bluuebunny/crossref_metadata_2025 huggingface dataset:
Role of MMPs and plasminogen activators in angiogenesis after transmyocardial laser revascularization in dogs
Authors: Li Wei , Tanaka Kuniyoshi , Chiba Yukio , Kimura Tetsuya , Morioka Kouichi , Uesaka Takahiko , Ihaya Akio , Sasaki Masato , Tsuda Takeshi , Yamada Narihisa
Published: March 2015

We examined the role of matrix metalloproteinases (MMPs), tissue inhibitors of MMP (TIMPs), and plasminogen activator (PA) in transmyocardial laser revascularization (TMLR)-induced angiogenesis. TMLR was accomplished with a carbon dioxide laser in seven dogs whose left anterior descending coronary artery (LAD) was ligated. Seven control dogs underwent only LAD ligation, and four dogs underwent a sham operation, consisting only of a left thoracotomy. Two weeks later, transmural myocardial samples were harvested from the distributions of the LAD and the left circumflex artery for substrate zymography, immunohistochemical staining, and in situ zymography. MMP-1, MMP-2, TIMP-1, TIMP-2, and urokinase-type PA levels in the distribution of the LAD were higher in the laser group than in the control or sham group. Counts of von Willebrand factor-positive microvessels and smooth muscle α-actin-positive arterioles demonstrated that the angiogenesis and ateriogenesis was promoted in the laser group and correlated directly with the number of MMP-stained microvessels. We conclude that TMLR induces the expression of MMPs, TIMPs, and urokinase-type PA and that these proteinases play an important role in angiogenesis after TMLR.
Attenuation of activity-induced increases in cerebellar blood flow by lesion of the inferior olive
Authors: Zhang Yi , Forster Colleen , Milner Teresa A. , Iadecola Costantino
Published: March 2015

We sought to define the contribution of the climbing fibers (CF), one of the major inputs to Purkinje neurons, to the increase in cerebellar blood flow (BFcrb) produced by activation of the cerebellar cortex. The neurotoxin 3-acetylpyridine was used to lesion the inferior olive, the site from which the CF originate. Crus II, an area of the cerebellar cortex that receives sensory afferents from the perioral region, was activated by low-intensity stimulation of the upper lip (5–25 V and 4–16 Hz) in sham-lesioned and lesioned mice. BFcrb was recorded in crus II using a laser-Doppler flow probe. The increase in BFcrb produced by harmaline, an alkaloid that activates the CF, was abolished in lesioned mice ( P > 0.05 vs. BFcrb before harmaline, n = 6), attesting to the effectiveness of the lesion. In sham-lesioned animals, upper lip stimulation increased BFcrb in crus II by 25 ± 2% (25 V and 10 Hz, n = 6). The rise in BFcrb was attenuated by 63 ± 7% (25 V and 10 Hz) in lesioned mice ( P < 0.05, n = 6). In contrast, the increase in BFcrb produced by hypercapnia was not affected ( P > 0.05). These data suggest that CF are responsible for a substantial portion of the increase in BFcrb produced by crus II activation. Thus the hemodynamic response evoked by functional activation of the cerebellar cortex reflects, in large part, CF activity.
Regulation of KATPchannels by P2Ypurinoceptors coupled to PIP2metabolism in guinea pig ventricular cells
Authors: Oketani Naoya , Kakei Masafumi , Ichinari Kotaro , Okamura Midori , Miyamura Akihiro , Nakazaki Mitsuhiro , Ito Seiki , Tei Chuwa
Published: March 2015

We used patch-clamp techniques to elucidate the regulatory mechanisms of ATP-sensitive K+(KATP) channels by stimulation of P2purinoceptors in guinea pig ventricular myocytes. Extracellular ATP at 0.1 mM transiently inhibited by 90.5 ± 5.0% the whole cell KATPchannel current evoked by a reduction in intracellular ATP concentration to 0.5 mM and exposure to 30 μM pinacidil. ADP and AMP (both 1 mM) also decreased the current by 42.8 ± 9.3% and 9.4 ± 4.8%, respectively, but adenosine did not, even at 10 mM. ATP-induced channel inhibition was hardly observed in the presence of 0.2 mM suramin, 0.2 mM guanosine 5'- O-(2-thiodiphosphate), or 0.1 mM compound 48/80, whereas it was not influenced by the presence of 0.1 μM staurosporine or 10 mM 1,2-bis(2-aminophenoxy)ethane- N,N,N′,N′-tetraacetic acid in the pipette. In the presence of 10 μM wortmannin or the absence of ATP in the cytosol, the ATP-induced channel inhibition was irreversible. Phosphatidylinositol 4,5-bisphosphate (PIP2) at 0.1 mM in the outside-out patch pipette prevented ATP-induced channel inhibition. The half-maximal internal ATP concentrations for inhibition of channel activity determined in inside-out membrane patches were 13.8 μM in the presence and 1.12 mM in the absence of 0.1 mM ATP at the external side. It is concluded that activity of KATPchannels is modulated by extracellular ATP by a mechanism involving P2Ypurinoceptors coupled to GTP-binding proteins associated with reduction of the sarcolemmal PIP2concentration via stimulation of phospholipase C.
Enhanced [Ca2+]iin renal arterial smooth muscle cells of pregnant rats with reduced uterine perfusion pressure
Authors: Murphy Jason G. , Herrington Jason N. , Granger Joey P. , Khalil Raouf A.
Published: March 2015

Reduction of uterine perfusion pressure (RUPP) during late pregnancy has been suggested to trigger increases in renal vascular resistance and lead to hypertension of pregnancy. We investigated whether the increased renal vascular resistance associated with RUPP in late pregnancy reflects increases in intracellular Ca2+concentration ([Ca2+]i) and contraction of renal arterial smooth muscle. Single smooth muscle cells were isolated from renal interlobular arteries of normal pregnant Sprague-Dawley rats and a rat model of RUPP during late pregnancy. The cells were loaded with fura 2 and both cell length and [Ca2+]iwere measured. In cells of normal pregnant rats incubated in Hanks' solution (1 mM Ca2+), ANG II (10−7M) caused an initial increase in [Ca2+]ito 414 ± 13 nM, a maintained increase to 149 ± 8 nM, and 21 ± 1% cell contraction. In RUPP rats, the initial ANG II-induced [Ca2+]i(431 ± 18 nM) was not different from pregnant rats, but both the maintained [Ca2+]i(225 ± 9 nM) and cell contraction (48 ± 2%) were increased. Membrane depolarization by 51 mM KCl and the Ca2+channel agonist BAY K 8644 (10−6M), which stimulate Ca2+entry from the extracellular space, caused maintained increases in [Ca2+]iand cell contraction that were greater in RUPP rats than control pregnant rats. In Ca2+-free (2 mM EGTA) Hanks' solution, the ANG II- and caffeine (10 mM)-induced [Ca2+]itransient and cell contraction were not different between normal pregnant and RUPP rats, suggesting no difference in Ca2+release from the intracellular stores. The enhanced maintained ANG II-, KCl- and BAY K 8644-induced [Ca2+]iand cell contraction in RUPP rats compared with normal pregnant rats suggest enhanced Ca2+entry mechanisms of smooth muscle contraction in resistance renal arteries and may explain the increased renal vascular resistance associated with hypertension of pregnancy.
TENS attenuates response to colon distension in paraplegic and quadriplegic rats
Authors: Collins Heidi L. , DiCarlo Stephen E.
Published: March 2015

Individuals with spinal cord injuries above thoracic level 6 experience episodic bouts of life-threatening hypertension as part of a condition termed autonomic dysreflexia (AD). The hypertension can be caused by stimulation of the skin, distension of the urinary bladder or colon, and/or muscle spasms. Transcutaneous electrical nerve stimulation (TENS) may reduce the severity of AD because TENS has been used to inhibit second-order neurons in the dorsal horn. Therefore, we tested the hypothesis that TENS attenuates the hemodynamic responses to colon distension. Eleven Wistar rats underwent spinal cord transection between thoracic vertebrae 4 and 5 (paraplegic, n = 6) or between cervical vertebra 7 and thoracic vertebra 1 (quadriplegic, n = 5). After recovery, all rats were instrumented with a radiotelemetry device for recording arterial pressure. Subsequently, the hemodynamic responses to graded colon distension were determined before and during TENS. During TENS the hemodynamic responses to colon distension were significantly attenuated. Thus TENS may be a preventive approach to reduce the severity of AD in paraplegic and quadriplegic individuals.
Activation time of myocardial oxidative phosphorylation in creatine kinase and adenylate kinase knockout mice
Authors: Gustafson Lori A. , Van Beek Johannes H. G. M.
Published: March 2015

Our goal was to determine whether mice genetically altered to lack either creatine kinase (M/MtCK−/−) or adenylate kinase (AK−/−) show altered properties in the dynamic regulation of myocardial oxygen consumption (MV˙o 2). We measured contractile function, oxygen consumption, and the mean response time of oxygen consumption to a step increase in heart rate [i.e., mitochondrial response time ( t mito)] in isolated Langendorff-perfused hearts from wild-type ( n = 6), M/MtCK−/− ( n = 6), and AK−/− ( n = 4) mice. Left ventricular developed pressure was higher in M/MtCK−/− hearts (88.2 ± 6.8 mmHg) and lower in AK−/− hearts (46.7 ± 9.4 mmHg) compared with wild-type hearts (60.7 ± 10.1 mmHg) at the basal pacing rate. Developed pressure fell slightly when heart rate was increased in all three groups. Basal MV˙o 2 at 300 beats/min was 19.1 ± 2.4, 19.4 ± 1.5, and 16.3 ± 1.9 μmol · min−1 · g dry wt−1for M/MtCK−/−, AK−/−, and wild type, respectively, which increased to 25.5 ± 3.7, 25.4 ± 2.6, and 22.0 ± 2.6 μmol · min−1 · g−1, when heart rate was increased to 400 beats/min. The t mito was significantly faster in M/MtCK−/− hearts: 3.0 ± 0.3 versus 7.3 ± 0.6 and 8.0 ± 0.4 s for M/MtCK−/−, AK−/−, and wild-type hearts, respectively. Our results demonstrate that MV˙o 2 of M/MtCK−/− hearts adapts more quickly to an increase in heart rate and thereby support the hypothesis that creatine kinase acts as an energy buffer in the cytosol, which delays the energy-related signal between sites of ATP hydrolysis and mitochondria.
Improvement in diastolic intraventricular pressure gradients in patients with HOCM after ethanol septal reduction
Authors: Rovner Aleksandr , Smith Rebecca , Greenberg Neil L. , Tuzcu E. Murat , Smedira Nicholas , Lever Harry M. , Thomas James D. , Garcia Mario J.
Published: March 2015

We sought to validate measurement of intraventricular pressure gradients (IVPG) and analyze their change in patients with hypertrophic obstructive cardiomyopathy (HOCM) after ethanol septal reduction (ESR). Quantitative analysis of color M-mode Doppler (CMM) images may be used to estimate diastolic IVPG noninvasively. Noninvasive IVPG measurement was validated in 10 patients undergoing surgical myectomy. Echocardiograms were then analyzed in 19 patients at baseline and after ESR. Pulsed Doppler data through the mitral valve and pulmonary venous flow were obtained. CMM was used to obtain the flow propagation velocity ( Vp) and to calculate IVPG off-line. Left atrial pressure was estimated with the use of previously validated Doppler equations. Data were compared before and after ESR. CMM-derived IVPG correlated well with invasive measurements obtained before and after surgical myectomy [ r = 0.8, P < 0.01, Δ(CMM – invasive IVPG) = 0.09 ± 0.45 mmHg]. ESR resulted in a decrease of resting LVOT systolic gradient from 62 ± 10 to 29 ± 5 mmHg ( P < 0.001). There was a significant increase in the Vp and IVPG (from 48 ± 5to 74 ± 7 cm/s and from 1.5 ± 0.2 to 2.6 ± 0.3 mmHg, respectively, P < 0.001 for both). Estimated left atrial pressure decreased from 16.2 ± 1.1 to 11.5 ± 0.9 mmHg ( P < 0.001). The increase in IVPG correlated with the reduction in the LVOT gradient ( r = 0.6, P < 0.01). Reduction of LVOT obstruction after ESR is associated with an improvement in diastolic suction force. Noninvasive measurements of IVPG may be used as an indicator of diastolic function improvement in HOCM.
Decreased Smad 7 expression contributes to cardiac fibrosis in the infarcted rat heart
Authors: Wang Baiqiu , Hao Jianming , Jones Stephen C. , Yee May-Sann , Roth Julie C. , Dixon Ian M. C.
Published: March 2015

We examined the role of the transforming growth factor (TGF)-β1signaling inhibitor Smad 7 in cardiac fibrosis. TGF-β1(10 ng/ml) was found to increase cytosolic Smad 7 expression in primary adult rat fibroblasts and induce rapid nuclear export of exogenous Smad 7 in COS-7 cells. Furthermore, overexpression of Smad 7 in primary adult fibroblasts was associated with suppressed collagen type I and III expression. We detected Smad 7, phosphorylated Smad 2, TGF-β type I receptor (TβRI), and TGF-β1proteins in postmyocardial infarct (MI) rat hearts. In 2 and 4 wk post-MI hearts, Smad 7 and TβRI expression were decreased in scar tissue, whereas TGF-β1expression was increased in scar and viable tissue. In the 8 wk post-MI heart, Smad 7 expression was decreased in both scar tissue and myocardium remote to the infarct scar. Finally, we confirmed that these changes are paralleled by decreased expression of cytosolic phosphorylated receptor-regulated Smad 2 in 4-wk viable myocardium and in 2- and 4-wk infarct scar tissues. Taken together, our data imply that decreased inhibitory Smad 7 signal in cardiac fibroblasts may play a role in the pathogenesis of cardiac fibrosis in the post-MI heart.
Reduced Ca2+-calmodulin-dependent protein kinase activity and expression in LV myocardium of dogs with heart failure
Authors: Mishra Sudhish , Sabbah Hani N. , Jain Jinesh C. , Gupta Ramesh C.
Published: March 2015

Studies on the status of multifunctional Ca2+-calmodulin (CaM)-dependent protein kinase-II (CaMKII) in failing hearts are limited and controversial. The study was performed in the left ventricular (LV) myocardium of six dogs with heart failure (HF) (LV ejection fraction, 23 ± 2%) and six normal (NL) dogs. In the LV homogenate, CaMKII activity and its protein level were determined by using the CaMKII peptide and antibody, respectively. Furthermore, the protein level of CaM and phosphorylated phospholamban (PLB) at threonine-17 (PLB-Thr17) and serine-16 (PLB-Ser16) were also determined in the LV homogenate using a specific antibody. In addition, the level of zinc, which inhibits protein kinase A activity, was determined in the LV tissue by inductively coupled plasma mass spectrometry. CaMKII activity and phosphorylated PLB-Thr17 and PLB-Ser16 levels, but not CaM and Zn levels, were significantly reduced in the LV homogenate of dogs with HF compared with NL dogs. These results suggest that CaMKII activity is reduced in the failing LV myocardium, and this abnormality is associated with reduced protein expression level of the enzyme but not due to changes in CaM and zinc levels. In conclusion, reduced CaMKII activity and phosphorylated PLB level may be partly responsible for impaired sarcoplasmic reticulum function in HF.
Extent of damage in ischemic, nonreperfused, and reperfused myocardium of anesthetized rats
Authors: Vetterlein Friedrich , Schrader Christina , Volkmann Rolf , Neckel Marion , Ochs Matthias , Schmidt Gerhard , Hellige Gerhard
Published: March 2015

To investigate the localization of the earliest damage in ischemic and ischemic-reperfused myocardium, anesthetized rats were subjected to coronary occlusion for 15, 30, 45, or 90 min. One-half of the animals in each group had no reperfusion, whereas the other half was reperfused for 14 min. With the use of histological methods, preferentially in the periphery of the area at risk, localized zones were detected that lacked the hypoxia-specific increase in NADH fluorescence. The extent of these areas displaying injured tissue was found to be significantly smaller in the ischemic-nonreperfused hearts than in the ischemic-reperfused organs (15-min ischemia: 0.22 ± 0.12% vs. 43.0 ± 5.0%; 30-min ischemia: 5.7 ± 2.7% vs. 64.6 ± 2.9%; 45-min ischemia: 5.6 ± 1.2% vs. 66.0 ± 7.5%; 90-min ischemia: 39.3 ± 5.5% vs. 86.7 ± 1.8% of the area at risk). The results point to a localized initiation of the damage close to the surrounding oxygen-supplied tissue during ischemia and an expansion of this injury by intercellular actions into yet-intact areas upon reperfusion.
Cardiac enkephalins attenuate vagal bradycardia: interactions with NOS-1-cGMP systems in canine sinoatrial node
Authors: Farias Martin , Jackson Keith , Johnson Michael , Caffrey James L.
Published: March 2015

Endogenous opioids and nitric oxide (NO) are recognized modulators of cardiac function. Enkephalins and inhibitors of NO synthase (NOS) both produce similar interruptions in the vagal control of heart rate. This study was conducted to test the hypothesis that NO systems within the canine sinoatrial (SA) node facilitate local vagal transmission and that the endogenous enkephalin methionine-enkephalin-arginine-phenylalanine (MEAP) attenuates vagal bradycardia by interrupting the NOS-cGMP pathway. Microdialysis probes were inserted into the SA node, and they were perfused with nonselective ( Nω-nitro-l-arginine methyl ester) and neuronal (7-nitroindazole) NOS inhibitors. The right vagus nerve was stimulated and both inhibitors gradually attenuated the resulting vagal bradycardia. The specificity of these inhibitions was verified by an equally gradual reversal of the inhibition with an excess of the NOS substrate l-arginine. Introduction of MEAP into the nodal interstitium produced a quickly developing but quantitatively similar interruption of vagal bradycardia that was also slowly reversed by the addition of l-arginine and not by d-arginine. Additional support for convergence of opioid and NO pathways was provided when the vagolytic effects of MEAP were also reversed by the addition of the NO donor S-nitroso- N-acetyl-penicillamine, the protein kinase G activator 8-bromo-cGMP, or the phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine. MEAP and 7-nitroindazole were individually combined with the direct acting muscarinic agonist methacholine to evaluate potential interactions with muscarinic receptors within the SA node. MEAP and 7-nitroindazole were unable to overcome the bradycardia produced by methacholine. These data suggest that NO and enkephalins moderate the vagal control of heart rate via interaction with converging systems that involve the regulation of cAMP within nodal parasympathetic nerve terminals.
Cardiac retention of [11C]HED in genotyped long QT patients: a potential amplifier role for severity of the disease
Authors: Mazzadi Alejandro N. , André-Fouët Xavier , Duisit Jérôme , Gebuhrer Véronique , Costes Nicolas , Chevalier Philippe , Rodriguez Claire , Schott Jean-Jacques , Le Marec Hervé , Guicheney Pascale , Le Bars Didier , Janier Marc
Published: March 2015

Although mutations in cardiac sodium and potassium channel genes are associated with congenital long QT syndrome (LQTS), a “modifier” role of the sympathetic nervous system was proposed to explain the distinct severity of the disease. We evaluated cardiac sympathetic innervation using [11C]hydroxyephedrine ([11C]HED) and positron emission tomography (PET) in genotyped LQTS patients. H215O and [11C]HED PET studies were performed in 11 patients (5 symptomatic) and 8 controls. Perfusion and [11C]HED images were depicted as 36-sector polar maps. Sectorial values of perfusion (H2O%), absolute (HEDRet) and relative retention (HED%Ret) of [11C]HED, and the ratio of HED%Ret to H2O% (HED%Ret/H2O%) were calculated. Normal databases were obtained from controls. Sectorial values below 2SD database values were defined as “outside sectors.” Controls and patients showed similar sectorial perfusion. Sectorial HEDRet did not differ between groups, but means of HED%Ret were lower in three sectors for patients ( P < 0.05). Three sectors from 3 controls had HED%Ret below 2SD, whereas 36 sectors in 9 patients were outside sectors ( P < 0.01). In patients, average HED%Ret/H2O% was lower in 9 sectors ( P < 0.05 vs. controls); 2 outside sectors were found in controls, but 43 outside sectors were found in patients ( P < 0.01), 77% of them in the 5 symptomatic patients. Heterogeneous [11C]HED retention was localized in the septal, anterior, and lateral walls. Most LQTS patients showed a localized and decreased pattern of [11C]HED retention. The larger number of heterogeneous sectors in symptomatic patients suggests that sympathetic function could play an amplifier role for severity of the disease.
Intracellular proton mobility and buffering power in cardiac ventricular myocytes from rat, rabbit, and guinea pig
Authors: Zaniboni Massimiliano , Swietach Pawel , Rossini Alessandra , Yamamoto Taku , Spitzer Kenneth W. , Vaughan-Jones Richard D.
Published: March 2015

Intracellular pH (pHi) is an important modulator of cardiac function. The spatial regulation of pH within the cytoplasm depends, in part, on intracellular H+ ([Formula: see text]) mobility. The apparent diffusion coefficient for [Formula: see text], was estimated in single ventricular myocytes isolated from the rat, guinea pig, and rabbit. [Formula: see text] was derived by best-fitting predictions of a two-dimensional model of H+ diffusion to the local rise of intracellular [H+], recorded confocally (ratiometric seminaphthorhodafluor fluorescence) downstream from an acid-filled, whole cell patch pipette. Under [Formula: see text] conditions, [Formula: see text] was similar in all three species (mean values: 8–12.5 × 10–7 cm2/s) and was over 200-fold lower than that for H+ in water. In guinea pig myocytes, [Formula: see text] was increased 2.5-fold in the presence of [Formula: see text] buffer, in agreement with previous observations in rabbit myocytes. [Formula: see text] mobility is therefore low in cardiac cells, a feature that may predispose them to the generation of pHi gradients in response to sarcolemmal acid/base transport or local cytoplasmic acid production. Low [Formula: see text] mobility most likely results from H+ shuttling among cytoplasmic mobile and fixed buffers. This hypothesis was explored by comparing the pHi dependence of intrinsic, intracellular buffering capacity, measured for all three species, and subdividing buffering into mobile and fixed fractions. The proportion of buffer that is mobile will be the main determinant of [Formula: see text]. At a given pHi, this proportion appeared to be similar in all three species, consistent with a common value for [Formula: see text]. Over the pHi range of 6.0–8.0, the proportion is expected to change, predicting that [Formula: see text] may display some pHi sensitivity.
Reversibility of electrophysiological changes induced by chronic high-altitude hypoxia in adult rat heart
Authors: Chouabe C. , Amsellem J. , Espinosa L. , Ribaux P. , Blaineau S. , Mégas P. , Bonvallet R.
Published: March 2015

Recent studies indicate that regression of left ventricular hypertrophy normalizes membrane ionic current abnormalities. This work was designed to determine whether regression of right ventricular hypertrophy induced by permanent high-altitude exposure (4,500 m, 20 days) in adult rats also normalizes changes of ventricular myocyte electrophysiology. According to the current data, prolonged action potential, decreased transient outward current density, and increased inward sodium/calcium exchange current density normalized 20 days after the end of altitude exposure, whereas right ventricular hypertrophy evidenced by both the right ventricular weight-to-heart weight ratio and the right ventricular free wall thickness measurement normalized 40 days after the end of altitude exposure. This morphological normalization occurred at both the level of muscular tissue, as shown by the decrease toward control values of some myocyte parameters (perimeter, capacitance, and width), and the level of the interstitial collagenous connective tissue. In the chronic high-altitude hypoxia model, the regression of right ventricular hypertrophy would not be a prerequisite for normalization of ventricular electrophysiological abnormalities.
Mechanisms underlying ischemic diastolic dysfunction: relation between rigor, calcium homeostasis, and relaxation rate
Authors: Varma Niraj , Morgan James P. , Apstein Carl S.
Published: March 2015

Increased diastolic chamber stiffness (↑DCS) during ischemia may result from increased diastolic calcium, rigor, or reduced velocity of relaxation. We tested these potential mechanisms during severe ischemia in isolated red blood cell-perfused isovolumic rabbit hearts. Ischemia (coronary flow reduced 83%) reduced left ventricular (LV) contractility by 70%, which then remained stable. DCS progressively increased. When LV end-diastolic pressure had increased 5 mmHg, myofilament calcium responsiveness was altered with 50 mmol/l NH4Cl or 10 mmol/l butanedione monoxime. These affected contractility (i.e., a calcium-mediated force) but not ↑DCS. Second, quick length changes reversed ↑DCS, supporting a rigor mechanism. Third, ischemia increased the time constant of isovolumic pressure decline from 47 ± 3 to 58 ± 3 ms ( P < 0.02) but concomitantly abbreviated the contraction-relaxation cycle, i.e., pressure dissipation occurred earlier without diastolic tetanization. Finally, to assess any link between rate of relaxation and ↑DCS, hearts were exposed to 10 mmol/l calcium. Calcium doubled contractility and accelerated relaxation velocity, but without affecting ↑DCS. Thus ↑DCS developed during ischemia despite severely reduced contractility via a rigor (and not calcium mediated) mechanism. Calcium resequestration capacity was preserved, and reduced relaxation velocity was not linked to ↑DCS.
Chronic shear induces caveolae formation and alters ERK and Akt responses in endothelial cells
Authors: Boyd Nolan L. , Park Heonyong , Yi Hong , Boo Yong Chool , Sorescu George P. , Sykes Michelle , Jo Hanjoong
Published: March 2015

Caveolae are plasmalemmal domains enriched with cholesterol, caveolins, and signaling molecules. Endothelial cells in vivo are continuously exposed to shear conditions, and their caveolae density and location may be different from that of static cultured cells. Here, we show that chronic shear exposure regulates formation and localization of caveolae and caveolin-1 in bovine aortic endothelial cells (BAEC). Chronic exposure (1 or 3 days) of BAEC to laminar shear increased the total number of caveolae by 45–48% above static control. This increase was due to a rise in the luminal caveolae density without changing abluminal caveolae numbers or increasing caveolin-1 mRNA and protein levels. Whereas some caveolin-1 was found in the plasma membrane in static-cultured cells, it was predominantly localized in the Golgi. In contrast, chronic shear-exposed cells showed intense caveolin-1 staining in the luminal plasma membrane with minimum Golgi association. The preferential luminal localization of caveolae may play an important role in endothelial mechanosensing. Indeed, we found that chronic shear exposure (preconditioning) altered activation patterns of two well-known shear-sensitive signaling molecules (ERK and Akt) in response to a step increase in shear stress. ERK activation was blunted in shear preconditioned cells, whereas the Akt response was accelerated. These results suggest that chronic shear stimulates caveolae formation by translocating caveolin-1 from the Golgi to the luminal plasma membrane and alters cell signaling responses.
Regulation of SM22α expression by arginine vasopressin and PDGF-BB in vascular smooth muscle cells
Authors: Kaplan-Albuquerque Nihal , Garat Chrystelle , Van Putten Vicki , Nemenoff Raphael A.
Published: March 2015

Vascular smooth muscle (SM) cells (VSMC) undergo phenotypic modulation in vivo and in vitro. This process involves coordinated changes in expression of multiple SM-specific genes. In cultured VSMC, arginine vasopressin (AVP) increases and PDGF decreases expression of SM α-actin (SMA), the earliest marker of SM cells (SMC). However, it is unknown whether these agents regulate other SM genes in a similar fashion. SM22α appears secondary to SMA during development and is also a marker for SMC. This study examined the regulation of SM22α expression by AVP and PDGF in cultured VSMC. Levels of SM22α mRNA and protein were increased by AVP and suppressed by PDGF. Consistent with these changes, AVP increased SM22α promoter activity, whereas PDGF inhibited basal promoter activity and blocked AVP-induced increase. Activation of both JNK and p38 MAPK pathways was necessary for AVP-mediated induction of SM22α promoter. Expression of constitutively active Ras produced similar suppressions on SM22α promoter activity as PDGF. Signaling relayed from PDGF/Ras activation involved Raf, or a protein that competes for this site, Ral-GDS, and phosphatidylinositol 3-kinase activation. Truncational analysis showed that the proximal location of three CArG boxes in the promoter was sufficient for AVP stimulation. Mutations in this CArG box reduced basal and AVP-stimulated promoter activity without effecting PDGF suppression. Overexpression of serum response factor enhanced basal and AVP-stimulated promoter activity but had no effect on PDGF-BB-induced suppression. These data indicate that AVP and PDGF initiate specific signaling pathways that control expression of multiple SM genes leading to phenotypic modulation.
Premature excitation and onset of reentrant ventricular tachycardia
Authors: Ciaccio Edward J.
Published: March 2015

It was hypothesized that quantitative sinus rhythm electrogram measurements could be used to predict conduction events that result from premature stimulation and reentrant ventricular tachycardia inducibility. Sinus rhythm activation and electrogram-duration maps were constructed from bipolar electrograms acquired at 196–312 sites in the epicardial border zone of 43 canine hearts (25 with and 18 without reentrant ventricular tachycardia inducible by premature stimulation). From these maps, lines of electrical discontinuity, where blocks would occur during premature excitation, were estimated. The mean error in distance between the estimated and actual block lines of premature excitation was 0.97 ± 0.49 cm. Based on the quantitative characteristics of the activation and electrogram-duration maps and the longest block line that formed during premature excitation, it was possible to predict whether reentry would occur (sensitivity, 94.7%; specificity, 79.6%). In reentry experiments, the breakthrough-point location along the unidirectional arc of the block that initiated reentry was also predictable (mean error, 0.79 ± 0.19 cm). Sinus rhythm measurements are useful to predict conduction events that result from premature stimulation and reentry inducibility.
Superoxide mediates endotoxin-induced platelet-endothelial cell adhesion in intestinal venules
Authors: Cerwinka Wolfgang H. , Cooper Dianne , Krieglstein Christian F. , Ross Chris R. , McCord Joe M. , Granger D. Neil
Published: March 2015

Platelets have been implicated in the pathogenesis of different diseases of the vascular system, including atherosclerosis, sepsis, and ischemia-reperfusion injury; however, relatively little is known about the factors that regulate the interactions between circulating platelets and the vessel wall. The objective of this study was to define the contribution of superoxide to LPS-induced platelet-endothelial cell (P/E) adhesion in murine intestinal venules. The adhesion of rhodamine-6G-labeled murine platelets was monitored by intravital fluorescence microscopy. Four hours after LPS administration in control [wild-type (WT)] mice, an ∼10-fold increase in P/E adhesion was detected. This response did not result from LPS-induced platelet activation. The LPS-induced P/E adhesion was greatly attenuated in NAD(P)H oxidase-deficient mice and in WT mice rendered neutropenic with anti-neutrophil serum, whereas the response was unchanged in WT mice receiving a CD18 blocking MAb or in CD18-deficient mice. A chimeric form of MnSOD that exhibits the binding properties of extracellular SOD also attenuated the LPS-induced response in WT mice. These findings indicate that neutrophil-derived superoxide plays a major role in the modulation of endotoxin-induced P/E adhesion.
Alterations in nitric oxide-cGMP pathway in ventricular myocytes from obese leptin-deficient mice
Authors: Su Jun , Zhang Shengjun , Tse James , Scholz Peter M. , Weiss Harvey R.
Published: March 2015

Leptin is a regulator of body weight and affects nitric oxide (NO) production. This study was designed to determine whether the myocardial NO-cGMP signal transduction system was altered in leptin-deficient obese mice. Contractile function, guanylyl cyclase activity, and cGMP-dependent protein phosphorylation were assessed in ventricular myocytes isolated from genetically obese (B6.V-Lepob) and age-matched lean (C57BL/6J) mice. There were no differences in baseline contraction between the lean and obese groups. After stimulation with the NO donor S-nitroso- N-acetyl-penicillamine (SNAP, 10–6and 10–5M) or a membrane-permeable cGMP analog 8-bromo-cGMP (8-Br-cGMP, 10–6and 10–5M), cell contractility was depressed. However, 8-Br-cGMP had significantly greater effects in obese mice than in lean controls with percent shortening reduced by 47 vs. 39% and maximal rate of shortening decreased by 46 vs. 36%. The negative effects of SNAP were similar between the two groups. Soluble guanylyl cyclase activity was not attenuated. This suggests that the activity of the cGMP-independent NO pathway may be enhanced in obesity. The phosphorylated protein profile of cGMP-dependent protein kinase showed that four proteins were more intensively phosphorylated in obese mice, which suggests an explanation for the enhanced effect of cGMP. These results indicate that the NO-cGMP signaling pathway was significantly altered in ventricular myocytes from the leptin-deficient obese mouse model.
Increased susceptibility to ventricular arrhythmias is associated with changes in Ca2+ regulatory proteins in paraplegic rats
Authors: Rodenbaugh David W. , Collins Heidi L. , Nowacek Dustin G. , DiCarlo Stephen E.
Published: March 2015

Paraplegia may increase susceptibility to ventricular arrhythmias by altering the autonomic control of the heart. Altered cardiac autonomic control has been documented to change the expression of genes that encode cardiac Ca2+ regulatory proteins. Therefore, we tested the hypothesis that paraplegia alters cardiac electrophysiology with concomitant changes in Ca2+ regulatory proteins in a manner that increases the susceptibility to ventricular arrhythmias. To test this hypothesis, intact ( n = 10) and paraplegic ( n = 6) male Wistar rats were chronically instrumented to measure atrioventricular (AV) interval, sinus cycle length, sinus node recovery time (SNRT), SNRT corrected for spontaneous sinus cycle (cSNRT), Wenckebach cycle length (WCL), and the electrical stimulation threshold to induce ventricular arrhythmias. In addition, relative protein abundance and mRNA expression for sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA), phospholamban, and the Na/Ca exchanger were determined in intact ( n = 8) and paraplegic ( n = 8) rats. Paraplegia significantly ( P < 0.05) reduced AV interval (–25%), sinus cycle length (–24%), SNRT (–28%), cSNRT (–53%), WCL (–19%), and the electrical stimulation threshold to induce ventricular arrhythmia (–48%). Paraplegia significantly increased the relative protein abundances of SERCA (45%) and the Na/Ca exchanger (40%) and decreased phospholamban levels (–28%). In contrast, only the relative mRNA expression of the Na/Ca exchanger was increased (25%) in paraplegic rats. These data demonstrate that paraplegia enhances cardiac electrophysiological properties and alters Ca2+ regulatory proteins in a manner that increases susceptibility to ventricular arrhythmias.
Coronary flow-induced inotropism is modulated by binding of dextrans to the endothelial luminal surface
Authors: Gonzalez-Castillo Carmen , Rubio Rafael , Zenteno-Savin Tania
Published: March 2015

In isolated perfused guinea pig hearts, coronary flow causes a positive inotropic effect [positive coronary flow-induced effect (+CFIE)] that could be altered by dextrans (Dx) in the coronary perfusion solution. To test this possibility, Dx of 20, 40, 70, and 500 kDa were infused and found to modulate +CFIE; however, when Dx infusion was terminated, the effect persisted, i.e., was irreversible/nonwashable, suggesting that Dx may bind to luminal endothelial lectinic structures. This hypothesis was tested when Dx [with fluorescent traces (D*)] bound to the vessel wall was hydrolyzed by dextranase infusion and washout of D* fragments completely reverted the +CFIE, and it was found that bound D* to be displaced by free Dx required concentrations 50–100 times that used during binding. In addition, dose-response curves for Dx on +CFIE show that the higher the Dx molecular mass, the lesser the concentration required to have an effect. Because a large Dx molecule has a greater number polymeric glucose branches, it can bind to a larger number of endothelial lectinic sites, requiring a lower concentration to affect +CFIE. Our results suggest that luminal endothelial lectinic structures are part of the flow-sensing assembly.
Evidence for fractal correlation properties in variations of peripheral arterial tone during REM sleep
Authors: Dvir I. , Adler Y. , Freimark D. , Lavie P.
Published: March 2015

Previous studies utilizing detrended fluctuation analysis (DFA) of heart rate variability during sleep revealed a higher fractal exponent during rapid eye movement (REM) sleep than non-REM sleep. The aim of this study was to determine whether the same difference exists in the variations of peripheral arterial tone (PAT). Finger pulse wave measured by a novel plethysmographic technique was monitored during sleep in 12 chronic heart failure patients, 8 heavy snorers, and 12 healthy volunteers. For each subject, at least two 15-min time series were constructed from the interpulse intervals and from pulse wave amplitudes during REM and non-REM sleep. Fractal scaling exponents of both types of time series were significantly higher for REM than non-REM sleep in all groups. In each of the groups and in both sleep stages, the fractal scaling exponents based on pulse wave amplitude were significantly higher than those based on pulse rate variability. A repeat of the analysis for short-, intermediate-, and long-term intervals revealed that the fractallike exponents were evident only in the short- and intermediate-term intervals. Because PAT is a surrogate of sympathetic activation, our results indicate that variations in sympathetic activation during REM sleep have a fractallike behavior.
Characterization of Ca2+ channels involved in ET-1-induced transactivation of EGF receptors
Authors: Kawanabe Yoshifumi , Hashimoto Nobuo , Masaki Tomoh
Published: March 2015

The purpose of this study was to demonstrate the involvement of Ca2+ influx through voltage-independent Ca2+ channels (VICCs) in endothelin-1 (ET-1)-induced transactivation of epidermal growth factor receptor protein tyrosine kinase (EGFR PTK) using the Ca2+ channel blockers LOE-908 and SK&F-96365 in rabbit internal carotid artery vascular smooth muscle cells. ET-1-induced EGFR PTK transactivation was completely inhibited by AG-1478, which is a specific inhibitor of EGFR PTK. In the absence of extracellular Ca2+, the magnitude of EGFR PTK transactivation was near the basal level. Based on sensitivity to nifedipine, which is a specific blocker of voltage-operated Ca2+ channels (VOCCs), VOCCs have minor roles in EGFR PTK transactivation. In contrast, Ca2+ influx through VICCs plays an important role in EGFR PTK transactivation. Moreover, based on the sensitivity of VICCs to SK&F-96365 and LOE-908, VICCs were shown to consist of two types of Ca2+-permeable nonselective cation channels (NSCCs), which are designated NSCC-1 and NSCC-2, and a store-operated Ca2+ channel. In summary, Ca2+influx through VICCs plays an essential role in ET-1-induced EGFR PTK transactivation in rabbit internal carotid artery vascular smooth muscle cells.
Border zone geometry increases wall stress after myocardial infarction: contrast echocardiographic assessment
Authors: Jackson Benjamin M. , Gorman Joseph H. , Salgo Ivan S. , Moainie Sina L. , Plappert Theodore , St. John-Sutton Martin , Edmunds L. Henry , Gorman Robert C.
Published: March 2015

After myocardial infarction (MI), the border zone expands chronically, causing ventricular dilatation and congestive heart failure (CHF). In an ovine model ( n = 4) of anteroapical MI that results in CHF, contrast echocardiography was used to image short-axis left ventricular (LV) cross sections and identify border zone myocardium before and after coronary artery ligation. In the border zone at end systole, the LV endocardial curvature ( K) decreased from 0.86 ± 0.33 cm−1 at baseline to 0.35 ± 0.19 cm−1 at 1 h ( P < 0.05), corresponding to a mean decrease of 55%. Also in the border zone, the wall thickness ( h) decreased from 1.14 ± 0.26 cm at baseline to 1.01 ± 0.25 cm at 1 h ( P < 0.05), corresponding to a mean decrease of 11%. By Laplace's law, wall stress is inversely proportional to the product K · h. Therefore, a 55% decrease in K results in a 122% increase in circumferential stress; a 11% decrease in h results in a 12% increase in circumferential stress. These findings indicate that after MI, geometric changes cause increased dynamic wall stress, which likely contributes to border zone expansion and remodeling.
Disruption of cardiac Ena-VASP protein localization in intercalated disks causes dilated cardiomyopathy
Authors: Eigenthaler Martin , Engelhardt Stefan , Schinke Birgitta , Kobsar Anna , Schmitteckert Eva , Gambaryan Stepan , Engelhardt Catherine M. , Krenn Veit , Eliava Marina , Jarchau Thomas , Lohse Martin J. , Walter Ulrich , Hein Lutz
Published: March 2015

Vasodilator-stimulated phosphoprotein (VASP) and mammalian enabled (Mena) are actin cytoskeleton and signaling modulators. Ena-VASP proteins share an identical domain organization with an NH2-terminal Ena VASP homology (EVH1) domain, which mediates the binding of these proteins to FPPPP-motif containing partners such as zyxin and vinculin. VASP and Mena are abundantly expressed in the heart. However, previous studies showed that disruption by gene targeting of VASP or Mena genes in mice did not reveal any cardiac phenotype, whereas mice lacking both VASP and Mena died during embryonic development. To determine the in vivo function of Ena-VASP proteins in the heart, we used a dominant negative strategy with cardiac-specific expression of the VASP-EVH1 domain. Transgenic mice with cardiac myocyte-restricted, α-myosin heavy chain promoter-directed expression of the VASP-EVH1 domain were generated. Overexpression of the EVH1 domain resulted in specific displacement of both VASP and Mena from cardiac intercalated disks. VASP-EVH1 transgenic mice developed dilated cardiomyopathy with myocyte hypertrophy and bradycardia, which resulted in early postnatal lethality in mice with high levels of transgene expression. The results demonstrate that Ena-VASP proteins may play an important role in intercalated disk function at the interface between cardiac myocytes.
Acute p38 MAPK activation decreases force development in ventricular myocytes
Authors: Chen Yi , Rajashree Ravi , Liu Qinghang , Hofmann Polly
Published: March 2015

Evidence suggests that p38 mitogen-activated protein kinase (MAPK) activation influences cardiac function on an acute basis. The characterization and mechanisms by which this occurs were investigated in the present study. Adult rat ventricular myocytes treated with 1 mM arsenite for 30 min had a 16-fold increase in p38 MAPK phosphorylation that was attenuated by SB-203580 (a p38 MAPK inhibitor). Extracellular signal-regulated protein kinase (ERK) and c-Jun NH2-terminal kinase (JNK) were also minimally activated, but this activation was not sensitive to SB-203580. In addition, arsenite caused a p38 MAPK-independent translocation/activation of protein phosphatase 2a (PP2a) and decrease in phosphorylation of myosin light chain 2 (LC2). Arsenite-p38 MAPK activation led to translocation of heat shock protein 27 but not αB-crystallin to the myofilaments. Using isolated cardiomyocytes, we determined that arsenite reduces isometric tension without a change in Ca2+sensitivity of tension via p38 MAPK and lowers myofibrillar actomyosin Mg2+-ATPase activity in a p38 MAPK-independent manner. Thus arsenite induces a p38 MAPK-independent change in PP2a and LC2 that may account for the arsenite-dependent decrease in ATPase and a p38 MAPK-dependent modification of the myofilaments that decreases myocardial force development.
Rat model of pulmonary arteriovenous malformations after right superior cavopulmonary anastomosis
Authors: Starnes Sandra L. , Duncan Brian W. , Fraga Charles H. , Desai Shailesh Y. , Jones Thomas K. , Mathur Sanjeev K. , Rosenthal Geoffrey L. , Lupinetti Flavian M.
Published: March 2015

We developed a rat model of pulmonary arteriovenous malformations after cavopulmonary anastomosis. We sought to determine whether this model reproduces the angiographic and histologic features seen in the human condition. Eight Sprague-Dawley rats underwent a right superior cavopulmonary anastomosis with the use of microsurgical techniques. Between 2 and 13 mo, pulmonary angiography was performed, the animals were euthanized, and the lungs were removed. Microscopic sections of the lung were stained with an endothelial-specific antibody (von Willebrand factor). Microvessel density was determined by counting vessels staining positively for von Willebrand factor, and the shunted and nonshunted (control) lungs were compared for each animal. Pulmonary angiography revealed time-dependent development of arteriovenous malformations. Microvessel density demonstrated a time-dependent increase in the shunted lung compared with the control lung (simple linear regression of the ratio of the microvessel density of the shunted lung divided by the microvessel density of the control lung on time; R 2 = 0.79, P = 0.003). This animal model reproduces the same angiographic and microscopic features of pulmonary arteriovenous malformations that develop in humans after cavopulmonary anastomosis. This appears to be a valid model that may be used to further study etiologic mechanisms for this phenomenon.
Regulation of N- and C-type inactivation of Kv1.4 by pHo and K+: evidence for transmembrane communication
Authors: Li Xiaoyan , Bett Glenna C. L. , Jiang Xuejun , Bondarenko Vladimir E. , Morales Michael J. , Rasmusson Randall L.
Published: March 2015

Kv1.4 encodes a slowly recovering transient outward current ( I to), which inactivates by a fast N-type (intracellular ball and chain) mechanism but has slow recovery due to C-type inactivation. C-type inactivation of the NH2-terminal deletion mutant (fKv1.4ΔN) was inhibited by 98 mM extracellular K+concentration ([K+]o), whereas N-type was unaffected. In 98 mM [K+]o, removal of intracellular K+ concentration ([K+]i) speeded C-type inactivation but had no effect on N-type inactivation, suggesting that C-type inactivation is sensitive to K+ binding to intracellular sites. C-type inactivation is thought to involve closure of the extracellular pore mouth. However, a valine to alanine mutation on the intracellular side of S6 (V561A) of fKv1.4ΔN alters recovery and results in anomalous speeding of C-type inactivation with increasing [K+]o. Extracellular pH (pHo) modulated both N- and C-type inactivation through an S5-H5 linker histidine (H508) with acidosis speeding both N- and C-type inactivation. Mutation of an extracellular lysine to a tyrosine (K532Y) slowed C-type inactivation and inhibited the pH dependence of both N- and C-type inactivation. These results suggest that mutations, [K+], and pH modulate inactivation through membrane-spanning mechanisms involving S6.