bluuebunny/crossref_metadata_2025 huggingface dataset:
Adenosine A3receptor activation protects the myocardium from reperfusion/reoxygenation injury
Authors: Maddock Helen L. , Mocanu Mihaela M. , Yellon Derek M.
Published: March 2015

Ischemia-reperfusion induces both necrotic and apoptotic cell death. The ability of adenosine to attenuate reperfusion-induced injury (RI) and the role played by adenosine receptors are unclear. We therefore studied the role of the A3receptor (A3R) in ameliorating RI using the specific A3R agonist 1-[2-chloro-6-[[(3-iodophenyl)methyl]amino]-9 H-purin-9-yl]-1-deoxi- N-methyl-b-d-ribofuranuronamide (2-Cl-IB-MECA). Isolated rat hearts and cardiomyocytes were subjected to ischemia or simulated ischemia, followed by reperfusion/reoxygenation. The end points were percent infarction/risk zone and annexin-V (apoptosis) and/or propidium iodide positivity (necrosis), respectively. In isolated hearts, 2-Cl-IB-MECA significantly limited infarct size (44.2 ± 2.7% in control vs. 21.9 ± 2.4% at 1 nM and 35.8 ± 3.3% at 0.1 nM, P < 0.05). In isolated myocytes, apoptosis and necrosis were significantly reduced compared with controls (5.7 ± 2.6% vs. 17.1 ± 1.3% and 13.7 ± 2.0% vs. 23.1 ± 1.5%, respectively, P < 0.0001). In both models, the beneficial effects were abrogated using the A3R antagonist MRS-1191. The involvement of A2areceptor activation was also examined. This is the first study to demonstrate that A3R activation at reperfusion limits myocardial injury in the isolated rat heart and improves survival in isolated myocytes, possibly by antiapoptotic and antinecrotic mechanisms.
Proteasome inhibition ablates activation of NF-κB in myocardial reperfusion and reduces reperfusion injury
Authors: Pye Joseph , Ardeshirpour Farhad , McCain Arlene , Bellinger Dwight A. , Merricks Elizabeth , Adams Julian , Elliott Peter J. , Pien Christine , Fischer Thomas H. , Baldwin Albert S. , Nichols Timothy C.
Published: March 2015

Both acute coronary occlusion and reperfusion of an infarct-related artery lead to significant myocardial cell death. Recent evidence has been presented that activation of the transcription factor nuclear factor-κB (NF-κB) plays a critical role in reperfusion injury. NF-κB is usually bound to its inhibitor, IκB, and classic activation of NF-κB occurs when the 20S proteasome degrades IκB that has been phosphorylated and ubiquitinated. In this study, activation of NF-κB was inhibited by systemic administration of a 20S proteasome inhibitor (PS-519) in a porcine model of myocardial reperfusion injury. The experimental protocol induced myocardial ischemia in the distribution of the left anterior descending coronary artery for 1 h with subsequent reperfusion for 3 h. A single systemic treatment with PS-519 reduced 20S proteasome activity; blocked activation of NF-κB induced by reperfusion; reduced creatine kinase, creatine kinase-muscle-brain fraction, and troponin I release from the myocardium; preserved regional myocardial function measured by segmental shortening; significantly reduced the size of myocardial infarction; and exhibited no acute toxicity. These data show that myocardial reperfusion injury can be inhibited by using proteasome inhibitors, which likely function through the inhibition of NF-κB activation.
Desipramine attenuates loss of cardiac sympathetic neurotransmitters produced by congestive heart failure and NE infusion
Authors: Liang Chang-seng , Yatani Akito , Himura Yoshihiro , Kashiki Michihiro , Stevens Susanne Y.
Published: March 2015

We reported recently that inhibition of neuronal reuptake of norepinephrine (NE) by desipramine prevented the reduction of sympathetic neurotransmitters in the failing right ventricle of right heart failure animals. In this study, we studied whether desipramine also reduced the sympathetic neurotransmitter loss in animals with left heart failure induced by rapid ventricular pacing (225 beats/min) or after chronic NE infusion (0.5 μg · kg−1 · min−1). Desipramine was given to the animals for 8 wk beginning with rapid ventricular pacing or NE infusion. Animals receiving no desipramine were studied as controls. We measured myocardial NE content, NE uptake activity, and sympathetic NE, tyrosine hydroxylase, and neuropeptide Y profiles by histofluorescence and immunocytochemical techniques. Effects of desipramine on NE uptake inhibition were evidenced by potentiation of the pressor response to exogenous NE and reduction of myocardial NE uptake activity. Desipramine treatment had no effect in sham or saline control animals but attenuated the reduction of sympathetic neurotransmitter profiles in the left ventricles of animals with rapid cardiac pacing and NE infusion. In contrast, the panneuronal marker protein gene product 9.5 profile was not affected by either rapid pacing or NE infusion, nor was it changed by desipramine treatment in the heart failure animals. The study confirms that excess NE contributes to the reduction of cardiac sympathetic neurotransmitters in heart failure. In addition, it shows that the anatomic integrity of the sympathetic nerves is relatively intact and that the neuronal damaging effect of NE involves the uptake of NE or its metabolites into the sympathetic nerves.
Differential response to myocardial reperfusion injury in eNOS-deficient mice
Authors: Sharp Brent R. , Jones Steven P. , Rimmer David M. , Lefer David J.
Published: March 2015

Two strains of endothelial nitric oxide synthase (eNOS)-deficient (−/−) mice have been developed that respond differently to myocardial ischemia-reperfusion (MI/R). We evaluated both strains of eNOS−/− mice in an in vivo model of MI/R. Harvard (Har) eNOS−/− mice ( n = 12) experienced an 84% increase in myocardial necrosis compared with wild-type controls ( P < 0.05). University of North Carolina (UNC) eNOS−/−( n = 10) exhibited a 52% reduction in myocardial injury versus wild-type controls ( P < 0.05). PCR analysis of myocardial inducible NO synthase (iNOS) mRNA levels revealed a significant ( P < 0.05) increase in the UNC eNOS−/− mice compared with wild-type mice, and there was no significant difference between the Har eNOS−/− and wild-type mice. UNC eNOS−/− mice treated with an iNOS inhibitor (1400W) exacerbated the extent of myocardial necrosis. When treated with 1400W, Har eNOS−/− did not exhibit a significant increase in myocardial necrosis. These data demonstrate that two distinct strains of eNOS−/− mice display opposite responses to MI/R. Although the protection seen in the UNC eNOS−/− mouse may result from compensatory increases in iNOS, other genes may be involved.
Upregulation of Cdc2 and cyclin A during apoptosis of endothelial cells induced by cleaved high-molecular-weight kininogen
Authors: Wang Shujie , Hasham Muneer G. , Isordia-Salas Irma , Tsygankov Alexander Y. , Colman Robert W. , Guo Yan-Lin
Published: March 2015

We (8) reported that the cleaved high-molecular-weight kininogen (HKa) and its domain 5 (D5) inhibited angiogenesis. Further studies (15) revealed that D5 could inhibit cell proliferation and induce apoptosis of proliferating endothelial cells, which together may represent a critical part of antiangiogenic activity of HKa and D5. In the present study, we further examined the effect of HKa on cell cycle progression and cell viability. We report that HKa induced a significant upregulation of Cdc2 and cyclin A in proliferating endothelial cells, concurrent with a marked increase of Cdc2 activity. The increased expression of Cdc2 and cyclin A by HKa was not associated with an apparent change in cell cycle profiles of basic fibroblast growth factor-stimulated proliferating cells, but closely correlated with a marked increase of apoptosis, suggesting that the elevated Cdc2 activity is involved in HKa-induced apoptosis of proliferating endothelial cells. Our results support an emerging hypothesis that Cdc2 and cyclin A are important regulators for cell cycle as well as for apoptosis.
Adenosine A1-receptor induced late preconditioning and myocardial infarction: reperfusion duration is critical
Authors: Tissier Renaud , Souktani Rachid , Bruneval Patrick , Giudicelli Jean-François , Berdeaux Alain , Ghaleh Bijan
Published: March 2015

We investigated the influence of coronary artery reperfusion (CAR) duration on the infarct-limiting properties of adenosine A1-receptor stimulation-induced delayed preconditioning (A1-DPC) compared with ischemia-induced delayed preconditioning (I-DPC). Sixty-one chronically instrumented conscious rabbits successfully underwent the following protocol. On day 1, rabbits were randomly divided into four groups: control (saline, iv), I-DPC (six 4-min coronary artery occlusion/4-min reperfusion cycles), A1-DPC100( N 6-cyclopentyladenosine, 100 μg/kg iv), and A1-DPC400( N 6-cyclopentyladenosine, 400 μg/kg iv). On day 2 (i.e., 24 h later), rabbits underwent a 30-min coronary artery occlusion after which CAR was started and maintained for either 3 or 72 h. Infarct size (percentage of the area at risk) was determined by triphenyltetrazolium chloride staining. After 3 h of CAR, I-DPC, A1-DPC100, and A1-DPC400 significantly decreased infarct size (36 ± 5, 41 ± 4, 38 ± 5%, respectively) compared with control (55 ± 3%). After 72 h of CAR, infarct sizes were not significantly different among the four groups. This result was confirmed by histologic analysis. Thus A1-DPC at the two investigated doses, as well as I-DPC, decreased infarct size after 3 h but not 72 h of CAR.
Role of Src protein tyrosine kinases in late preconditioning against myocardial infarction
Authors: Dawn Buddhadeb , Takano Hitoshi , Tang Xian-Liang , Kodani Eitaro , Banerjee Supratim , Rezazadeh Arash , Qiu Yumin , Bolli Roberto
Published: March 2015

Although Src protein tyrosine kinases (PTKs) have been shown to be essential in late preconditioning (PC) against myocardial stunning, their role in triggering versus mediating late PC against myocardial infarction remains unclear. Four groups of conscious rabbits were subjected to a 30-min coronary occlusion on day 2, with or without PC ischemia on day 1. Administration of the Src PTK inhibitor lavendustin A (LD-A; 1 mg/kg iv) before the PC ischemia on day 1 ( group III, n = 7) failed to block the delayed protective effect against myocardial infarction 24 h later. Late PC against infarction, however, was completely abrogated when LD-A was given 24 h after the PC ischemia, prior to the 30-min occlusion on day 2 ( group IV, n = 8). We conclude that, in conscious rabbits, Src PTK activity is necessary for the mediation of late PC protection against myocardial infarction on day 2, but not for the initiation of this phenomenon on day 1. Taken together with previous studies in the setting of stunning, these findings reveal heretofore unrecognized differences in the roles of Src PTKs in late PC against stunning versus late PC against infarction.
Type II secretory phospholipase A2binds to ischemic flip-flopped cardiomyocytes and subsequently induces cell death
Authors: Nijmeijer R. , Willemsen M. , Meijer C. J. L. M. , Visser C. A. , Verheijen R. H. , Gottlieb R. A. , Hack C. E. , Niessen H. W. M.
Published: March 2015

Type II secretory phospholipase A2(sPLA2) is a cardiovascular risk factor. We recently found depositions of sPLA2in the necrotic center of infarcted human myocardium and normally appearing cardiomyocytes adjacent to the border zone. The consequences of binding of sPLA2to ischemic cardiomyocytes are not known. To explore a potential effect of sPLA2on ischemic cardiomyocytes at a cellular level we used an in vitro model. The cardiomyocyte cell line H9c2 or adult cardiomyocytes were isolated from rabbits that were incubated with sPLA2in the presence of metabolic inhibitors to mimic ischemia-reperfusion conditions. Cell viability was established with the use of annexin V and propidium iodide or 7-aminoactinomycin D. Metabolic inhibition induced an increase of the number of flip-flopped cells, including a population that did not stain with propidium iodide and that was caspase-3 negative. sPLA2bound to the flip-flopped cells, including those negative for caspase-3. sPLA2binding induced cell death in these latter cells. In addition, sPLA2potentiated the binding of C-reactive protein (CRP) to these cells. We conclude that by binding to flip-flopped cardiomyocytes, including those that are caspase-3 negative and presumably reversibly injured, sPLA2may induce cell death and tag these cells with CRP.
Effect of aggregation and shear rate on the dispersion of red blood cells flowing in venules
Authors: Bishop Jeffrey J. , Popel Aleksander S. , Intaglietta Marcos , Johnson Paul C.
Published: March 2015

Previous in vitro studies of blood flow in small glass tubes have shown that red blood cells exhibit significant erratic deviations in the radial position in the laminar flow regime. The purpose of the present study was to assess the magnitude of this variability and that of velocity in vivo and the effect of red blood cell aggregation and shear rate upon them. With the use of a gated image intensifier and fluorescently labeled red blood cells in tracer quantities, we obtained multiple measurements of red blood cell radial and longitudinal positions at time intervals as short as 5 ms within single venous microvessels (diameter range 45–75 μm) of the rat spinotrapezius muscle. For nonaggregating red blood cells in the velocity range of 0.3–14 mm/s, the mean coefficient of variation of velocity was 16.9 ± 10.5% and the SD of the radial position was 1.98 ± 0.98 μm. Both quantities were inversely related to shear rate, and the former was significantly lowered on induction of red blood cell aggregation by the addition of Dextran 500 to the blood. The shear-induced random movements observed in this study may increase the radial transport of particles and solutes within the bloodstream by orders of magnitude.
Ca2+activation and tension cost in myofilaments from mouse hearts ectopically expressing enteric γ-actin
Authors: Martin Anne F. , Phillips Ronald M. , Kumar Ajit , Crawford Kelly , Abbas Zainab , Lessard James L. , de Tombe Pieter , Solaro R. John
Published: March 2015

To determine the significance of actin isoforms in chemomechanical coupling, we compared tension and ATPase rate in heart myofilaments from nontransgenic (NTG) and transgenic (TG) mice in which enteric γ-actin replaced >95% of the cardiac α-actin. Enteric γ-actin was expressed against three backgrounds: mice expressing cardiac α-actin, heterozygous null cardiac α-actin mice, and homozygous null cardiac α-actin mice. There were no differences in maximum Ca2+activated tension or maximum rate of tension redevelopment after a quick release and rapid restretch protocol between TG and NTG skinned fiber bundles. However, compared with NTG controls, Ca2+sensitivity of tension was significantly decreased and economy of tension development was significantly increased in myofilaments from all TG hearts. Shifts in myosin isoform population could not fully account for this increase in the economy of force production of TG myofilaments. Our results indicate that an exchange of cardiac α-actin with an actin isoform differing in only five amino acids has a significant impact on both Ca2+regulation of cardiac myofilaments and the cross-bridge cycling rate.
Structural mechanisms of acute VEGF effect on microvessel permeability
Authors: Fu Bingmei M. , Shen Shang
Published: March 2015

To investigate the ultrastructural mechanisms of acute microvessel hyperpermeability by vascular endothelial growth factor (VEGF), we combined a mathematical model ( J Biomech Eng 116: 502–513, 1994) with experimental data of the effect of VEGF on microvessel hydraulic conductivity ( Lp) and permeability of various-sized solutes. We examined the effect of VEGF on microvessel permeability to a small solute (sodium fluorescein, Stokes radius 0.45 nm), an intermediate solute (α-lactalbumin, Stokes radius 2.01 nm), and a large solute [albumin (BSA), Stokes radius 3.5 nm]. Exposure to 1 nM VEGF transiently increased apparent permeability to 2.3, 3.3, and 6.2 times their baseline values for sodium fluorescein, α-lactalbumin, and BSA, respectively, within 30 s, and all returned to control within 2 min. On the basis of Lp(DO Bates and FE Curry. Am J Physiol Heart Circ Physiol 271: H2520–H2528, 1996) and permeability data, the prediction from the model suggested that the most likely structural changes in the interendothelial cleft induced by VEGF would be a ∼2.5-fold increase in its opening width and partial degradation of the surface glycocalyx.
Increased vulnerability to inducible atrial fibrillation caused by partial cellular uncoupling with heptanol
Authors: Ohara Toshihiko , Qu Zhilin , Lee Moon-Hyoung , Ohara Keiko , Omichi Chikaya , Mandel William J. , Chen Peng-Sheng , Karagueuzian Hrayr S.
Published: March 2015

We hypothesized that partial cellular uncoupling produced by low concentrations of heptanol increases the vulnerability to inducible atrial fibrillation (AF). The epicardial surface of 12 isolated-perfused canine left atria was optically mapped before and after 1–50 μM heptanol infusion. At baseline, no sustained (>30 s) AF could be induced in any of the 12 tissues. However, after 2 μM heptanol infusion, sustained AF was induced in 9 of 12 tissues ( P < 0.001). Heptanol >5 μM caused loss of 1:1 capture during rapid pacing, causing no AF to be induced. AF was initiated by conduction block across the fiber leading to reentry, which broke up after one to two rotations into two to four independent wavelets that sustained the AF. Heptanol at 2 μM had no effect on the cellular action potential duration restitution or on the maximal velocity rate over time of the upstroke. The effects of heptanol were reversible. We conclude that partial cellular uncoupling by heptanol without changing atrial active membrane properties promotes wavebreak, reentry, and AF during rapid pacing.
Cardiac troponin T isoforms demonstrate similar effects on mechanical performance in a regulated contractile system
Authors: VanBuren Peter , Alix Shari L. , Gorga Joseph A. , Begin Kelly J. , LeWinter Martin M. , Alpert Norman R.
Published: March 2015

Alteration of troponin T (TnT) isoform expression has been reported in human and animal models of myocardial failure. The two adult beef cardiac TnT isoforms (TnT3and TnT4) were isolated for comparative functional analysis. Thin filaments were reconstituted containing pure populations of the isoforms. The in vitro motility assay was used to directly compare the effect of the two TnT isoforms on force and unloaded shortening as a function of free calcium. We found no significant differences between the two isoforms in terms of calcium sensitivity, cooperativity, or maximal activation (velocity and force) as assessed in a fully calcium-regulated system. Activation by myosin strong binding was similar for thin filaments containing either of the two TnT isoforms. Whereas maximally activated velocity and cooperativity was depressed at pH 6.5, no difference between thin filaments containing the two isoforms was detected. From the small magnitude of the TnT isoform shifts detected in myocardial failure and the lack of significant mechanical effect detected in the motility assay, variable TnT isoform expression is unlikely to be any functional significance in heart failure.
Syndecan-4 modulates basic fibroblast growth factor 2 signaling in vivo
Authors: Zhang Yufeng , Li Jianyi , Partovian Chohreh , Sellke Frank W. , Simons Michael
Published: March 2015

Syndecan-4 is one of the principal heparan sulfate-carrying proteins on the cell surface. Unlike other members of syndecan family, syndecan-4 mediates phosphatidylinositol 4,5-bisphosphate 2 (PIP2)-dependent PKC-α activation, and overexpression of syndecan-4 in vitro results in enhanced FGF2 signaling. The present study was designed to test the functional effect of increased syndecan-4 expression in endothelial cells in transgenic mice. Several transgenic mice lines expressing syndecan-4 cDNA under control of human endothelial nitric oxide (NO) synthase (eNOS) promoter were generated. Exogenous syndecan-4 was mainly expressed in the heart, brain, and lungs. In particular, the heart demonstrated the greatest increase in the ratio of transgenic-to-native syndecan-4 gene expression. Vessels from the eNOS-syndecan-4 mice demonstrated more pronounced vasodilation to FGF2 but not to VEGF-A165, sodium nitroprusside, and A 23187 compared with wild-type mice. To elucidate the mechanism of this effect, we measured NO release from primary cardiac endothelial cells isolated from transgenic or wild-type adult mice. Cells from the eNOS-syndecan-4 transgenic mice had a significant increase in FGF2- and VEGF-A165-induced NO release compared with endothelial cells from the wild-type mice. However, the absolute magnitude of this increase was higher for FGF2 than VEGF-A165. In conclusion, enhanced syndecan-4 expression in mouse cardiac endothelial cells results in preferential augmentation of FGF2 but not VEGF-A165-induced NO release.
Metalloendopeptidases EC 3.4.24.15/16 regulate bradykinin activity in the cerebral microvasculature
Authors: Norman M. Ursula , Lew Rebecca A. , Smith A. Ian , Hickey Michael J.
Published: March 2015

Bradykinin is a vasoactive peptide that has been shown to increase the permeability of the cerebral microvasculature to blood-borne macromolecules. The two zinc metalloendopeptidases EC 3.4.24.15 (EP 24.15) and EC 3.4.24.16 (EP 24.16) degrade bradykinin in vitro and are highly expressed in the brain. However, the role that these enzymes play in bradykinin metabolism in vivo remains unclear. In the present study, we investigated the role of EP 24.15 and EP 24.16 in the regulation of bradykinin-induced alterations in microvascular permeability. Permeability of the cerebral microvasculature was assessed in anesthetized Sprague-Dawley rats by measuring the clearance of 70-kDa FITC dextran from the brain. Inhibition of EP 24.15 and EP 24.16 by the specific inhibitor N-[1-( R, S)-carboxy-3-phenylpropyl]-Ala-Aib-Tyr- p-aminobenzoate (JA-2) resulted in the potentiation of bradykinin-induced increases in cerebral microvessel permeability. The level of potentiation was comparable to that achieved by the inhibition of angiotensin-converting enzyme. These findings provide the first evidence of an in vivo role for EP 24.15/EP 24.16 in brain function, specifically in regulating alterations in microvessel permeability induced by exogenous bradykinin.
β-Adrenergic and antiadrenergic modulation of cardiac adenylyl cyclase is influenced by phosphorylation
Authors: Dobson James G. , Shea Lynne G. , Fenton Richard A.
Published: March 2015

Adenosine protects the myocardium of the heart by exerting an antiadrenergic action via the adenosine A1receptor (A1R). Because β1-adrenergic receptor (β1R) stimulation elicits myocardial protein phosphorylation, the present study investigated whether protein kinase A (PKA) catalyzed rat heart ventricular membrane phosphorylation affects the β1R adrenergic and A1R adenosinergic actions on adenylyl cyclase activity. Membranes were either phosphorylated with PKA in the absence/presence of a protein kinase inhibitor (PKI) or dephosphorylated with alkaline phosphatase (AP) and assayed for adenylyl cyclase activity (AC) in the presence of the β1R agonist isoproterenol (ISO) and/or the A1R agonist 2-chloro- N6-cyclopentyladenosine (CCPA).32P incorporation into the protein substrates of 140–120, 43, and 29 kDa with PKA increased both the ISO-elicited activation of AC by 51–54% and the A1R-mediated reduction of the ISO-induced increase in AC by 29–50%, thereby yielding a total antiadrenergic effect of ∼78%. These effects of PKA were prevented by PKI. AP reduced the ISO-induced increase in AC and eliminated the antiadrenergic effect of CCPA. Immunoprecipitation of the solubilized membrane adenylyl cyclase with the use of a polyclonal adenylyl cyclase VI antibody indicated that the enzyme is phosphorylated by PKA. These results indicate that the cardioprotective effect of adenosine afforded by its antiadrenergic action is facilitated by cardiac membrane phosphorylation.
Block of the background K+channel TASK-1 contributes to arrhythmogenic effects of platelet-activating factor
Authors: Barbuti Andrea , Ishii Satoshi , Shimizu Takao , Robinson Richard B. , Feinmark Steven J.
Published: March 2015

Platelet-activating factor (PAF), an inflammatory phospholipid, induces ventricular arrhythmia via an unknown ionic mechanism. We can now link PAF-mediated cardiac electrophysiological effects to inhibition of a two-pore domain K+channel [TWIK-related acid-sensitive K+background channel (TASK-1)]. Superfusion of carbamyl-PAF (C-PAF), a stable analog of PAF, over murine ventricular myocytes causes abnormal automaticity, plateau phase arrest of the action potential, and early afterdepolarizations in paced and quiescent cells from wild-type but not PAF receptor knockout mice. C-PAF-dependent currents are insensitive to Cs+and are outwardly rectifying with biophysical properties consistent with a K+-selective channel. The current is blocked by TASK-1 inhibitors, including protons, Ba2+, Zn2+, and methanandamide, a stable analog of the endogenous lipid ligand of cannabanoid receptors. In addition, when TASK-1 is expressed in CHO cells that express an endogenous PAF receptor, superfusion of C-PAF decreases the expressed current. Like C-PAF, methanandamide evoked spontaneous activity in quiescent myocytes. C-PAF- and methanandamide-sensitive currents are blocked by a specific protein kinase C (PKC) inhibitor, implying overlapping signaling pathways. In conclusion, C-PAF blocks TASK-1 or a closely related channel, the effect is PKC dependent, and the inhibition alters the electrical activity of myocytes in ways that would be arrhythmogenic in the intact heart.
Sex differences in repolarization homogeneity and its circadian pattern
Authors: Smetana Peter , Batchvarov Velislav N. , Hnatkova Katerina , Camm A. John , Malik Marek
Published: March 2015

The reason for sex differences in arrhythmic risk remains unclear. Heterogeneity of ventricular repolarization is directly linked to arrhythmogenesis; thus we investigated repolarization homogeneity and its circadian pattern in men and women. During 24-h Holter recordings in 60 healthy subjects (27 males), a 12-lead electrocardiogram (ECG) was obtained every 30 s. RR and QT intervals, and, after singular-value decomposition, two characteristics of repolarization homogeneity were calculated in each ECG. Corrected QT (QTc) values were obtained using an individually optimized heart rate (HR) correction formula. All values were averaged over 10-min time bands from 0000 to 2400. There were substantial sex differences in both global repolarization homogeneity (measured by the total cosine of the angle between QRS and T wave vectors) and regional homogeneity of repolarization (quantified independently by the relative T wave residuum). Whereas women throughout the 24 h followed more closely the pattern of inverse sequence between depolarization and repolarization, they also showed much higher localized repolarization heterogeneity than men. In both women and men, repolarization irregularity was greatest during morning hours. A sex difference was also observed for HR and QTc interval; however, the circadian patterns of the repolarization homogeneity descriptors were different from those of HR and QTc intervals.
Targeted deletion of adenosine A3 receptors augments adenosine-induced coronary flow in isolated mouse heart
Authors: Talukder M. A. Hassan , Morrison R. Ray , Jacobson Marlene A. , Jacobson Kenneth A. , Ledent Catherine , Mustafa S. Jamal
Published: March 2015

To determine whether adenosine A3 receptors participate in adenosine-induced changes in coronary flow, isolated hearts from wild-type (WT) and A3 receptor knockout (A3KO) mice were perfused under constant pressure and effects of nonselective and selective agonists were examined. Adenosine and the selective A2A agonist 2-[ p-(2-carboxyethyl)]phenylethylamino-5′- N-ethylcarboxamidoadenosine (CGS-21680) produced augmented maximal coronary vasodilation in A3KO hearts compared with WT hearts. Selective activation of A3 receptors with 2-chloro- N 6-(3-iodobenzyl)-adenosine-5′- N-methyluronamide (Cl-IB-MECA) at nanomolar concentrations did not effect coronary flow, but at higher concentrations it produced coronary vasodilation both in WT and A3KO hearts. Cl-IB-MECA-induced increases in coronary flow were susceptible to both pharmacological blockade and genetic deletion of A2A receptors. Because deletion or blockade of adenosine A3 receptors augmented coronary flow induced by nonselective adenosine and the selective A2A receptor agonist CGS-21680, we speculate that this is due to removal of an inhibitory influence associated with the A3 receptor subtype. These data indicate that the presence of adenosine A3 receptors may either inhibit or negatively modulate coronary flow mediated by other adenosine receptor subtypes.
Pathophysiology of orthostatic hypotension after bed rest: paradoxical sympathetic withdrawal
Authors: Kamiya Atsunori , Michikami Daisaku , Fu Qi , Iwase Satoshi , Hayano Junichiro , Kawada Toru , Mano Tadaaki , Sunagawa Kenji
Published: March 2015

Although orthostatic hypotension is a common clinical syndrome after spaceflight and its ground-based simulation model, 6° head-down bed rest (HDBR), the pathophysiology remains unclear. The authors' hypothesis that a decrease in sympathetic nerve activity is the major pathophysiology underlying orthostatic hypotension after HDBR was tested in a study involving 14-day HDBR in 22 healthy subjects who showed no orthostatic hypotension during 15-min 60° head-up tilt test (HUT) at baseline. After HDBR, 10 of 22 subjects demonstrated orthostatic hypotension during 60° HUT. In subjects with orthostatic hypotension, total activity of muscle sympathetic nerve activity (MSNA) increased less during the first minute of 60° HUT after HDBR (314% of resting supine activity) than before HDBR (523% of resting supine activity, P < 0.05) despite HDBR-induced reduction in plasma volume (13% of plasma volume before HDBR). The postural increase in total MSNA continued during several more minutes of 60° HUT while arterial pressure was maintained. Thereafter, however, total MSNA was paradoxically suppressed by 104% of the resting supine level at the last minute of HUT ( P < 0.05 vs. earlier 60° HUT periods). The suppression of total MSNA was accompanied by a 22 ± 4-mmHg decrease in mean blood pressure (systolic blood pressure <80 mmHg). In contrast, orthostatic activation of total MSNA was preserved throughout 60° HUT in subjects who did not develop orthostatic hypotension. These data support the hypothesis that a decrease in sympathetic nerve activity is the major pathophysiological factor underlying orthostatic hypotension after HDBR. It appears that the diminished sympathetic activity, in combination with other factors associated with HDBR (e.g., hypovolemia), may predispose some individuals to postural hypotension.
Expression of kinin B1 receptor in fresh or cultured rabbit aortic smooth muscle: role of NF-κB
Authors: Sabourin Thierry , Morissette Guillaume , Bouthillier Johanne , Levesque Luc , Marceau François
Published: March 2015

Kinin B1receptor (B1R) expression and the importance of the transcription factor nuclear factor (NF)-κB in this process were evaluated in models based on the rabbit aorta: freshly isolated tissue (postisolation induction) and cultured smooth muscle cells (SMCs). A 3-h incubation of freshly isolated tissues determined a sharp B1R mRNA increase (RT-PCR). Coincubation of tissues with a stimulus (interleukin-1β, fetal bovine serum, epidermal growth factor, or cycloheximide) further increased mRNA levels. Cultured SMCs possessed a basal population of surface B1Rs ([3H]Lys-des-Arg9-bradykinin binding) that was upregulated by treatments with the same set of stimuli (binding, mRNA, nuclear runon). Pharmacological inhibitors of NF-κB (MG-132, BAY 11-7082, dexamethasone) or actinomycin D reduced the postisolation induction of B1Rs in fresh aortic tissue (contractility or mRNA) and the cytokine effect on cells (mRNA, binding). NF-κB may be a common mediator of various stimuli that increase B1R gene transcription in the rabbit aorta, including tissue isolation, but cycloheximide also stabilizes B1R mRNA. The SMC models faithfully mimic the in vivo situation with regard to B1R regulation.
ANG II increases TIMP-1 expression in rat aortic smooth muscle cells in vivo
Authors: Castoldi Giovanna , di Gioia Cira R. T. , Pieruzzi Federico , D'Orlando Cristina , van de Greef Willy M. M. , Busca Giuseppe , Sperti Giovanni , Stella Andrea
Published: March 2015

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) are involved in tissue remodeling processes. TIMP-1 is the main native inhibitor of MMPs and it contributes to the development of tissue fibrosis. It is known that ANG II plays a fundamental role in vascular remodeling. In this study, we investigated whether ANG II modulates TIMP-1 expression in rat aortic smooth muscle cells. In vitro, ANG II induces TIMP-1 mRNA expression in a dose-dependent manner. The maximal increase in TIMP-1 expression was present after 3 h of ANG II stimulation. The ANG II increase in TIMP-1 expression was mediated by the ANG type 1 receptors because it was blocked by losartan. The increase in TIMP-1 expression was present after the first ANG II treatment, whereas repeated treatments (3 and 5 times) did not modify TIMP-1 expression. In vivo, exogenous ANG II was administered to Sprague-Dawley rats (200 ng · kg−1· min−1sc) for 6 and 25 days. Control rats received physiological saline. After treatment, systolic blood pressure was significantly higher ( P < 0.01), whereas plasma renin activity was suppressed ( P < 0.01), in ANG II-treated rats. ANG II increased TIMP-1 expression in the aorta of ANG II-treated rats both at the mRNA ( P < 0.05) and protein levels as evaluated by Western blotting ( P < 0.05) and/or immunohistochemistry. Neither histological modifications at the vascular wall nor differences in collagen content in the tunica media were present in both the ANG II- and saline-treated groups. Our data demonstrate that ANG II increases TIMP-1 expression in rat aortic smooth muscle cells. In vivo, both short- and long-term chronic ANG II treatments increase TIMP-1 expression in the rat aorta. TIMP-1 induction by ANG II in aortic smooth muscle cells occurs in the absence of histological changes at the vascular wall.
Natriuretic peptide receptor-B in adult rat ventricle is predominantly confined to the nonmyocyte population
Authors: Doyle Donald D. , Upshaw-Earley Judy , Bell Eric L. , Palfrey H. Clive
Published: March 2015

We assessed the cellular localization and relative concentration of the C-type natriuretic peptide (CNP) guanylate cyclase-B (GC-B) receptor in the adult rat heart ventricle by several techniques. In frozen sections of the ventricle, anti-receptor antibody stained the vasculature and cells interstitial to myocytes, but not the myocytes themselves. The same antibody detected GC-B in immunoblots of protein extracts of nonmyocytes, but not myocytes and recognized an equivalent protein in extracts of cultured cardiac fibroblasts, but not A7r5 rat smooth muscle cells. In functional assays, CNP-induced cGMP accumulation per milligram cell protein was an order of magnitude greater in cultured cardiac fibroblasts than in A7r5 smooth muscle cells and two orders of magnitude greater than in freshly isolated cardiac myocytes. Modulation of cGMP accumulation by phosphodiesterases (PDEs) was cell specific as determined by antagonist pharmacological profiles, PDE1 in fibroblasts, PDE2 in A7r5 cells, and PDE3 in myocytes, suggesting that significant but low-level cGMP response to CNP measured in heart myocytes is not due to nonmyocyte contamination. Fibroblasts of cardiac origin do not show an interactive relationship between receptor responsiveness to CNP, cGMP levels, and proliferation-related mitogen-activated signal transduction pathways. Whereas previous reports suggest CNP exerts significant effects in neonatal rat cardiomyocytes, our results suggest that fibroblasts are likely the most responsive cell type (cGMP production) in the adult rat heart.
Intravital microscopy comparing T lymphocyte trafficking to the spleen and the mesenteric lymph node
Authors: Grayson Mitchell H. , Hotchkiss Richard S. , Karl Irene E. , Holtzman Michael J. , Chaplin David D.
Published: March 2015

Lymphocyte rolling velocity is determined largely by interactions between leukocyte α4-integrin (CD49d) and L-selectin and mucosal addressin cell adhesion molecule-1 (MAdCAM-1) in mesenteric postcapillary venules and Peyer's patch high endothelial venules (HEVs). The role of these interactions in other tissue sites of lymphocyte emigration is not known. With the use of real-time intravital confocal microscopy, we found that rolling velocities of T lymphocytes in the murine mesenteric lymph node (MLN) HEV also depend on L-selectin and CD49d. However, in the murine spleen, rolling velocities of T lymphocytes are not influenced by the loss of L-selectin and CD49d. With the use of FITC-dextran and TIE2-GFP mice, we further defined the microvascular compartments of the spleen and showed that adherence of T cells is localized to regions in the white pulp that are not lined by endothelial cells and have shear rates similar to bone marrow sinusoids. These results establish that T cell trafficking to the spleen differs from trafficking to other secondary lymphoid organs and suggest that the mechanical properties of the blood-filtering role of the spleen are important in T cell accumulation in the organ.
Electrophysiological response of rat atrial myocytes to acidosis
Authors: Komukai Kimiaki , Brette Fabien , Orchard Clive H.
Published: March 2015

The effect of acidosis on the electrical activity of isolated rat atrial myocytes was investigated using the patch-clamp technique. Reducing the pH of the bathing solution from 7.4 to 6.5 shortened the action potential. Acidosis had no significant effect on transient outward or inward rectifier currents but increased steady-state outward current. This increase was still present, although reduced, when intracellular Ca2+ was buffered by 1,2-bis(2-aminophenoxy)ethane- N,N,N′,N′-tetraacetic acid (BAPTA); BAPTA also inhibited acidosis-induced shortening of the action potential. Ni2+ (5 mM) had no significant effect on the acidosis-induced shortening of the action potential. Acidosis also increased inward current at −80 mV and depolarized the resting membrane potential. Acidosis activated an inwardly rectifying Cl− current that was blocked by 4,4′-diisothiocyanostilbene-2,2′-disulfonic acid (DIDS), which also inhibited the acidosis-induced depolarization of the resting membrane potential. It is concluded that an acidosis-induced increase in steady-state outward K+ current underlies the shortening of the action potential and that an acidosis-induced increase in inwardly rectifying Cl− current underlies the depolarization of the resting membrane potential during acidosis.
Rapid upregulation of endothelial P-selectin expression via reactive oxygen species generation
Authors: Takano Manabu , Meneshian Avedis , Sheikh Emran , Yamakawa Yasuhiko , Wilkins Kirsten Bass , Hopkins Elise A. , Bulkley Gregory B.
Published: March 2015

Endothelial cell ICAM-1 upregulation in response to TNF-α is mediated in part by reactive oxygen species (ROS) generated by the endothelial membrane-associated NADPH oxidase and occurs maximally after 4 h as the synthesis of new protein is required. However, thrombin-stimulated P-selectin upregulation is bimodal, the first peak occurring within minutes. We hypothesize that this early peak, which results from the release of preformed P-selectin from within Weibel-Palade bodies, is mediated in part by ROS generated from the endothelial membrane-associated xanthine oxidase. We found that this rapid expression of P-selectin on the surface of endothelial cells was accompanied by qualitatively parallel increases in ROS generation. Both P-selectin expression and ROS generation were inhibited, dose dependently, by the exogenous administration of disparate cell-permeable antioxidants and also by the inhibition of either of the known membrane-associated ROS-generating enzymes NADPH oxidase or xanthine oxidase. This rapid, posttranslational cell signaling response, mediated by ROS generated not only by the classical NADPH oxidase but also by xanthine oxidase, may well represent an important physiological trigger of the microvascular inflammatory response.
MCSF expression is induced in healing myocardial infarcts and may regulate monocyte and endothelial cell phenotype
Authors: Frangogiannis Nikolaos G. , Mendoza Leonardo H. , Ren Guofeng , Akrivakis Spyridon , Jackson Peggy L. , Michael Lloyd H. , Smith C. Wayne , Entman Mark L.
Published: March 2015

Myocardial infarction is associated with the rapid induction of mononuclear cell chemoattractants that promote monocyte infiltration into the injured area. Monocyte-to-macrophage differentiation and macrophage proliferation allow a long survival of monocytic cells, critical for effective healing of the infarct. In a canine infarction-reperfusion model, newly recruited myeloid leukocytes were markedly augmented during early reperfusion (5–72 h). By 7 days, the number of newly recruited myeloid cells was reduced, and the majority of the inflammatory cells remaining in the infarct were mature macrophages. Macrophage colony-stimulating factor (MCSF) is known to facilitate monocyte survival, monocyte-to-macrophage conversion, and macrophage proliferation. We demonstrated marked induction of MCSF mRNA in ischemic segments persisting for at least 5 days after reperfusion. MCSF expression was predominantly localized to mature macrophages infiltrating the infarcted myocardium; the expression of the MCSF receptor, c-Fms, a protein with tyrosine kinase activity, was found in these macrophages but was also observed in a subset of microvessels within the infarct. Many infarct macrophages expressed proliferating cell nuclear antigen, a marker of proliferative activity. In vitro MCSF induced monocyte chemoattractant protein-1 synthesis in canine venous endothelial cells. MCSF-induced endothelial monocyte chemoattractant protein-1 upregulation was inhibited by herbimycin A, a tyrosine kinase inhibitor, and by LY-294002, a phosphatidylinositol 3′-kinase inhibitor. We suggest that upregulation of MCSF in the infarcted myocardium may have an active role in healing not only through its effects on cells of monocyte/macrophage lineage, but also by regulating endothelial cell chemokine expression.
Sites of action of adenosine in interorgan preconditioning of the heart
Authors: Liem David A. , Verdouw Pieter D. , Ploeg Harald , Kazim Shahla , Duncker Dirk J.
Published: March 2015

The mechanism underlying interorgan preconditioning of the heart remains elusive, although a role for adenosine and activation of a neurogenic pathway has been postulated. We tested in rats the hypothesis that adenosine released by the remote ischemic organ stimulates local afferent nerves, which leads to activation of myocardial adenosine receptors. Preconditioning with a 15-min mesenteric artery occlusion (MAO15) reduced infarct size produced by a 60-min coronary artery occlusion (60-min CAO) from 68 ± 2% to 48 ± 4% ( P < 0.05). Pretreatment with the ganglion blocker hexamethonium or 8-( p-sulfophenyl)theophylline (8-SPT) abolished the protection by MAO15. Intramesenteric artery (but not intraportal vein) infusion of adenosine (10 μg/min) was as cardioprotective as MAO15, which was also abolished by hexamethonium. Whereas administration of hexamethonium at 5 min of reperfusion following MAO15 had no effect, 8-SPT at 5 min of reperfusion abolished the protection. Permanent reocclusion of the mesenteric artery before the 60-min CAO enhanced the cardioprotection by MAO15 (30 ± 5%), but all protection was abolished when 8-SPT was administered after reocclusion of the mesenteric artery. Together, these findings demonstrate the involvement of myocardial adenosine receptors. We therefore conclude that locally released adenosine during small intestinal ischemia stimulates afferent nerves in the mesenteric bed during early reperfusion, initiating a neurogenic pathway that leads to activation of myocardial adenosine receptors.
Glucocorticoids reduce responses to AMPA receptor activation and blockade in nucleus tractus solitarius
Authors: Shank Sylvan S. , Scheuer Deborah A.
Published: March 2015

We tested the hypothesis that glucocorticoids attenuate changes in arterial pressure and renal sympathetic nerve activity (RSNA) in response to activation and blockade of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors within the nucleus of the solitary tract (NTS). Experiments were performed in Inactin-anesthetized male Sprague-Dawley rats treated for 7 ± 1 days with a subcutaneous corticosterone (Cort) pellet or in control rats. Baseline mean arterial pressure (MAP) was significantly higher in Cort-treated rats (109 ± 2 mmHg, n = 39) than in control rats (101 ± 1 mmHg, n = 48, P < 0.05). In control rats, microinjection of AMPA (0.03, 0.1, and 0.3 pmol/100 nl) into the NTS significantly decreased MAP at all doses and decreased RSNA at 0.1 and 0.3 pmol/100 nl. Responses to AMPA in Cort-treated rats were attenuated at all doses of AMPA ( P < 0.05). Responses to the AMPA-kainate receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) were also significantly reduced in Cort-treated rats relative to control rats. Blockade of glucocorticoid type II receptors with mifepristone significantly enhanced responses to CNQX in both control and Cort rats. We conclude that glucocorticoids attenuate MAP and RSNA responses to activation and blockade of AMPA receptors in the NTS.
Parathyroid hormone-related peptide improves contractile function of stunned myocardium in rats and pigs
Authors: Jansen Johanna , Gres Petra , Umschlag Christian , Heinzel Frank R. , Degenhardt Heike , Schlüter Klaus-Dieter , Heusch Gerd , Schulz Rainer
Published: March 2015

The effect of synthetic parathyroid hormone (PTH)-related peptide [PTHrP(1–34)] on regional myocardial function was studied in 11 anesthetized pigs. Intracoronary infusion of PTHrP (cumulative dose: 14 ± 1 μg) decreased coronary resistance to 33 ± 2% of baseline ( P < 0.05) and regional myocardial function to 90 ± 3% of baseline (not significant). Ischemia-reperfusion alters the activity of several kinases and therefore possibly the myocardial effects of PTHrP. In stunned myocardium, induced by 20-min ischemia and 30-min reperfusion, the dose of PTHrP reducing coronary resistance to a minimum of 29 ± 2% was decreased to 8 ± 2 μg ( P < 0.05). Regional myocardial function was no longer decreased but increased to 132 ± 9% ( P < 0.05). The increase in regional myocardial function during PTHrP was inversely related to baseline function at 30-min reperfusion in vivo ( r = 0.9) as well as in myocytes isolated from stunned pig hearts ( r = 0.7). In isolated rat hearts subjected to 30-min global ischemia followed by 30-min reperfusion, blockade of endogenous PTHrP byd-Trp12-Tyr34-PTH(7–34) attenuated the recovery of left ventricular developed pressure by 30 ± 14% ( P < 0.05). Thus endogenous and exogenous PTHrP impact on the function of stunned myocardium.