bluuebunny/crossref_metadata_2025 huggingface dataset:
A chronic mouse model of myocardial ischemia-reperfusion: essential in cytokine studies
Authors: Nossuli T. O. , Lakshminarayanan V. , Baumgarten G. , Taffet G. E. , Ballantyne C. M. , Michael L. H. , Entman M. L.
Published: December 2017

Reperfusion of the ischemic myocardium is associated with a cytokine cascade that reflects a cellular response to injury. We studied this cascade in the mouse and found that acute surgical trauma in sham-operated animals obscured early changes in cytokine induction that occur during myocardial ischemia-reperfusion (MI/R). Therefore, we utilized a new implantable device that allows occlusion and reperfusion of the left anterior descending coronary artery in a closed-chest mouse at any time after instrumentation. Induction of interleukin (IL)-6 and tumor necrosis factor (TNF)-α mRNA in the whole heart was examined by RNase protection assay and quantitated by Phosphor- Imager. At 3 h after instrumentation, levels of IL-6 mRNA in sham-operated animals increased above those of control naive hearts, whereas this increase did not occur until after 1 day for TNF-α mRNA. The surgical trauma led to exaggeration of I/R cytokine induction with greater variance in response. At 3 days and 1 wk after instrumentation, levels of both IL-6 and TNF-α mRNA in sham-operated animals were comparable to those of naive hearts and induction responses in I/R were much less variant. We also found that 1 h of ischemia and 2 h of reperfusion at all time points of recovery (i.e., 3 h and 1, 3, and 7 days after instrumentation) led to a significant increase in IL-6 and TNF-α mRNA levels. In addition, 3 h of permanent occlusion, which did not induce any mRNA increase after 1 wk postinstrumentation, caused marked upregulation of IL-6 mRNA in an acutely prepared animal. This study of early cytokine responses evoked by MI/R highlights the need for dissipation of acute surgical trauma by using a chronic, closed-chest mouse preparation.
Dietary coenzyme Q10 supplement renders swine hearts resistant to ischemia-reperfusion injury
Authors: Maulik Nilanjana , Yoshida Tetsuya , Engelman Richard M. , Bagchi Debasis , Otani Hajime , Das Dipak K.
Published: December 2017

To examine whether nutritional supplementation of coenzyme Q10(CoQ10) can reduce myocardial ischemia-reperfusion injury, a group of swine was fed a regular diet supplemented with CoQ10 (5 mg ⋅ kg− 1 ⋅ day− 1) for 30 days. Another group of pigs that were fed a regular diet supplemented with placebo served as a control. After 30 days, isolated in situ pig hearts were prepared and hearts were perfused with a cardiopulmonary pump system. Each heart was subjected to 15 min of regional ischemia by snaring of the left anterior descending coronary artery, followed by 60 min of hypothermic cardioplegic global ischemia and 120 min of reperfusion. After the experiments were completed, myocardial infarct size was measured by triphenyltrazolium chloride staining methods. Postischemic left ventricular contractile function was better recovered in the CoQ10 group than in the control group of pigs. CoQ10-fed pigs revealed less myocardial infarction and less creatine kinase release from the coronary effluent compared with control pigs. The experimental group also demonstrated a smaller amount of malonaldehyde in the coronary effluent and a higher content of the endogenous antioxidants ascorbate and thiol. Significant induction of the expression of ubiquitin mRNA was also found in the hearts of the CoQ10-fed group. The results of this study demonstrate that nutritional supplementation of CoQ10 renders the hearts resistant to ischemia-reperfusion injury, probably by reducing the oxidative stress.
In spontaneously hypertensive rats alterations in aortic wall properties precede development of hypertension
Authors: van Gorp Ad W. , Schenau Dorette S. Van Ingen , Hoeks Arnold P. G. , Boudier Harry A. J. Struijker , de Mey Jo G. R. , Reneman Robert S.
Published: December 2017

In hypertension arterial wall properties do not necessarily depend on increased blood pressure alone. The present study investigates the relationship between the development of hypertension and thoracic aortic wall properties in 1.5-, 3-, and 6-mo-old spontaneously hypertensive rats (SHR); Wistar-Kyoto rats (WKY) served as controls. During ketamine-xylazine anesthesia, compliance and distensibility were assessed by means of a noninvasive ultrasound technique combined with invasive blood pressure measurements. Morphometric measurements provided in vivo media cross-sectional area and thickness, allowing the calculation of the incremental elastic modulus. Extracellular matrix protein contents were determined as well. Blood pressure was not significantly different in 1.5-mo-old SHR and WKY, but compliance and distensibility were significantly lower in SHR. Incremental elastic modulus was not significantly different between SHR and WKY at this age. Media thickness and media cross-sectional area were significantly larger in SHR than in WKY, but there was no consistent difference in collagen density and content between the strains. Blood pressure was significantly higher in 3- and 6-mo-old SHR than in WKY, and compliance was significantly lower in SHR. The findings in this study show that in SHR, in which hypertension develops over weeks, alterations in functional aortic wall properties precede the development of hypertension. The decrease in compliance and distensibility at a young age most likely results from media hypertrophy rather than a change in intrinsic elastic properties.
Immunochemical evidence for a unique GPI-anchored carbonic anhydrase isozyme in human cardiomyocytes
Authors: Knüppel-Ruppert Anja Sylvia , Gros Gerolf , Harringer Wolfgang , Kubis Hans-Peter
Published: December 2017

To clarify the controversial question of cell-specific distribution of carbonic anhydrase (CA) in the heart, endothelial cells and cardiomyocytes were isolated from porcine and human hearts and were characterized with cell-specific markers. CA activity was found in the microsomal fraction of both cell types. It was shown by Triton X-114 phase separation that both cell types possess a membrane-bound form of CA. These CAs share the same mechanism of membrane-anchoring via glycosylphosphatidylinositol (GPI), which excludes identity with transmembrane isoforms CA IX or CA XII. Western blotting analysis of human microsomes with anti-human CA IV antibodies revealed a marked difference in immunoreactivity. Endothelial CA activity resulted in 11-fold stronger CA IV bands compared with identical amounts of myocytic CA activity, indicating that cardiac endothelium and cardiomyocytes possess immunologically distinct forms of CA. We conclude that in human hearts CA IV is associated with the endothelium, whereas most of the CA in myocytes is not identical with one of the known CA isozymes. This suggests that cardiomyocytic CA is a novel isozyme.
Myocardial substrate metabolism influences left ventricular energetics in vivo
Authors: Korvald Christian , Elvenes Odd Petter , Myrmel Truls
Published: December 2017

The myocardial oxygen consumption (MV˙o2) to left ventricular pressure-volume area (PVA) relationship is assumed unaltered by substrates, despite varying phosphate-to-oxygen ratios and possible excess MV˙o2associated with fatty acid consumption. The validity of this assumption was tested in vivo. Left ventricular volumes and pressures were assessed with a combined conductance-pressure catheter in eight anesthetized pigs. MV˙o2was calculated from coronary flow and arterial-coronary sinus O2differences. Metabolism was altered by glucose-insulin-potassium (GIK) or Intralipid-heparin (IH) infusions in random order and monitored with [14C]glucose and [3H]oleate tracers. Profound shifts in glucose and fatty acid oxidation were observed. Contractility, coronary flow, and slope of the MV˙o2-PVA relationship were unchanged during GIK and IH infusions. MV˙o2at zero PVA (unloaded MV˙o2) was 0.16 ± 0.13 J ⋅ beat−1⋅ 100 g−1higher during IH compared with GIK infusion ( P = 0.001), a 48% increase. The study demonstrates a marked energetic advantage of glucose oxidation in the myocardium, profoundly affecting the MV˙o2-PVA relationship. This may in part explain the “oxygen-wasting” effect of lipid-enhancing interventions such as adrenergic drugs and ischemia.
Fluid shear stress increases membrane fluidity in endothelial cells: a study with DCVJ fluorescence
Authors: Haidekker Mark A. , L'Heureux Nicolas , Frangos John A.
Published: December 2017

Fluid shear stress (FSS) has been shown to be an ubiquitous stimulator of mammalian cell metabolism. Although many of the intracellular signal transduction pathways have been characterized, the primary mechanoreceptor for FSS remains unknown. One hypothesis is that the cytoplasmic membrane acts as the receptor for FSS, leading to increased membrane fluidity, which in turn leads to the activation of heterotrimetric G proteins (13). 9-(Dicyanovinyl)-julolidine (DCVJ) is a fluorescent probe that integrates into the cell membrane and changes its quantum yield with the viscosity of the environment. In a parallel-plate flow chamber, confluent layers of DCVJ-labeled human endothelial cells were exposed to different levels of FSS. With increased FSS, a reduced fluorescence intensity was observed, indicating an increase of membrane fluidity. Step changes of FSS caused an approximately linear drop of fluorescence within 5 s, showing fast and almost full recovery after shear cessation. A linear dose-response relationship between shear stress and membrane fluidity changes was observed. The average fluidity increase over the entire cell monolayer was 22% at 26 dyn/cm2. This study provides evidence for a link between FSS and membrane fluidity, and suggests that the membrane is an important flow mechanosensor of the cell.
Energy-wasteful total Ca2+handling underlies increased O2cost of contractility in canine stunned heart
Authors: Lee Shinyu , Araki Junichi , Imaoka Takeshi , Maesako Masaki , Iribe Gentaro , Miyaji Katsumasa , Mohri Satoshi , Shimizu Juichiro , Harada Mine , Ohe Tohru , Hirakawa Masahisa , Suga Hiroyuki
Published: December 2017

Postischemic myocardial stunning halved left ventricular contractility [end-systolic maximum elastance ( Emax)] and doubled the O2cost of Emaxin excised cross-circulated canine heart. We hypothesized that this increased O2cost derived from energy-wasteful myocardial Ca2+handling consisting of a decreased internal Ca2+recirculation, some futile Ca2+cycling, and a depressed Ca2+reactivity of Emax. We first calculated the internal Ca2+recirculation fraction (RF) from the exponential decay component of postextrasystolic potentiation. Stunning significantly accelerated the decay and decreased RF from 0.63 to 0.43 on average. We then combined the decreased RF with the halved Emaxand its doubled O2cost and analyzed total Ca2+handling using our recently developed integrative method. We found a decreased total Ca2+transport and a considerable shift of the relation between futile Ca2+cycling and Ca2+reactivity in an energy-wasteful direction in the stunned heart. These changes in total Ca2+handling reasonably account for the doubled O2cost of Emaxin stunning, supporting the hypothesis.
Role of EDHF in conduction of vasodilation along hamster cheek pouch arterioles in vivo
Authors: Welsh Donald G. , Segal Steven S.
Published: December 2017

We tested whether local and conducted responses to ACh depend on factors released from endothelial cells (EC) in cheek pouch arterioles of anesthetized hamsters. ACh was delivered from a micropipette (1 s, 500 nA), while arteriolar diameter (rest, ∼40 μm) was monitored at the site of application (local) and at 520 and 1,040 μm upstream (conducted). Under control conditions, ACh elicited local (22–65 μm) and conducted (14–44 μm) vasodilation. Indomethacin (10 μM) had no effect, whereas N ω-nitro-l-arginine (100 μM) reduced local and conducted vasodilation by 5–8% ( P < 0.05). Miconazole (10 μM) or 17-octadecynoic acid (17-ODYA; 10 μM) diminished local vasodilation by 15–20% and conducted responses by 50–70% ( P < 0.05), suggesting a role for cytochrome P-450 (CYP) metabolites in arteriolar responses to ACh. Membrane potential ( E m) was recorded in smooth muscle cells (SMC) and in EC identified with dye labeling. At rest (control E m, typically −30 mV), ACh evoked local (15–32 mV) and conducted (6–31 mV) hyperpolarizations in SMC and EC. Miconazole inhibited SMC and EC hyperpolarization, whereas 17-ODYA inhibited hyperpolarization of SMC but not of EC. Findings indicate that ACh-induced release of CYP metabolites from arteriolar EC evoke SMC hyperpolarization that contributes substantively to conducted vasodilation.
Endoplasmic reticulum contribution to the relaxant effect of cGMP- and cAMP-elevating agents in feline aorta
Authors: Mundiña-Weilenmann Cecilia , Vittone Leticia , Rinaldi Gustavo , Said Matilde , de Cingolani Gladys Chiappe , Mattiazzi Alicia
Published: December 2017

The contribution of endoplasmic reticulum (ER) and phosphorylation of phospholamban (PLB) to the relaxant effect of cGMP- and cAMP-elevating agents was studied in feline aorta. Sodium nitroprusside (NP, 100 μM) completely relaxed contracture induced by 10 μM norepinephrine. This NP-induced relaxation was partially prevented by tetraethylammonium, suggesting that a fraction of NP-induced relaxation was mediated by activation of K+channels. In the absence and presence of tetraethylammonium, the relaxant effect of NP was associated with a significant increase in Ser16phosphorylation of PLB immunodetected by phosphorylation site-specific antibodies. The relaxant effect of NP on aortic strips precontracted with 80 mM KCl was significantly reduced by 1 μM thapsigargin. This decrease, which represents the ER contribution to the relaxant effect of NP, reached 23 ± 9% at 100 μM NP and was closely associated with a dose-dependent increase in Ser16phosphorylation (128 ± 49% over control at 100 μM NP). Effects of NP were associated with a significant increase in activity of protein kinase G and were mimicked by 8-bromo-cGMP. Forskolin produced a dose-dependent relaxant effect on KCl-induced contracture, which reached 64 ± 8% at 50 μM and was associated with an increase in phosphorylation of Ser16residue of PLB (88 ± 18% over control). Thapsigargin reduced this relaxant effect by 38 ± 9%. 8-Bromo-cAMP mimicked effects of forskolin. The ER-mediated relaxant effect and the increase in Ser16phosphorylation produced by forskolin were partially blocked by the protein kinase A inhibitor H-89 (5 μM). The results indicate that ER partially contributes to the relaxant effect of NP and forskolin in feline aorta. This effect may be mediated by the associated increase in Ser16phosphorylation of PLB.
Peroxynitrite reversibly inhibits Ca2+-activated K+channels in rat cerebral artery smooth muscle cells
Authors: Brzezinska Anna K. , Gebremedhin Debebe , Chilian William M. , Kalyanaraman Balaraman , Elliott Stephen J.
Published: December 2017

Peroxynitrite (ONOO−) is a contractile agonist of rat middle cerebral arteries. To determine the mechanism responsible for this component of ONOO−bioactivity, the present study examined the effect of ONOO−on ionic current and channel activity in rat cerebral arteries. Whole cell recordings of voltage-clamped cells were made under conditions designed to optimize K+current. The effects of iberiotoxin, a selective inhibitor of large-conductance Ca2+-activated K+(BK) channels, and ONOO−(10–100 μM) were determined. At a pipette potential of +50 mV, ONOO−inhibited 39% of iberiotoxin-sensitive current. ONOO−was selective for iberiotoxin-sensitive current, whereas decomposed ONOO−had no effect. In excised, inside-out membrane patches, channel activity was recorded using symmetrical K+solutions. Unitary currents were sensitive to increases in internal Ca2+concentration, consistent with activity due to BK channels. Internal ONOO−dose dependently inhibited channel activity by decreasing open probability and mean open times. The inhibitory effect of ONOO−could be overcome by reduced glutathione. Glutathione, added after ONOO−, restored whole cell current amplitude to control levels and reverted single-channel gating to control behavior. The inhibitory effect of ONOO−on membrane K+current is consistent with its contractile effects in isolated cerebral arteries and single myocytes. Taken together, our data suggest that ONOO−has the potential to alter cerebral vascular tone by inhibiting BK channel activity.
Evidence for modulation of smooth muscle force by the p38 MAP kinase/HSP27 pathway
Authors: Yamboliev Ilia A. , Hedges Jason C. , Mutnick Jack L.-M. , Adam Leonard P. , Gerthoffer William T.
Published: December 2017

Mitogen-activated protein (MAP) kinases signal to proteins that could modify smooth muscle contraction. Caldesmon is a substrate for extracellular signal-related kinases (ERK) and p38 MAP kinases in vitro and has been suggested to modulate actin-myosin interaction and contraction. Heat shock protein 27 (HSP27) is downstream of p38 MAP kinases presumably participating in the sustained phase of muscle contraction. We tested the role of caldesmon and HSP27 phosphorylation in the contractile response of vascular smooth muscle by using inhibitors of both MAP kinase pathways. In intact smooth muscle, PD-098059 abolished endothelin-1 (ET-1)-stimulated phosphorylation of ERK MAP kinases and caldesmon, but p38 MAP kinase activation and contractile response remained unaffected. SB-203580 reduced muscle contraction and inhibited p38 MAP kinase and HSP27 phosphorylation but had no effect on ERK MAP kinase and caldesmon phosphorylation. In permeabilized muscle fibers, SB-203580 and a polyclonal anti-HSP27 antibody attenuated ET-1-dependent contraction, whereas PD-098059 had no effect. These results suggest that ERK MAP kinases phosphorylate caldesmon in vivo but that activation of this pathway is unnecessary for force development. The generation of maximal force may be modulated by the p38 MAP kinase/HSP27 pathway.
Effects of estrogen on venous function in rats with chronic heart failure
Authors: Nekooeian Ali Akbar , Pang Catherine C. Y.
Published: December 2017

The effect of 17β-estradiol on venous function was investigated in ovariectomized rats with heart failure. Rats (50–60 days old) were ovariectomized and implanted with 60-day-release pellets that contain 17β-estradiol (1.5 mg) or vehicle. The left coronary artery was ligated 7 days later. Another group of ovariectomized rats was given vehicle pellets and then a sham operation was performed. The rats were studied while under pentobarbital anesthesia at 7 wk after ligation. Ligated rats, relative to sham groups, had lower mean arterial pressure (MAP, −34 mmHg) and cardiac output (CO, −38%); higher arterial resistance ( R A, +12%) and venous resistance ( R V, +116%); mean circulatory filling pressure (MCFP, +40%) and left ventricular end-diastolic pressure (LVEDP, +11 mmHg); and similar cardiovascular responses to norepinephrine (NE). Treatment of ligated rats with 17β-estradiol increased CO (+16%); reduced R A (−16%), R V(−35%), MCFP (−23%), and LVEDP (−3 mmHg); and augmented MAP, R V, and MCFP responses to NE. Therefore, 17β-estradiol reduced MCFP, and this reduced preload (LVEDP). 17β-Estradiol decreased R V, which, along with decreased R A (afterload), led to an increase in CO. 17β-Estradiol likely augmented vasoconstriction to NE through an improvement on the cardiovascular status.
Systemic and renal hemodynamics after moderate hemodilution with HbOCs in anesthetized rabbits
Authors: Caron Alexis , Menu Patrick , Faivre-Fiorina Béatrice , Labrude Pierre , Alayash Abdu , Vigneron Claude
Published: December 2017

Hb-based O2-carrying solutions (HbOCs) have been developed as red blood cell substitutes for use in patients undergoing hemodilution. Variously modified Hb with diverse solution properties have been shown to produce variable hemodynamic responses. We examined, through pulsed-Doppler velocimetry, the systemic and renal hemodynamic effects of dextran-benzene-tetracarboxylate-conjugated (Hb-Dex-BTC), bis(3,5-dibromosalicyl)fumarate cross-linked (αα-Hb), and o-raffinose-polymerized ( o-raffinose-Hb) Hb perfused in rabbits after moderate hemodilution (30% hematocrit), and we compared the effects of these Hb solutions with the effects elicited by plasma volume expanders. In addition, vascular hindrance (resistance/blood viscosity at 128.5 s− 1) was calculated to determine whether a moderate decrease in the viscosity of blood mixed with HbOCs may impair vasoconstriction as a result of autoregulation after infusion of cell-free Hb. No changes were observed in renal hemodynamics after hemodilution with reference or Hb solutions. Increase in blood pressure and vascular resistance was found with Hb-Dex-BTC and αα-Hb (for 180 min) and, to a lesser extent, with o-raffinose-Hb (for 120 min). Furthermore, Hb-Dex-BTC (high viscosity) and o-raffinose-Hb (medium viscosity) induced comparable increases in vascular hindrance (from 0.091 to 0.159 and from 0.092 to 0.162 cm− 1, respectively) but far less than that produced by αα-Hb (low viscosity, from 0.092 to 0.200 cm− 1). These results suggest that maintaining the viscosity of blood by infusing solutions with high viscosity makes it possible to limit vasoconstriction due to autoregulation mechanisms and mainly caused by hemodilution per se.
ET-1 in the myocardial interstitium: relation to myocyte ECE activity and expression
Authors: Ergul Adviye , Walker C. Allyson , Goldberg Aron , Baicu Simona C. , Hendrick Jennifer W. , King Mary K. , Spinale Francis G.
Published: December 2017

Increased plasma levels of endothelin-1 (ET-1) have been identified in congestive heart failure (CHF), but local myocardial interstitial ET-1 levels and the relation to determinants of ET-1 synthesis remain to be defined. Accordingly, myocardial interstitial ET-1 levels and myocyte endothelin-converting enzyme (ECE)-1 activity and expression with the development of CHF were examined. Pigs were instrumented with a microdialysis system to measure myocardial interstitial ET-1 levels with pacing CHF (240 beats/min, 3 wk; n = 9) and in controls ( n = 14). Plasma ET-1 was increased with CHF (15 ± 1 vs. 9 ± 1 fmol/ml, P < 0.05) as was total myocardial ET-1 content (90 ± 15 vs. 35 ± 5 fmol/g, P < 0.05). Paradoxically, myocardial interstitial ET-1 was decreased in CHF (32 ± 4 vs. 21 ± 2 fmol/ml, P < 0.05), which indicated increased ET-1 uptake by the left ventricular (LV) myocardium with CHF. In isolated LV myocyte preparations, ECE-1 activity was increased by twofold with CHF ( P < 0.05). In LV myocytes, both ECE-1a and ECE-1c mRNAs were detected, and ECE-1a expression was upregulated fivefold in CHF myocytes ( P < 0.05). In conclusion, this study demonstrated compartmentalization of ET-1 in the myocardial interstitium and enhanced ET-1 uptake with CHF. Thus a local ET-1 system exists at the level of the myocyte, and determinants of ET-1 biosynthesis are selectively regulated within this myocardial compartment in CHF.
Regulation of types I and III NOS in ovine uterine arteries by daily and acute estrogen exposure
Authors: Salhab Walid A. , Shaul Philip W. , Cox Blair E. , Rosenfeld Charles R.
Published: December 2017

Nitric oxide contributes to estrogen-mediated uterine vasodilation; however, the nitric oxide synthases (NOS) involved and their location within uterine arteries are incompletely documented. We investigated the effects of repetitive daily and acute estradiol-17β (E2β) exposure on uterine hemodynamics and NOS abundance and localization in uterine arteries from nonpregnant ovariectomized ewes receiving daily intravenous E2β (1 μg/kg, n = 5) or no E2β ( n = 7) for 5 days to determine NOS abundance, cGMP contents, and NOS immunohistochemistry. Daily E2β increased basal and E2β-mediated rises in uterine blood flow (UBF) 36 and 43% (<0.01), respectively, calcium-dependent NOS activity 150% ( P < 0.02) in endothelium-intact and -denuded (∼40% of total NOS) arteries, and cGMP contents 39% ( P < 0.05). Endothelial (eNOS) was detected in luminal endothelium, whereas neuronal NOS (nNOS) protein was only in the media. A second group of ewes received E2β (1 μg/kg iv) for 4 days and acute intravenous E2β ( n = 8) or vehicle ( n= 4) on day 5. UBF rose 5.5-fold ( P < 0.001) 115 min after E2β, at which time only endothelium-derived calcium-dependent NOS activity increased 30 ± 13% ( P < 0.05). Daily E2β enhances basal and E2β-mediated increases in UBF, which parallel increases in endothelium-derived eNOS and smooth muscle-derived nNOS. Acute E2β, however, selectively increases endothelium-derived eNOS.
Cardiac interstitial bradykinin release during ischemia is enhanced by ischemic preconditioning
Authors: Pan Hui-Lin , Chen Shao-Rui , Scicli Gloria M. , Carretero Oscar A.
Published: December 2017

Ischemic preconditioning is known to protect the myocardium from ischemia-reperfusion injury. We examined the transmural release of bradykinin during myocardial ischemia and the influence of ischemic preconditioning on bradykinin release during subsequent myocardial ischemia. Myocardial ischemia was induced by occlusion of the left anterior descending coronary artery in anesthetized cats. Cardiac microdialysis was performed by implantation and perfusion of dialysis probes in the epicardium and endocardium. In eight animals, bradykinin release was greater in the endocardium than in the epicardium (14.4 ± 2.8 vs. 7.3 ± 1.7 ng/ml, P < 0.05) during 30 min of ischemia. In seven animals subjected to preconditioning, myocardial bradykinin release was potentiated significantly from 2.4 ± 0.6 ng/ml during the control period to 23.1 ± 2.5 ng/ml during 30 min of myocardial ischemia compared with the non-preconditioning group (from 2.7 ± 0.6 to 13.4 ± 1.9 ng/ml, P < 0.05, n = 6). Thus this study provides further evidence that transmural gradients of bradykinin are produced during ischemia. The results also suggest that ischemic preconditioning enhances bradykinin release in the myocardial interstitial fluid during subsequent ischemia, which is likely one of the mechanisms of cardioprotection of ischemic preconditioning.
Effect of the NADPH oxidase inhibitor apocynin on ischemia-reperfusion lung injury
Authors: Dodd-O Jeffrey M. , Pearse David B.
Published: December 2017

Apocynin (4-hydroxy-3-methoxy-acetophenone) inhibits NADPH oxidase in activated polymorphonuclear (PMN) leukocytes, preventing the generation of reactive oxygen species. To determine if apocynin attenuates ischemia-reperfusion lung injury, we examined the effects of apocynin (0.03, 0.3, and 3 mM) in isolated in situ sheep lungs. In diluent-treated lungs, reperfusion with blood (180 min) after 30 min of ischemia (ventilation 28% O2, 5% CO2) caused leukocyte sequestration in the lung and increased vascular permeability [reflection coefficient for albumin (ςalb) 0.47 ± 0.10, filtration coefficient ( Kf) 0.14 ± 0.03 g · min−1· mmHg−1· 100 g−1] compared with nonreperfused lungs (ςalb0.77 ± 0.03, Kf0.03 ± 0.01 g · min−1· mmHg−1· 100 g−1; P < 0.05). Apocynin attenuated the increased protein permeability at 0.3 and 3 mM (ςalb0.69 ± 0.05 and 0.91 ± 0.03, respectively, P < 0.05); Kfwas decreased by 3 mM apocynin (0.05 ± 0.01 g · min−1· mmHg−1· 100 g−1, P < 0.05). Diphenyleneiodonium (DPI, 5 μM), a structurally unrelated inhibitor of NADPH oxidase, worsened injury ( Kf0.32 ± 0.07 g · min−1· mmHg−1· 100 g−1, P < 0.05). Neither apocynin nor DPI affected leukocyte sequestration. Apocynin and DPI inhibited whole blood chemiluminescence and isolated PMN leukocyte-induced resazurin reduction, confirming NADPH oxidase inhibition. Apocynin inhibited pulmonary artery hypertension and perfusate concentrations of cyclooxygenase metabolites, including thromboxane B2. The cyclooxygenase inhibitor indomethacin had no effect on the increased vascular permeability, suggesting that cyclooxygenase inhibition was not the explanation for the apocynin results. Apocynin prevented ischemia-reperfusion lung injury, but the mechanism of protection remains unclear.
Retinoic acid upregulates β1-integrin in vascular smooth muscle cells and alters adhesion to fibronectin
Authors: Medhora Meetha M.
Published: December 2017

Retinoic acid has an established physiological role in differentiation, development, and cellular growth. This study investigated the action of all- trans retinoic acid (ATRA) on vascular integrins, cell-surface receptors that control growth and remodeling of blood vessels. The β1-integrin subunit mRNA and protein was induced after treatment with ATRA in two different rat vascular smooth muscle cell lines. To relate this result to the in vivo state, the aortas from adult rats fed with therapeutic doses of ATRA were examined for β1-integrin protein. A significant upregulation of the integrin subunit was observed in vivo. To assess if this increase contributed to physiological changes in cellular function, cells treated with ATRA were tested for alterations in adhesion to extracellular matrix proteins. The cells exposed to the retinoid were seen to adhere more strongly to fibronectin, via the β1-integrin. These results showed that modulation of vascular integrins by ATRA in adult rats contributes to functional changes that can cause remodeling of blood vessels.
Reactive oxygen species may contribute to reduced endothelium-dependent dilation in rats fed high salt
Authors: Lenda Deborah M. , Sauls Bryan A. , Boegehold Matthew A.
Published: December 2017

In normotensive rats, an increase in dietary salt leads to decreased arteriolar responsiveness to acetylcholine (ACh) because of suppressed local nitric oxide (NO) activity. We evaluated the possibility that generation of reactive oxygen species in the arteriolar wall is responsible for this loss of NO activity. Arteriolar responses to iontophoretically applied ACh were examined in the superfused spinotrapezius muscle of Sprague-Dawley rats fed a low-salt (LS; 0.45%) or high-salt diet (HS; 7%) for 4–5 wk. Responses to ACh were significantly depressed in HS rats but returned to normal in the presence of the oxidant scavengers superoxide dismutase + catalase or 2,2,6,6-tetamethylpiperidine- N-oxyl (TEMPO) + catalase. Arteriolar responses to the NO donor sodium nitroprusside were similar in HS and LS rats. Arteriolar and venular wall oxidant activity, as determined by reduction of tetranitroblue tetrazolium, was significantly greater in HS rats than in LS rats. Exposure to TEMPO + catalase reduced microvascular oxidant levels to normal in HS rats. These data suggest that a high-salt diet leads to increased generation of reactive oxygen species in striated muscle microvessels, and this increased oxidative state may be responsible for decreased endothelium-dependent responses associated with high salt intake.
Estrogen reduces mouse cerebral artery tone through endothelial NOS- and cyclooxygenase-dependent mechanisms
Authors: Geary Greg G. , Krause Diana N. , Duckles Sue P.
Published: December 2017

Gender and estrogen status are known to influence the incidence and severity of cerebrovascular disease. The vasoprotective effects of estrogen are thought to include both nitric oxide-dependent and independent mechanisms. Therefore, using small, resistance-sized arteries pressurized in vitro, the present study determined the effect of gender and estrogen status on myogenic reactivity of mouse cerebral arteries. Luminal diameter was measured in middle cerebral artery segments from males and from females that were either untreated, ovariectomized (OVX), or OVX with estrogen replacement (OVX + EST). The maximal passive diameters of arteries from all four groups were similar. In response to increases in transmural pressure, diameters of arteries from males and OVX females were smaller compared with diameters of arteries from either untreated or OVX + EST females. In the presence of N G-nitro-l-arginine methyl ester, artery diameters decreased in all groups, but diameters remained significantly smaller in arteries from males and OVX females compared with untreated and OVX + EST females. After endothelium removal or when inhibition of nitric oxide synthase and cyclooxygenase were combined, differences in diameters of arteries from OVX and OVX + EST were abolished. These data suggest that chronic estrogen treatment modulates myogenic reactivity of mouse cerebral arteries through both endothelium-derived cyclooxygenase- and nitric oxide synthase-dependent mechanisms.
Effects of aging on single cardiac myocyte function in Fischer 344 x Brown Norway rats
Authors: Wahr Philip A. , Michele Daniel E. , Metzger Joseph M.
Published: December 2017

The Fischer 344 x Brown Norway (F344xBN) rat has been demonstrated to have a lower incidence of age-related pathology than other rat strains. Therefore, to elucidate the effects of aging on cardiac function, uncomplicated by compensatory effects caused by age-related pathology, cardiac myocytes were isolated from female F344xBN rats at 6 (young) and 32–33 (old) mo of age. Myocytes showed an increase in the relative amount of β-myosin heavy chain with advanced age and a significant rightward shift in the tension-pCa curve from 5.78 ± 0.02 pCa units in young adult myocytes to 5.66 ± 0.03 pCa units. Consistent with a shift to a slower myosin isoform, the time from stimulation to peak sarcomere shortening increased with age from 50.5 ± 1.3 to 58.9 ± 1.0 ms. In contrast, no age-related difference was found in either the relengthening parameters or the Ca2+transient, indicating that relaxation is not directly altered by aging. This latter finding is at variance with previous studies in rat strains with higher rates of pathology. We conclude, therefore, that the primary effect of aging in isolated cardiac myocytes from the F344xBN rat model is a shift in the myosin heavy chain isoform. Changes in relaxation seen in other rat strains may result from compensatory mechanisms induced by pathological conditions.
Adverse effects of constitutively active α1B-adrenergic receptors after pressure overload in mouse hearts
Authors: Wang Bing H. , Du Xiao-Jun , Autelitano Dominic J. , Milano Carmelo A. , Woodcock Elizabeth A.
Published: December 2017

Cardiac hypertrophy and function were studied 6 wk after constriction of the thoracic aorta (TAC) in transgenic (TG) mice expressing constitutively active mutant α1B-adrenergic receptors (ARs) in the heart. Hearts from sham-operated TG animals and nontransgenic littermates (WT) were similar in size, but hearts from TAC/TG mice were larger than those from TAC/WT mice, and atrial natriuretic peptide mRNA expression was also higher. Lung weight was markedly increased in TAC/TG animals, and the incidence of left atrial thrombus formation was significantly higher. Ventricular contractility in anesthetized animals, although it was increased in TAC/WT hearts, was unchanged in TAC/TG hearts, implying cardiac decompensation and progression to failure in TG mice. There was no increase in α1A-AR mRNA expression in TAC/WT hearts, and expression was significantly reduced in TAC/TG hearts. These findings show that cardiac expression of constitutively actively mutant α1B-ARs is detrimental in terms of hypertrophy and cardiac function after pressure overload and that increased α1A-AR mRNA expression is not a feature of the hypertrophic response in this murine model.
H2O2mediates Ca2+- and MLC20phosphorylation-independent contraction in intact and permeabilized vascular muscle
Authors: Pelaez Nancy J. , Braun Tracey R. , Paul Richard J. , Meiss Richard A. , Packer C. Subah
Published: December 2017

One purpose of the current study was to establish whether vasoconstriction occurs in all vessel types in response to H2O2. Isometric force was measured in pulmonary venous and arterial rings, and isobaric contractions were measured in mesenteric arteries and veins in response to H2O2. A second purpose was to determine whether H2O2-induced contraction is calcium independent. The addition of H2O2to calcium-depleted (using the Ca2+ionophore ionomycin in zero calcium EGTA buffer) muscle caused contraction. Furthermore, permeabilized muscle contracted in response to H2O2even in zero Ca2+. The final purpose was to determine whether the 20-kDa regulatory myosin light chain (MLC20) phosphorylation plays a role in H2O2-induced contraction. Pulmonary arterial strips were freeze-clamped at various time points during H2O2-induced contractions, and the relative amounts of phosphorylated MLC20were measured. H2O2caused dose-dependent contractions that were independent of MLC20phosphorylation. ML-9, a myosin light chain kinase inhibitor, had no effect on the H2O2contractile response. In conclusion, H2O2induces Ca2+- and MLC20phosphorylation-independent contraction in pulmonary and systemic arterial and venous smooth muscle.
Dilated cardiomyopathy in transgenic mice expressing a mutant A subunit of protein phosphatase 2A
Authors: Brewis Neil , Ohst Kim , Fields Katherine , Rapacciuolo Antonio , Chou Danny , Bloor Colin , Dillmann Wolfgang , Rockman Howard , Walter Gernot
Published: December 2017

The protein phosphatase 2A (PP2A) holoenzyme consists of a catalytic subunit, C, and two regulatory subunits, A and B. The PP2A core enzyme is composed of subunits A and C. Both the holoenzyme and the core enzyme are similarly abundant in heart tissue. Transgenic mice were generated expressing high levels of a dominant negative mutant of the A subunit (AΔ5) in the heart, skeletal muscle, and smooth muscle that competes with the endogenous A subunit for binding the C subunit but does not bind B subunits. We found that the ratio of core enzyme to holoenzyme was increased in AΔ5-expressing hearts. Importantly, already at day 1 after birth, AΔ5-transgenic mice had an increased heart weight-to-body weight ratio that persisted throughout life. Echocardiographic analysis of AΔ5-transgenic hearts revealed increased end-diastolic and end-systolic dimensions and decreased fractional shortening. In addition, the thickness of the septum and of the left ventricular posterior wall was significantly reduced. On the basis of these findings, we consider the heart phenotype of AΔ5-transgenic mice to be a form of dilated cardiomyopathy that frequently leads to premature death.
SAPKs regulation of ischemic preconditioning
Authors: Sato Motoaki , Cordis Gerald A. , Maulik Nilanjana , Das Dipak K.
Published: December 2017

The role of stress-activated protein kinases (SAPKs), c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein (MAP) kinase, in preconditioning (PC) was examined with the use of isolated rat hearts subjected to four cyclic episodes of 5-min ischemia and 10-min reperfusion followed by 30-min ischemia and 2-h reperfusion (I/R). A group of hearts was preperfused with 100 μM curcumin, a c-Jun and JNK1 inhibitor, or 5 μM SB 203580, a p38 MAP kinase inhibitor. Another group of hearts was preperfused with 20 μM anisomycin, a stimulator for both JNK and p38 MAP kinases. I/R increased the protein levels of JNK1, c-Jun, and p38 MAP kinase. PC also enhanced the induction of these kinases, but subsequent I/R-mediated increase was blocked by PC. Curcumin blocked I/R- and PC-mediated increase in JNK1 and c-Jun protein levels, whereas it had no effects on p38 MAP kinase. SB 203580, on the other hand, was equally effective in reducing the p38 MAP kinase activation but exerted no effects on JNK1 and c-Jun induction. I/R-mediated increased myocardial infarction was reduced by any of the following compounds: anisomycin, curcumin, and SB 203580. The cardioprotective effects of PC were abolished by either curcumin or SB 203580. The results demonstrate that PC is mediated by a signal-transduction pathway involving both JNK1 and p38 MAP kinase. Activation of SAPKs, although transient, is obligatory for PC.
Ca2+ handling in isolated human atrial myocardium
Authors: Maier Lars S. , Barckhausen Paul , Weisser Jutta , Aleksic Ivo , Baryalei Mersa , Pieske Burkert
Published: December 2017

Physiologically, human atrial and ventricular myocardium are coupled by an identical beating rate and rhythm. However, contractile behavior in atrial myocardium may be different from that in ventricular myocardium, and little is known about intracellular Ca2+handling in human atrium under physiological conditions. We used rapid cooling contractures (RCCs) to assess sarcoplasmic reticulum (SR) Ca2+ content and the photoprotein aequorin to assess intracellular Ca2+ transients in atrial and ventricular muscle strips isolated from nonfailing human hearts. In atrial myocardium ( n = 19), isometric twitch force frequency dependently (0.25–3 Hz) increased by 78 ± 25% (at 3 Hz; P < 0.05). In parallel, aequorin light signals increased by 111 ± 57% ( P < 0.05) and RCC amplitudes by 49 ± 13% ( P < 0.05). Similar results were obtained in ventricular myocardium ( n = 13). SR Ca2+ uptake (relative to Na+/Ca2+ exchange) frequency dependently increased in atrial and ventricular myocardium ( P < 0.05). With increasing rest intervals (1–240 s), atrial myocardium ( n = 7) exhibited a parallel decrease in postrest twitch force (at 240 s by 68 ± 5%, P < 0.05) and RCCs (by 49 ± 10%, P < 0.05). In contrast, postrest twitch force and RCCs significantly increased in ventricular myocardium ( n = 6). We conclude that in human atrial and ventricular myocardium the positive force-frequency relation results from increased SR Ca2+ turnover. In contrast, rest intervals in atrial myocardium are associated with depressed contractility and intracellular Ca2+ handling, which may be due to rest-dependent SR Ca2+ loss (Ca2+ leak) and subsequent Ca2+ extrusion via Na+/Ca2+ exchange. Therefore, the influence of rate and rhythm on mechanical performance is not uniform in atrial and ventricular myocardium.
Aldose reductase inhibition alone or combined with an adenosine A3 agonist reduces ischemic myocardial injury
Authors: Tracey W. Ross , Magee William P. , Ellery Craig A. , MacAndrew Joseph T. , Smith Andrew H. , Knight Delvin R. , Oates Peter J.
Published: December 2017

This study investigated whether aldose reductase (AR) inhibition with zopolrestat, either alone or in combination with an adenosine A3-receptor agonist (CB-MECA), reduced myocardial ischemic injury in rabbit hearts subjected to 30 min of regional ischemia and 120 min of reperfusion. Zopolrestat reduced infarct size by up to 61%, both in vitro (2 nM to 1 μM; EC50 = 24 nM) and in vivo (50 mg/kg). Zopolrestat reduced myocardial sorbitol concentration (index of AR activity) by >50% (control, 15.0 ± 2.2 nmol/g; 200 nM zopolrestat, 6.7 ± 1.3 nmol/g). A modestly cardioprotective concentration of CB-MECA (0.2 nM) allowed a 50-fold reduction in zopolrestat concentration while providing a similar reduction in infarct size (infarct area/area at risk: control, 62 ± 2%; 1 μM zopolrestat, 24 ± 5%; 20 nM zopolrestat plus 0.2 nM CB-MECA, 20 ± 4%). In conclusion, AR inhibition is cardioprotective both in vitro and in vivo. Furthermore, combining zopolrestat with an A3 agonist allows a reduction in the zopolrestat concentration while maintaining an equivalent degree of cardioprotection.
Adenosine A2a-receptor activation increases contractility in isolated perfused hearts
Authors: Monahan Thomas S. , Sawmiller Darrell R. , Fenton Richard A. , Dobson James G.
Published: December 2017

Adenosine A2a-receptor activation enhances shortening of isolated cardiomyocytes. In the present study the effect of A2a-receptor activation on the contractile performance of isolated rat hearts was investigated by recording left ventricular pressure (LVP) and the maximal rate of LVP development (+dP/d tmax). With constant-pressure perfusion, adenosine caused concentration-dependent increases in LVP and +dP/d tmax, with detectable increases of 4.1 and 4.8% at 10−6M and maximal increases of 12.0 and 11.1% at 10−4M, respectively. The contractile responses were prevented by the A2a-receptor antagonists chlorostyryl-caffeine and aminofuryltriazolotriazinyl-aminoethylphenol (ZM-241385) but were not affected by the β1-adrenergic antagonist atenolol. The adenosine A1-receptor antagonist dipropylcyclopentylxanthine and pertussis toxin potentiated the positive inotropic effects of adenosine. The A2a-receptor agonists ethylcarboxamidoadenosine and dimethoxyphenyl-methylphenylethyl-adenosine also enhanced contractility. With constant-flow perfusion, 10−5M adenosine increased LVP and +dP/d tmaxby 5.5 and 6.0%, respectively. In the presence of the coronary vasodilator hydralazine, adenosine increased LVP and +dP/d tmaxby 7.5 and 7.4%, respectively. Dipropylcyclopentylxanthine potentiated the adenosine contractile responses with constant-flow perfusion in the absence and presence of hydralazine. These increases in contractile performance were also antagonized by chlorostyryl-caffeine and ZM-241385. The results indicate that adenosine increases contractile performance via activation of A2areceptors in the intact heart independent of β1-adrenergic receptor activation or changes in coronary flow.
Three-week neonatal hypoxia reduces blood CGRP and causes persistent pulmonary hypertension in rats
Authors: Keith I. M. , Tjen-A-Looi S. , Kraiczi H. , Ekman R.
Published: December 2017

To increase understanding of persistent pulmonary hypertension, we examined chronic pulmonary effects of hypoxia at birth and their relationships with immunoreactive levels of the potent vasodilator, calcitonin gene-related peptide (CGRP). Rats were born in 10% hypobaric hypoxia, where they remained for 1–2 days, or in 15% hypoxia, where they remained for 21 days. All were then reared in normoxia for 3 mo followed by reexposure to 10% hypoxia for 7 days (H→H) or continued normoxia (H→N); age-matched normoxic rats were hypoxic for the last 7 days (N→H) or normoxic throughout (N→N). Results are as follows. Pulmonary arterial pressure (PPA) in 10% H→N rats was normal at the end of the experiment (13 wk), but in rats reexposed to hypoxia (H→H), pressure rose to 19% above N→H controls. In 15% H→N rats, PPAremained high, similar to that of N→H rats, and increased further by 40% on reexposure (H→H). Medial thickness of small pulmonary arteries in 10% H→H rats also increased by 40% over N→H controls and was equally high in 15% H→N and H→H rats. In N→H rats from both experiments, right ventricular hypertrophy index (RVH) was increased after hypoxia at 15–16 wk. Also, in the 15% study, RVH remained elevated in H→N rats and increased in H→H rats by 19% above N→H controls. Blood CGRP was reduced by neonate and adult hypoxia, and hypoxic reexposure (H→H) further lowered blood CGRP in the 15% but not 10% study. Declining left ventricular blood CGRP correlated highly with logarithmically increasing PPAin the 15% study ( r = −0.81, P = 0.000). In conclusion, 1) short perinatal exposure to 10% O2exacerbated pulmonary hypertension with hypoxia later in life, 2) 15% O2at birth and for 21 days caused persistent pulmonary hypertension and exacerbation with reexposure, and 3) PPAcorrelated highly with declining blood CGRP levels in the 15% study.
Skin microvascular adaptations during maturation and aging of hairless mice
Authors: Vollmar Brigitte , Morgenthaler Martin , Amon Michaela , Menger Michael D.
Published: December 2017

Using intravital fluorescence microscopy in the ears of hairless mice, we determined skin microvascular adaptations during the process of aging from juvenile to adult and senescent life (6–78 wk). Despite an increase of ear area within the first 36 wk, the number and branching pattern of both arteriolar and venular microvessels remained constant during the whole life period. Both arterioles and venules exhibited an increase in length, diameter, and intervascular distance up to the age of 36 wk. With the increase of the size of the ears, the observation that cutaneous capillary density remained unchanged implied new capillary formation. During aging to 78 wk, capillary density in the ears was reduced to ∼40%. Functional analysis revealed an appropriate hyperemic response to a 2-min period of ischemia during late juvenile and adult life, which, however, was markedly reduced during senescence. Thus, except for capillaries, there is no indication for age-related new vessel formation. The process of aging from adult to senescent life does not cause any significant remodeling but is associated with a decrease of nutritive perfusion and a functional impairment to respond to stimuli such as ischemia.