NS)]. Brain swelling (ipsilateral minus contralateral hemispheric volumes) was similar in proestrus and metestrus for SHRSP [138 ± 9 mm3 ( n = 6) and 136 ± 10 mm3 ( n = 7), respectively] and for WKY [103 ± 15 mm3 ( n = 5) and 90 ± 11 mm3 ( n = 8), respectively]. Thus the reduction in infarct size in SHRSP is caused by a true attenuation of the infarct volume and not simply by a reduction in brain edema.
Na+/H+ exchange inhibition attenuates hypertrophy and heart failure in 1-wk postinfarction rat myocardium
Authors: Yoshida Hiroyuki , Karmazyn Morris
Published: December 2017

Na+/H+ exchange (NHE) represents a major mechanism for intracellular pH regulation, particularly in the ischemic myocardium. NHE has also been shown to be important in the regulation of cell proliferation and growth. We examined whether inhibition of NHE results in an attenuation of early postinfarction myocyte remodeling responses in the rat. Male Sprague-Dawley rats were randomized to receive either a control diet or an identical diet supplemented with the NHE inhibitor cariporide. After 1 wk, animals were anesthetized, subjected to ligation of the left main coronary artery, and maintained for an additional week, after which time they were anesthetized and intraventricular pressures were obtained. Hearts were removed, and myocytes were isolated to obtain cell dimensions and determine the response to isoproterenol. Body, heart, and lung weights were obtained. Coronary artery ligation in control animals resulted in a significant elevation in left ventricular end-diastolic pressure, as well as increased heart weight- and lung weight-to-body weight ratios, both of which were abrogated by cariporide. Cell length and area significantly increased by 14 and 19.2%, respectively, whereas cell width increased by 4.1% ( P> 0.05). These cells exhibited a significant hyporesponsiveness to the positive inotropic responses to isoproterenol at the lower drug concentrations (3 and 10 nM). A <1% dimensional change occurred in myocytes from cariporide-fed animals, and the hyporesponse to isoproterenol was reversed. Cariporide had no effect on infarct size or blood pressure. These studies suggest that the early adaptive hypertrophic response of surviving myocytes is dependent on NHE activity. As such, it is attractive to suggest that NHE inhibition could be an effective therapeutic strategy for prevention of postinfarction remodeling, independent of infarct size or afterload reduction.
Role of adenosine A2B receptors in vasodilation of rat pial artery and cerebral blood flow autoregulation
Authors: Shin Hwa Kyoung , Shin Yung Woo , Hong Ki Whan
Published: December 2017

This study was aimed to investigate the underlying mechanism of vasodilation induced by the activation of A2B adenosine receptors in relation to cerebral blood flow (CBF) autoregulation. Changes in pial arterial diameters were observed directly through a closed cranial window. N ω-nitro-l-arginine methyl ester (l-NAME, nitric oxide synthase inhibitor) significantly suppressed the concentration-dependent vasodilations induced by adenosine and 5′- N-ethylcarboxamido-adenosine (NECA) but not the vasodilation by CGS-21680 (A2A-receptor agonist). Moreover, NECA-induced vasodilation was suppressed by alloxazine (1 μmol/l) but not by ZM-241385 (1 μmol/l, A2Aantagonist), which suggests mediation by A2B- receptor activation. Otherwise, the level of nitrite/nitrate was concentration dependently increased in the artificial cerebrospinal fluid (CSF) when adenosine and NECA were suffused over the cortical surface.l-NAME and alloxazine, but not ZM-241385, largely inhibited their releases. The lower limit of CBF autoregulation was little affected following pretreatment with l-NAME or alloxazine. Thus it is suggested that adenosine-induced vasodilation via activation of A2B-adenosine receptors of the rat pial artery is coupled to the production of nitric oxide, which contributes little to CBF autoregulation.
Transient increases in diameter and [Ca2+]i are not obligatory for myogenic constriction
Authors: Hill Michael A. , Zou Hui , Davis Michael J. , Potocnik Simon J. , Price Simone
Published: December 2017

Studies were performed to determine the significance of temporal variation in vascular smooth muscle Ca2+ signaling during acute arteriolar myogenic constriction and, in particular, the importance of the stretch-induced intracellular Ca2+concentration ([Ca2+]i) transient in attaining a steady-state mechanical response. Rat cremaster arterioles (diameter ∼100 μm) were dissected from surrounding tissues, and vessel segments were pressurized in the absence of intraluminal flow. For [Ca2+]imeasurements, vessels were loaded with fura 2 and fluorescence emitted by excitation at 340 and 380 nm was measured using video-based image analysis. Ca2+ and diameter responses were examined after increases in intravascular pressure were applied as an acute step increase or a ramp function. Additional studies examined the effect of longitudinal vessel stretch on [Ca2+]i and arteriolar diameter. Step increase in intraluminal pressure (from 50 to 120 mmHg) caused biphasic change in [Ca2+]i and diameter. [Ca2+]i transiently increased to 114.0 ± 2.0% of basal levels and subsequently declined to 106.7 ± 4.4% at steady state. Diameter initially distended to 125.4 ± 2.1% of basal levels before constricting to 71.1 ± 1.2%. In contrast, when the same pressure increase was applied as a ramp function (over 5 min) transient vessel distension and transient increase in [Ca2+]i were prevented, yet at steady state vessels constricted to 71.3 ± 2.5%. Longitudinal stretch resulted in a large [Ca2+]i transient (158 ± 19% of basal) that returned to baseline despite maintenance of the stretch stimulus. The data demonstrate that the initial vessel distension (reflecting myocyte stretch) and associated global [Ca2+]i transient are not obligatory for myogenic contraction. Thus, although arteriolar smooth muscle cells are responsive to acute stretch, the resulting changes in myogenic tone may be more closely related to other mechanical variables such as wall tension.
B1and B2bradykinin receptors on adventitial fibroblasts of cerebral arteries are coupled to recombinant eNOS
Authors: Tsutsui Masato , Onoue Hisashi , Iida Yasuhiko , Smith Leslie , O'Brien Timothy , Katusic Zvonimir S.
Published: December 2017

Our previous ex vivo and in vivo studies reported that expression of the recombinant endothelial nitric oxide (NO) synthase (eNOS) gene in adventitial fibroblasts recovers NO production in arteries without endothelium in response to bradykinin. The present study was designed to characterize subtypes of bradykinin receptors on adventitial fibroblasts coupled to the activation of recombinant eNOS. Endothelium-denuded segments of canine basilar arteries were transduced with β-galactosidase (β-Gal) gene or eNOS gene ex vivo, using a replication-defective adenoviral vector (1010plaque-forming units/ml) for 30 min at 37°C. Twenty-four hours later, isometric force recording or cGMP measurement was carried out. B1bradykinin receptor agonist (des-Arg9-bradykinin, 10−10-10−8mol/l) did not significantly affect vascular tone in control or β-Gal gene-transduced canine basilar arteries without endothelium. In contrast, this agonist caused concentration-dependent relaxations in recombinant eNOS gene-transduced arteries without endothelium. Relaxations to B1receptor agonist in the eNOS arteries were abolished by B1receptor antagonist (des-Arg9-[Leu8]bradykinin, 6 × 10−9mol/l) but not by B2receptor antagonist (Hoe-140, 5 × 10−8mol/l). Bradykinin did not significantly alter vascular tone in control or β-gal arteries without endothelium, whereas this peptide (10−11-10−8mol/l) induced concentration-dependent relaxations, as well as an increase in cGMP formation in endothelium-denuded eNOS-transduced arteries. Stimulatory effects of bradykinin were prevented in the presence of a B2receptor antagonist but not in the presence of a B1receptor antagonist. B1and B2receptor antagonists had no effect on relaxations to substance P, confirming the selectivity of the compounds. Our results suggest that B1and B2bradykinin receptors are coupled to activation of recombinant eNOS expressed in adventitial fibroblasts.
Reduced NO-dependent arteriolar dilation during the development of cardiomyopathy
Authors: Sun Dong , Huang An , Zhao Gong , Bernstein Robert , Forfia Paul , Xu Xiaobin , Koller Akos , Kaley Gabor , Hintze Thomas H.
Published: December 2017

Our previous studies have suggested that there is reduced nitric oxide (NO) production in canine coronary blood vessels after the development of pacing-induced heart failure. The goal of these studies was to determine whether flow-induced NO-mediated dilation is altered in coronary arterioles during the development of heart failure. Subepicardial coronary arterioles (basal diameter 80 μm) were isolated from normal canine hearts, from hearts with dysfunction but no heart failure, and from hearts with severe cardiac decompensation. Arterioles were perfused at increasing flow or administered agonists with no flow in vitro. In arterioles from normal hearts, flow increased arteriolar diameter, with one-half of the response being NO dependent and one-half prostaglandin dependent. Shear stress-induced dilation was eliminated by removing the endothelium. Arterioles from normal hearts and hearts with dysfunction but no failure responded to increasing shear stress with dilation that reached a maximum at a shear stress of 20 dyn/cm2. In contrast, arterioles from failing hearts showed a reduced dilation, reaching only 55% of the dilation seen in vessels of normal hearts at a shear stress of 100 dyn/cm2. This remaining dilation was eliminated by indomethacin, suggesting that the NO-dependent component was absent in coronary microvessels after the development of heart failure. Similarly, agonist-induced NO-dependent coronary arteriolar dilation was markedly attenuated after the development of heart failure. After the development of severe dilated cardiomyopathy and heart failure, the NO-dependent component of both shear stress- and agonist-induced arteriolar dilation is reduced or entirely absent.
Effects of central infusion of ANG II and losartan on the cardiac baroreflex in rabbits
Authors: Gaudet Elisabeth , Godwin Shirley J. , Head Geoffrey A.
Published: December 2017

The effect of chronic activation or inhibition of central ANG II receptors on cardiac baroreflex function in conscious normotensive rabbits was examined. Animals received a fourth ventricular (4V) infusion of ANG II (30 and 100 ng/h), losartan (3 and 30 μg/h), or Ringer solution (2 μl/h) for 2 wk. After 1 and 2 wk, ANG II (100 ng/h) decreased cardiac baroreflex gain by 20 and 37%, respectively ( P = 0.015), whereas losartan (30 μg/h) increased baroreflex gain by 24 and 58%, respectively ( P = 0.02). Within 1 wk of the end of the infusions, cardiac baroreflex gain had returned to control. Ringer solution or the lower doses of ANG II or losartan did not modify the cardiac baroreflex function. Blood pressure and heart rate were not altered by any treatment, nor was their variability affected. These data demonstrate a novel long-term modulation of cardiac baroreflexes by endogenous ANG II that is independent of blood pressure level.
Developmental changes of cardiac function and mass assessed with MRI in neonatal, juvenile, and adult mice
Authors: Wiesmann Frank , Ruff Jan , Hiller Karl-Heinz , Rommel Eberhard , Haase Axel , Neubauer Stefan
Published: December 2017

Cardiovascular transgenic mouse models with an early phenotype or even premature death require noninvasive imaging methods that allow for accurate visualization of cardiac morphology and function. Thus the purpose of our study was to assess the feasibility of magnetic resonance imaging (MRI) to characterize cardiac function and mass in newborn, juvenile, and adult mice. Forty-five C57bl/6 mice from seven age groups (3 days to 4 mo after birth) were studied by MRI under isoflurane anesthesia. Electrocardiogram-gated cine MRI was performed with an in-plane resolution of (78–117 μm)2. Temporal resolution per cine frame was 8.6 ms. MRI revealed cardiac anatomy in mice from all age groups with high temporal and spatial resolution. There was close correlation between MRI- and autopsy-determined left ventricular (LV) mass ( r = 0.95, SE of estimate = 9.5 mg). The increase of LV mass (range 9.6–101.3 mg), cardiac output (range 1.1–14.3 ml/min), and stroke volume (range 3.2–40.2 μl) with age could be quantified by MRI measurements. Ejection fraction and cardiac index did not change with aging. However, LV mass index decreased with increasing age ( P < 0.01). High-resolution MRI allows for accurate in vivo assessment of cardiac function in neonatal, juvenile, and adult mice. This method should be useful when applied in transgenic mouse models.
LPS tolerance in human endothelial cells: reduced PMN adhesion, E-selectin expression, and NF-κB mobilization
Authors: Lush Cameron W. , Cepinskas Gediminas , Kvietys Peter R.
Published: December 2017

Cytokine release from inflammatory (CD14+) cells is reduced after repeated stimulation with lipopolysaccharide (LPS; LPS tolerance). However, it is not known whether LPS tolerance can be induced in CD14−cells. The aim of the present study was to determine whether endothelial cells [human umbilical vein endothelial cells (HUVEC)] could be rendered tolerant to LPS with respect to LPS-induced polymorphonuclear neutrophil (PMN) adhesion. LPS stimulation (0.5 μg/ml; 4 h) of naive HUVEC increased PMN adhesion. Pretreatment of HUVEC with LPS (0.5 μg/ml) for 24 h resulted in a reduction in the proadhesive effects of a subsequent LPS challenge. The initial LPS stimulation increased 1) mobilization of the nuclear transcription factor NF-κB to the nucleus and 2) surface levels of the adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and E-selectin. In LPS-tolerant HUVEC, a second LPS challenge resulted in 1) less accumulation of NF-κB in the nucleus, 2) a reduction in E-selectin expression, and 3) unchanged ICAM-1 expression. LPS-tolerant cells were still capable of mobilizing NF-κB in response to stimulation with either interleukin-1β or tumor necrosis factor-α, resulting in elevated E-selectin levels and increased PMN adhesion. These studies show for the first time that LPS tolerance can be induced in endothelial cells with respect to PMN adhesion. This tolerance is specific for LPS and is associated with an inability of LPS to mobilize NF-κB, resulting in less E-selectin expression.
Cardiorespiratory interactions during periodic breathing in awake chronic heart failure patients
Authors: Pinna G. D. , Maestri R. , Mortara A. , Rovere M. T. La
Published: December 2017

We applied spectral techniques to the analysis of cardiorespiratory signals [instantaneous lung volume (ILV), instantaneous tidal volume (ITV), arterial O2 saturation ([Formula: see text]) at the ear, heart rate (HR), systolic (SAP), and diastolic (DAP) arterial pressure] during nonapneic periodic breathing (PB) in 29 awake chronic heart failure (CHF) patients and estimated the timing relationships between respiratory and slow cardiovascular (<0.04 Hz) oscillations. Our aim was 1) to elucidate major mechanisms involved in cardiorespiratory interactions during PB and 2) to test the hypothesis of a central vasomotor origin of PB. All cardiovascular signals were characterized by a dominant (≥84% of total power) oscillation at the frequency of PB (mean ± SE: 0.022 ± 0.0008 Hz), highly coherent (≥0.89), and delayed with respect to ITV (ITV-HR, 2.4 ± 0.72 s; ITV-SAP, 6.7 ± 0.65 s; ITV-DAP, 3.2 ± 0.61 s; P < 0.01). [Formula: see text] was highly coherent with (coherence function = 0.96 ± 0.009) and almost opposite in phase to ITV. These findings demonstrate the existence of a generalized cardiorespiratory rhythm led by the ventilatory oscillation and suggest that 1) the cyclic increase in inspiratory drive and cardiopulmonary reflexes and 2) mechanical effects of PB-induced changes in intrathoracic pressure are the more likely sources of the HR and blood pressure oscillations, respectively. The timing relationship between ITV and blood pressure signals excludes the possibility that PB represents the effect of a central vasomotor rhythm.
Protective effect of lung inflation in reperfusion-induced lung microvascular injury
Authors: Srinivasan Hari B. , Vogel Stephen M. , Vidyasagar Dharmapuri , Malik Asrar B.
Published: December 2017

We used the isolated-perfused rat lung model to study the influence of pulmonary ventilation and surfactant instillation on the development of postreperfusion lung microvascular injury. We hypothesized that the state of lung inflation during ischemia contributes to the development of the injury during reperfusion. Pulmonary microvascular injury was assessed by continuously monitoring the wet lung weight and measuring the vessel wall125I-labeled albumin (125I-albumin) permeability-surface area product ( PS). Sprague-Dawley rats ( n = 24) were divided into one control group and five experimental groups ( n = 4 rats per group). Control lungs were continuously ventilated with 20% O2and perfused for 120 min. All lung preparations were ventilated with 20% O2before the ischemia period and during the reperfusion period. The various groups differed only in the ventilatory gas mixtures used during the flow cessation: group I, ventilated with 20% O2; group II, ventilated with 100% N2; group III, lungs remained collapsed and unventilated; group IV, same as group IIIbut pretreated with surfactant (4 ml/kg) instilled into the airway; and group V, same as group III but saline (4 ml/kg) was instilled into the airway. Control lungs remained isogravimetric with baseline125I-albumin PS value of 4.9 ± 0.3 × 10−3ml ⋅ min−1⋅ g wet lung wt−1. Lung wet weight in group III increased by 1.45 ± 0.35 g and albumin PSincreased to 17.7 ± 2.3 × 10−3, indicating development of vascular injury during the reperfusion period. Lung wet weight and albumin PS did not increase in groups I and II, indicating that ventilation by either 20% O2or 100% N2prevented vascular injury. Pretreatment of collapsed lungs with surfactant before cessation of flow also prevented the vascular injury, whereas pretreatment with saline vehicle had no effect. These results indicate that the state of lung inflation during ischemia (irrespective of gas mixture used) and supplementation of surfactant prevent reperfusion-induced lung microvascular injury.
TGF-β1 downregulates CD36 and scavenger receptor A but upregulates LOX-1 in human macrophages
Authors: Draude Georg , Lorenz Reinhard L.
Published: December 2017

Transforming growth factor-β1 (TGF-β1), a key cytokine for control of cell growth, extracellular matrix formation, and inflammation control, is secreted by many cells present in the arteriosclerotic plaque. Lipid accumulation in the vessel wall is regarded as an early step in atherogenesis and depends on uptake of modified low-density lipoprotein (LDL) by macrophages through scavenger receptors and their transformation into foam cells. Prominent members of the scavenger receptor family are the class A type I and II receptors (ScR-A), the class B receptor CD36, and the recently detected lectin-like oxidized LDL receptor-1 (LOX-1), which, unlike the native LDL receptor (LDL-R), are not feedback controlled. CD36 is responsible for >50% of modified LDL uptake into human monocyte-derived macrophages. We therefore studied whether TGF-β1 influences expression and function of ScR-A, CD36, and LOX-1 in monocytes using RT-PCR and flow cytometry. Total uptake of oxidized LDL by monocytoid cells, reflecting the combined function of all scavenger receptors, was significantly reduced by TGF-β1. At initially low picomolar concentrations, TGF-β1 decreased CD36 mRNA and protein surface expression and ScR-A mRNA levels in the human monocytic cell line THP-1 and in freshly isolated and cultivated human monocytes, whereas LOX-1 mRNA was increased. Expression of LDL-R and β-actin was not affected by TGF-β1. In conclusion, depression of scavenger receptor function in monocytes by TGF-β1 in low concentrations reduces foam cell formation. Together with matrix control by TGF-β1, this may be important for atherogenesis and plaque stabilization.
Activation of HSF and selective increase in heat-shock proteins by acute dexamethasone treatment
Authors: Sun L. , Chang J. , Kirchhoff S. R. , Knowlton A. A.
Published: December 2017

Heat-shock proteins (HSPs) are an important family of endogenous protective proteins, which increase in response to myocardial ischemia and other stresses. Overexpression of HSP72 is cardioprotective. We were interested in the regulation of heat-shock factor (HSF), the transcription factor for HSP genes. Previously we have observed that the inflammatory cytokine tumor necrosis factor-α increases HSP72 levels and postulated that dexamethasone might effect the heat shock response. In the adult rat cardiac myocyte we found that treatment with either low (10 μM)- or high (100 μM)-dose dexamethasone activated HSF by 2–6 h as determined by gel shift assay without evidence of cytotoxicity. Although HSF activation is a key step in expression of HSP72, this may not result in an increase in HSP72. We found that 10 μM dexamethasone increased HSP72 38%, and 100 μM dexamethasone increased HSP72 62% ( P < 0.05). HSP27 and HSP60 were unchanged. The selective increase in HSP72 was associated with protection of the cardiac myocytes from hypoxia and reoxygenation. We conclude that dexamethasone is a novel inducer of the heat shock response.
Regulation of DHP receptor expression by elements in the 5′-flanking sequence
Authors: Liu Lei , Fan Q. Ivy , El-Zaru Mohamad R. , Vanderpool Kathleen , Hines Ronald N. , Marsh James D.
Published: December 2017

The α1-subunit of the cardiac/vascular Ca2+channel, which is the dihydropyridine (DHP)-binding site (the DHP receptor), provides the pore structure for Ca2+entry. It contains the binding sites for multiple classes of drugs collectively known as Ca2+antagonists. As an initial step toward understanding the mechanisms controlling transcription of the rat cardiac α1C-subunit gene, we have cloned a 2.3-kb fragment containing the 5′-flanking sequences and identified the α1C-subunit gene transcription start site. The rat α1C-subunit gene promoter belongs to the TATA-less class of such basal elements. Using deletion analysis of α1C-subunit promoter-luciferase reporter gene constructs, we have characterized the transcriptional modulating activity of the 5′-flanking region and conducted transient transfections in cultured neonatal rat cardiac ventricular myocytes and vascular smooth muscle cells. Sequence scanning identified several potential regulatory elements, including five consensus sequences for the cardiac-specific transcription factor Nkx2.5, an AP-1 site, a cAMP response element, and a hormone response element. Transient transfection experiments with the promoter-luciferase reporter fusion gene demonstrate that the 2-kb 5′-flanking region confers tissue specificity and hormone responsiveness to expression of the Ca2+channel α1C-subunit gene. Electrophoretic mobility shift assays identified a region of the α1C-subunit gene promoter that can bind transcription factors and appears to be important for gene expression.
Increased flow precedes remote arteriolar dilations for some microapplied agonists
Authors: Frame Mary D.
Published: December 2017

This study asks which occurs first in time for remote responses: a dilation or a remote change in flow. Arteriolar diameter (∼20 μm) and fluorescently labeled red blood cell (RBC) velocity were measured in the cremaster muscle of anesthetized (pentobarbital sodium, 70 mg/kg) hamsters ( n = 51). Arterioles were locally stimulated for 60 s with micropipette-applied 10 μg/ml LM-609 (αvβ3-integrin agonist), 10−3M adenosine, or 10−3M 3-morpholinosydnonimine (SIN-1, nitric oxide donor) as remote response agonists or with 10−3M papaverine, which dilates only locally. Observations were made at a remote site 1,200 μm upstream. With LM-609 or adenosine, the RBC velocity increased first (within 5 s), and the remote dilation followed 5–7 s later. N-nitro-l-arginine (100 μM) blocked the LM-609 (100%) and adenosine (60%) remote dilations. SIN-1 induced a concurrent remote dilation and decrease in RBC velocity (∼10 s), suggesting the primary signal was to dilate. Papaverine had no remote effects. This study suggests that, although remote responses to some agonists are induced by primary signals to dilate, additionally, network changes in flow can stimulate extensive remote changes in diameter.
Contribution of malonyl-CoA decarboxylase to the high fatty acid oxidation rates seen in the diabetic heart
Authors: Sakamoto Jun , Barr Rick L. , Kavanagh Katherine M. , Lopaschuk Gary D.
Published: December 2017

Myocardial glucose oxidation is markedly reduced in the uncontrolled diabetic. We determined whether this was due to direct biochemical changes in the heart or whether this was due to altered circulating levels of insulin and substrates that can be seen in the diabetic. Isolated working hearts from control or diabetic rats (streptozotocin, 55 mg/kg iv administered 6 wk before study) were aerobically perfused with either 5 mM [14C]glucose and 0.4 mM [3H]palmitate (low-fat/low-glucose buffer) or 20 mM [14C]glucose and 1.2 mM [3H]palmitate (high-fat/high-glucose buffer) ±100 μU/ml insulin. The presence of insulin increased glucose oxidation in control hearts perfused with low-fat/low-glucose buffer from 553 ± 85 to 1,150 ± 147 nmol ⋅ g dry wt−1⋅ min−1( P < 0.05). If control hearts were perfused with high-fat/high-glucose buffer, palmitate oxidation was significantly increased by 112% ( P < 0.05), but glucose oxidation decreased to 55% of values seen in the low-fat/low-glucose group ( P < 0.05). In diabetic hearts, glucose oxidation was very low in hearts perfused with low-fat/low-glucose buffer (9 ± 1 nmol ⋅ g dry wt−1⋅ min−1) and was not altered by insulin or high-fat/high-glucose buffer. These results suggest that neither circulating levels of substrates nor insulin was responsible for the reduced glucose oxidation in diabetic hearts. To determine if subcellular changes in the control of fatty acid oxidation contribute to these changes, we measured the activity of three enzymes involved in the control of fatty acid oxidation; AMP-activated protein kinase (AMPK), acetyl-CoA carboxylase (ACC), and malonyl-CoA decarboxylase (MCD). Although AMPK and ACC activity in control and diabetic hearts was not different, MCD activity and expression in all diabetic rat heart perfusion groups were significantly higher than that seen in corresponding control hearts. These results suggest that an increased MCD activity contributes to the high fatty acid oxidation rates and reduced glucose oxidation rates seen in diabetic rat hearts.
Regulation of ET: pulmonary release of ET contributes to increased plasma ET levels and vasoconstriction in CHF
Authors: Kjekshus Harald , Smiseth Otto A. , Klinge Randi , Øie Erik , Hystad Marit E. , Attramadal Håvard
Published: December 2017

Endothelin (ET) contributes to the increased systemic vascular resistance and elevated cardiac filling pressures seen in congestive heart failure (CHF). We investigated to what extent ET-mediated vasoconstriction in CHF occurs through an endocrine action of elevated plasma ET or by an autocrine/paracrine mechanism related to induction of vascular ET gene expression. Three weeks of pacing (225 beats/min) induced a marked release of ET-1 from the pulmonary circulation with a sixfold elevation of arterial plasma ET in CHF pigs compared with sham-operated pigs. Arterial plasma ET was the strongest and only independent predictor of systemic vascular resistance. In contrast, vascular preproET-1 and ET-receptor mRNA expression were unaltered or decreased in CHF pigs and did not correlate with indexes of vascular tone. However, myocardial preproET-1 mRNA expression increased twofold in CHF pigs. PreproET-2 and preproET-3 mRNAs were not detectable in cardiovascular tissues. In conclusion, plasma ET was markedly increased because of an augmented release from the pulmonary circulation during CHF, and arterial plasma ET correlated with systemic vascular resistance. The absence of ET induction in the peripheral vasculature suggests that ET increases vascular tone during CHF by an endocrine, not an autocrine/paracrine, mechanism.
In vivo expression profile of an endothelial nitric oxide synthase promoter-reporter transgene
Authors: Teichert Anouk-Martine , Miller Tricia L. , Tai Sharon C. , Wang Yang , Bei Xie , Robb G. Brett , Phillips M. James , Marsden Philip A.
Published: December 2017

Endothelium-derived nitric oxide (NO) is primarily attributable to constitutive expression of the endothelial nitric oxide synthase (eNOS) gene. Although a more comprehensive understanding of transcriptional regulation of eNOS is emerging with respect to in vitro regulatory pathways, their relevance in vivo warrants assessment. In this regard, promoter-reporter insertional transgenic murine lines were created containing 5,200 bp of the native murine eNOS promoter directing transcription of nuclear-localized β-galactosidase. Examination of β-galactosidase expression in heart, lung, kidney, liver, spleen, and brain of adult mice demonstrated robust signal in large and medium-sized blood vessels. Small arterioles, capillaries, and venules of the microvasculature were notably negative, with the exception of the vasa recta of the medullary circulation of the kidney, which was strongly positive. Only in the brain was the reporter expressed in non-endothelial cell types, such as the CA1 region of the hippocampus. Epithelial cells of the bronchi, bronchioles, and alveoli were scored as negative, as was renal tubular epithelium. Cardiac myocytes, skeletal muscle, and smooth muscle of both vascular and nonvascular sources failed to demonstrate β-galactosidase staining. Expression was uniform across multiple founders and was not significantly affected by genomic integration site. These transgenic eNOS promoter-reporter lines will be a valuable resource for ongoing studies addressing the regulated expression of eNOS in vivo in both health and disease.
Sphingomyelinase and ceramide analogs induce contraction and rises in [Ca2+]iin canine cerebral vascular muscle
Authors: Zheng Tao , Li Wenyan , Wang Jun , Altura Bella T. , Altura Burton M.
Published: December 2017

Studies were designed to investigate effects of neutral sphingomyelinase ( N-SMase) and ceramide analogs as well as phosphorylcholine on vascular tone and Ca2+mobilization in isolated canine cerebral arterial smooth muscle. N-SMase (0.001–0.1 U/ml) provoked a gradual but sustained vasoconstriction of arterial rings in a concentration-related manner that was endothelium independent. Incubation of denuded arterial rings in Ca2+-free medium or pretreatment with verapamil in extracellular Ca2+resulted in a reduction of the N-SMase-evoked constriction. Exposure of arterial rings to 1,2-bis(2-aminophenoxy)ethane- N, N, N′, N′-tetraacetic acid (BAPTA)-AM did not, however, result in a reduction of N-SMase-induced constriction. Both staurosporine and bisindolymaleimide I attenuated N-SMase-induced contractions to 66% and 72% of control, respectively. N-SMase caused gradual and sustained rises in intracellular Ca2+concentration ([Ca2+]i) in primary cultured cerebral vascular smooth muscle cells. Pretreatment of these cultured cells with nimodipine and verapamil caused a steady decline in N-SMase-induced rises in [Ca2+]i. Exposure of the cells to Ca2+-free solution reversed the [Ca2+]i-induced rise triggered by N-SMase to the resting baseline. Both C8and C16ceramide (10−9–10−6M), but not phosphorylcholine, constricted denuded canine arterial rings in a concentration-related manner and elevated [Ca2+]i. Our results suggest that the sphingomyelin-signaling pathway, via a probable release of ceramide molecules, may play an important role in regulation of cerebral arterial wall tone.
Recombinant human, active site-blocked factor VIIa reduces infarct size and no-reflow phenomenon in rabbits
Authors: Golino Paolo , Ragni Massimo , Cirillo Plinio , Scognamiglio Annalisa , Ravera Amelia , Buono Chiara , Guarino Angela , Piro Orlando , Lambiase Catello , Botticella Filomena , Ezban Mirella , Condorelli Mario , Chiariello Massimo
Published: December 2017

Oxygen free radicals induce de novo synthesis of tissue factor (TF), the initiator of the extrinsic pathway of coagulation, within the coronary vasculature during postischemic reperfusion. In the present study we wanted to assess whether TF expression might cause myocardial injury during postischemic reperfusion. Anesthetized rabbits underwent 30 min of coronary occlusion followed by 5.5 h of reperfusion. At reperfusion the animals received 1) saline ( n = 8), 2) human recombinant, active site-blocked activated factor VII (FVIIai, 1 mg/kg, n = 8), or 3) human recombinant activated FVII (FVIIa, 1 mg/kg, n = 8). FVIIai binds to TF as native FVII, but with the active site blocked it inhibits TF procoagulant activity. The area at risk of infarction (AR), the infarct size (IS), and the no-reflow area (NR) were determined at the end of the experiment. FVIIai resulted in a significant reduction in IS and NR with respect to control animals (28.1 ± 11.3 and 11.1 ± 6.1% of AR vs. 59.8 ± 12.8 and 24.4 ± 2.7% of AR, respectively, P < 0.01), whereas FVIIa resulted in a significant increase in IS and NR to 80.1 ± 13.1 and 61.9 ± 13.8% of AR, respectively ( P < 0.01). In conclusion, TF-mediated activation of the extrinsic coagulation pathway makes an important contribution to myocardial injury during postischemic reperfusion.
Bradykinin mediates cardiac preconditioning at a distance
Authors: Schoemaker Regien G. , van Heijningen Caroline L.
Published: December 2017

Preconditioning the heart by brief coronary (CAO) or mesenteric artery occlusion (MAO) can protect against damage during subsequent prolonged CAO and reperfusion. The role of bradykinin (BK) in remote cardiac preconditioning by MAO is investigated by antagonizing the BK B2 receptor [Hoechst 140 (HOE-140)] or simulating local BK release by mesenteric intra-arterial infusion. Anesthetized male Wistar rats ( n = 6–8) were treated with HOE-140 or saline before starting the preconditioning protocol, CAO, MAO, or non-preconditioned control. Infarct size related to risk area [ratio of infarct area to area at risk (IA/AR)] was determined after 3 h of reperfusion following a 60-min CAO. IA/AR was 62 ± 5% in controls and not affected by HOE-140 (58 ± 6%). CAO as well as MAO significantly protected the heart (IA/AR, 37 ± 3 and 35 ± 5%), which was prevented by HOE-140 (IA/AR, 71 ± 6 and 65 ± 7%, respectively). Brief intramesenteric BK infusion mimicked MAO (IA/AR, 26 ± 3%). Pretreatment with hexamethonium could abolish this protection (IA/AR, 67 ± 4%). These data indicate an important role for BK in remote preconditioning by MAO. Results support the hypothesis that remote preconditioning acts through sensory nerve stimulation in the ischemic organ.
Orosomucoid has a cAMP-dependent effect on human endothelial cells and inhibits the action of histamine
Authors: Sörensson Jenny , Ohlson Maria , Björnson Anna , Haraldsson Börje
Published: December 2017

The plasma protein orosomucoid (α1-acid glycoprotein) has previously been shown to constitute a critical component of the capillary barrier. The protein has also been suggested to act as an anti-inflammatory mediator in a diversity of experimental situations. Recently we reported that orosomucoid is synthesized by the microvascular endothelial cells per se. In the present study, the effects of orosomucoid on primary cultures of human umbilical vein endothelial cells (HUVEC) were studied using the Cytosensor microphysiometer. We found that 1) orosomucoid (0.01 g/l) increased the metabolic activity of HUVEC as reflected by the increased acidification rate of +14 ± 1%; 2) pretreatment with 0.5 mM 8-bromo-cAMP for 20 min markedly and reversibly inhibited the effect of orosomucoid, whereas 8-bromo-cGMP did not; 3) histamine elicited a dose-dependent response that was abolished by pretreatment with either cAMP or cGMP; and finally, 4) pretreatment of HUVEC for 6 min with orosomucoid (0.01 g/l) inhibited the action of histamine. In summary, this is the first report demonstrating that orosomucoid affects human endothelial cells and that it does so by using cAMP as a second messenger. This provides an explanation for previous findings of anti-inflammatory effects of the protein and shows that orosomucoid affects the endothelium during both normal and pathophysiological conditions.
Metabolism of VLDL is increased in streptozotocin-induced diabetic rat hearts
Authors: Sambandam Nandakumar , Abrahani Mohammed A. , Craig Scott , Al-Atar Osama , Jeon Esther , Rodrigues Brian
Published: December 2017

In streptozotocin (STZ)-induced diabetic rats, we previously showed an increased heparin-releasable (luminal) lipoprotein lipase (LPL) activity from perfused hearts. To study the effect of this enlarged LPL pool on triglyceride (TG)-rich lipoproteins, we examined the metabolism of very-low-density lipoprotein (VLDL) perfused through control and diabetic hearts. Diabetic rats had elevated TG levels compared with control. However, fasting for 16 h abolished this difference. When the plasma lipoprotein fraction of density <1.006 g/ml from fasted control and diabetic rats was incubated in vitro with purified bovine or rat LPL, VLDL from diabetic animals was hydrolyzed as proficiently as VLDL from control animals. Post-heparin plasma lipolytic activity was comparable in control and diabetic animals. However, perfusion of control and diabetic rats with heparinase indicated that diabetic hearts had larger amounts of LPL bound to heparan sulfate proteoglycan-binding sites. [3H]VLDL obtained from control rats, when recirculated through the isolated heart, disappeared at a significantly faster rate from diabetic than from control rat hearts. This increased VLDL-TG hydrolysis was essentially abolished by prior perfusion of the diabetic heart with heparin, implicating LPL in this process. These findings suggest that the enlarged LPL pool in the diabetic heart is present at a functionally relevant location (at the capillary lumen) and is capable of hydrolyzing VLDL. This could increase the delivery of free fatty acid to the heart, and the resultant metabolic changes could induce the subsequent cardiomyopathy that is observed in the chronic diabetic rat.
SERCA2a activity correlates with the force-frequency relationship in human myocardium
Authors: Münch Götz , Bölck Birgit , Brixius Klara , Reuter Hannes , Mehlhorn Uwe , Bloch Wilhelm , Schwinger Robert H. G.
Published: December 2017

The present investigation addresses whether protein expression and function of sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA2a) and phospholamban (PLB) correlate in failing and nonfailing human myocardium. SERCA2a activity and protein expression, PLB phosphorylation, and the force-frequency relationship (FFR) have been determined in right atrium (RA) and left ventricle (LV) from nonfailing (NF, n = 12) and terminally failing [dilated cardiomyopathy (DCM), n = 12] human hearts. Only in LV of DCM hearts was SERCA2a activity significantly decreased [maximal turnover rate ( Vmax) = 196 ± 11 and 396 ± 30 nmol ⋅ mg−1⋅ min−1in LV and RA, respectively], whereas protein expression of SERCA2a in the different chambers was unchanged in NF (3.9 ± 0.3 and 3.2 ± 0.4 densitometric units in LV and RA, respectively) and DCM hearts (4.8 ± 0.8 and 3.4 ± 0.1 densitometric units in LV and RA, respectively). Phosphorylation of PLB was higher in LV than in RA in NF (Ser16: 180.5 ± 19.0 vs. 56.8 ± 6.0 densitometric units; Thr17: 174.6 ± 11.2 vs. 37.4 ± 8.9 densitometric units) and DCM hearts (Ser16: 132.0 ± 5.4 vs. 22.4 ± 3.5 densitometric units; Thr17: 131.2 ± 10.9 vs. 9.2 ± 2.4 densitometric units). SERCA2a function, but not protein expression, correlated well with the functional parameters of the FFR in DCM and NF human hearts. Regulation of SERCA2a function depends on the phosphorylation of PLB at Ser16and Thr17. However, direct SERCA2a regulation might also be affected by an unknown mechanism.
Nitric oxide modulation of TNF-α-induced cardiac contractile dysfunction is concentration dependent
Authors: Horton Jureta W. , Maass David , White Jean , Sanders Billy
Published: December 2017

Whereas previous studies suggest that tumor necrosis factor-α (TNF-α) induces cardiac contraction-relaxation deficits, the mechanisms remain unclear. Our recent studies have implicated cardiac-derived nitric oxide (NO). This study examined the detrimental and protective effects of NO donors S-nitroso- N-acetyl-penicillamine (SNAP) or ( Z)-1- [ N-(3-ammonio-propyl)- N-( n-propyl)amino]diazen-1-ium- 1,2diolate (PAPA/NO) on TNF-α-related changes in cardiac contractile function (Langendorff), cellular injury, and intracellular myocyte Ca2+concentration ([Ca2+]i). Myocytes were incubated in the presence/absence of TNF-α (200–500 pg/ml × 105cells) for 3 h; subsets of myocytes were incubated with one of several concentrations of SNAP or PAPA/NO (0.1, 0.3, 0.5, and 1.5 mM) for 15 min before TNF-α challenge. Supernatant creatine kinase (CK), cell viability (Trypan blue dye exclusion), and myocyte [Ca2+]i(fura 2-acetoxymethyl ester) were measured. In parallel experiments, cardiac function (Langendorff) was examined after TNF-α challenge in the presence or absence of SNAP or PAPA/NO (0.1 and 1.5 mM). TNF-α in the absence of an NO donor impaired cardiac contraction and relaxation and produced cardiomyocyte injury. Pretreating perfused hearts or isolated cardiomyocytes with a low concentration of either SNAP or PAPA/NO decreased TNF-α-mediated cardiac injury and improved contractile dysfunction, whereas high concentrations of NO donor exacerbated TNF-α-mediated cardiac effects. These data provide one explanation for the conflicting reports of beneficial versus detrimental effects of NO in the face of inflammation and suggest that the effects of NO on organ function are concentration dependent; low concentrations of NO are cardioprotective, whereas high concentrations of NO are deleterious.
Influence of glycogen storage on vascular smooth muscle metabolism
Authors: Allen Tara J. , Hardin Christopher D.
Published: December 2017

The role of glycogen as an oxidative substrate for vascular smooth muscle (VSM) remains controversial. To elucidate the importance of glycogen as an oxidative substrate and the influence of glycogen flux on VSM substrate selection, we systematically altered glycogen levels and measured metabolism of glucose, acetate, and glycogen. Hog carotid arteries with glycogen contents ranging from 1 to 11 μmol/g were isometrically contracted in physiological salt solution containing 5 mM [1-13C]glucose and 1 mM [1,2-13C]acetate at 37°C for 6 h. [1-13C]glucose, [1,2-13C]acetate, and glycogen oxidation were simultaneously measured with the use of a 13C-labeled isotopomer analysis of glutamate. Although oxidation of glycogen increased with the glycogen content of the tissue, glycogen oxidation contributed only ∼10% of the substrate oxidized by VSM. Whereas [1-13C]glucose flux, [3-13C]lactate production from [1-13C]glucose, and [1,2-13C]acetate oxidation were not regulated by glycogen content, [1-13C]glucose oxidation was significantly affected by the glycogen content of VSM. However, [1-13C]glucose remained the primary (∼40–50%) contributor to substrate oxidation. Therefore, we conclude that glucose is the predominate substrate oxidized by VSM, and glycogen oxidation contributes minimally to substrate oxidation.