ß(3) adrenergic stimulation of the cardiac Na+-K+ pump by reversal of an inhibitory oxidative modification.

BACKGROUND: inhibition of L-type Ca(2+) current contributes to negative inotropy of ß(3) adrenergic receptor (ß(3) AR) activation, but effects on other determinants of excitation-contraction coupling are not known. Of these, the Na(+)-K(+) pump is of particular interest because of adverse effects attributed to high cardiac myocyte Na(+) levels and upregulation of the ß(3) AR in heart failure. METHODS AND RESULTS: we voltage clamped rabbit ventricular myocytes and identified electrogenic Na(+)-K(+) pump current (I(p)) as the shift in holding current induced by ouabain. The synthetic ß(3) AR agonists BRL37344 and CL316,243 and the natural agonist norepinephrine increased I(p). Pump stimulation was insensitive to the ß(1)/ß(2) AR antagonist nadolol and the protein kinase A inhibitor H-89 but sensitive to the ß(3) AR antagonist L-748,337. Blockade of nitric oxide synthase abolished pump stimulation and an increase in fluorescence of myocytes loaded with a nitric oxide-sensitive dye. Exposure of myocytes to ß(3) AR agonists decreased ß(1) Na(+)-K(+) pump subunit glutathionylation, an oxidative modification that causes pump inhibition. The in vivo relevance of this was indicated by an increase in myocardial ß(1) pump subunit glutathionylation with elimination of ß(3) AR-mediated signaling in ß(3) AR(-/-) mice. The in vivo effect of BRL37344 on contractility of the nonfailing and failing heart in sheep was consistent with a beneficial effect of Na(+)-K(+) pump stimulation in heart failure. CONCLUSIONS: the ß(3) AR mediates decreased ß(1) subunit glutathionylation and Na(+)-K(+) pump stimulation in the heart. Upregulation of the receptor in heart failure may be a beneficial mechanism that facilitates the export of excess Na(+).

Mesenchymal stem cells: isolation, characterisation and in vivo fluorescent dye tracking.

Cell therapies have been used to regenerate the heart by direct myocardial delivery, by coronary infusion and by surface attached scaffolds. Multipotent mesenchymal stem cells (MSC) with capacity to differentiate into cardiomyocytes and other cell lines have been predominantly trialled in rodents. However, large animal models are increasingly needed to translate basic research into new, safe regenerative therapies. Understanding the mode of action of cell therapies in the mammalian heart has been limited by cell tracking capability. This study examined the ability to track the fate of allogeneic MSC in sheep using various fluorescent dyes. MSC isolated from sheep bone marrow were grown in culture following extraction and flow cytometric characterisation. After labelling with fluorescent tracking dyes (e.g. CFSE and DiI) cells were tested for in vitro and in vivo signal up to six weeks. Labelling effect on cell division and differentiation was studied. Several dyes lost fluorescence and slowed cell division. However, the thiol reactive dye CM-DiI showed detectable in vivo fluorescence in labelled MSC six weeks after injection into sheep skeletal muscle and two weeks after implantation of an MSC coated biomaterial scaffold. CM-DiI labelled MSC differentiated in vitro showed label retention over four weeks. The fluorescent membrane dye CM-DiI tracks implanted sheep MSC and provides an alternative to traditional cell markers such as gene modified GFP.

Environmental stress and vestibular inputs modulate cardiovascular responses to orthostasis in hypertensive rats.

The frequent accompaniment of hypertension by orthostatic circulatory disorders prompted us to investigate the effect of repeated and sustained head-up and head-down tilt positions on cardiovascular responses in spontaneously hypertensive rats vs. Wistar rats using radiotelemetric implants. Repeated orthostasis caused a transient elevation in blood pressure (7.3±1.7 mmHg) and heart rate (39.7±10.5 BPM), while repeated antiorthostasis led only to reversible tachycardia (85.6±11.7-54.3±16.8 BPM) in spontaneously hypertensive rats. In contrast to the Wistar rats, sustained tilt failed to affect the blood pressure or heart rate in spontaneously hypertensive rats because the environmental stress of being placed in horizontal tilt cages prior to the sustained tilt test induced marked changes in cardiovascular parameters. Non-specific stress responses were eliminated both by the anxiolytic diazepam and a sub-anesthetic dose of chloralose. Unlike diazepam, chloralose amplified the orthostatic pressor responses in the Wistar rats. In contrast to diazepam preventing the pressor response and associated tachycardia in spontaneously hypertensive rats, chloralose elicited this effect during both sustained orthostasis (36.0±7.3 mmHg, 63.7±21.8 BPM) and antiorthostasis (42.9±10.9 mmHg, 82.8±25.4 BPM), with a reduced baroreflex sensitivity. However, during sustained orthostasis, removal of the vestibular input led to a depressor response with bradycardia (12.5±3.2 mmHg, 59.3±17.3 BPM), whereas antiorthostasis only reduced blood pressure (20.5±7.1 mmHg) in the spontaneously hypertensive rats. We conclude that repeated tilts induce a transient pressor response and/or tachycardia in spontaneously hypertensive rats. Cardiovascular parameters are suppressed by diazepam, whereas chloralose evokes both blood pressure and heart rate responses during sustained tilts, which are primarily elicited by baroreflex suppression in hypertension. Vestibular inputs support cardiovascular tolerance to sustained postural changes in a rat model of human 'essential' hypertension.

Myocardial extracellular matrix remodeling in ischemic heart failure.

Left ventricular (LV) remodeling is a process whereby structural alterations attempt to compensate altered hemodynamic load. In the chronic setting this process becomes maladaptive, self-sustaining and is associated with worsened survival. The extracellular matrix (ECM) of the heart, once believed an inert scaffold for cardiomyocytes, is now known to play an important role in LV remodeling. The enzyme system primarily responsible for ECM turnover is the matrix metalloproteinases (MMPs), and these enzymes are robustly altered in cardiovascular pathologies, including myocardial infarction (MI) and ischemic heart failure. A cause-and-effect relationship has been established between MMPs and LV remodeling post MI, as MMP inhibition prevents LV dilation and preserves cardiac function in animal models of infarction. In spite of this, initial clinical experience with MMP inhibition post MI has been disappointing. This review examines the structural and functional roles of the myocardial ECM, the evidence for MMP involvement in LV remodeling, and recent investigations into MMPs as prognostic markers and therapeutic targets.

Cardiomyoplasty reduces myocardial oxygen consumption: implications for direct mechanical compression.

BACKGROUND: This study investigates the possibility of reducing myocardial oxygen consumption by dynamic cardiomyoplasty in chronic heart failure. The sheep model used is relevant for cardiac assist using direct mechanical cardiac compression. METHODS: In 7 sheep, heart failure was induced by staged intracoronary microembolization followed by dynamic cardiomyoplasty. Six months later, the effect of latissimus dorsi muscle stimulation in the 2:1 mode (on, cardiomyoplasty; off, control) was studied. Left ventricular pressure-volume loops were obtained by conductance, micromanometer, and inferior vena cava occlusion catheter. Myocardial oxygen consumption was derived from left main coronary artery blood flow and oxygen content of arterial and coronary sinus blood. RESULTS: Cardiomyoplasty had no significant effect on left ventricular hemodynamic variables such as end-systolic pressure. However, cardiomyoplasty increased stroke volume and ejection fraction significantly by 11% +/- 12% and 11% +/- 10%, respectively. Although pressure-volume area and external work did not increase with cardiomyoplasty, myocardial oxygen consumption decreased by 21% +/- 11%. Therefore, cardiomyoplasty increased myocardial efficiency (external work/myocardial oxygen consumption) by 16% +/- 13%. CONCLUSIONS: Despite limited hemodynamic improvement from dynamic cardiac compression by cardiomyoplasty in sheep with chronic heart failure, myocardial oxygen consumption was significantly reduced. These findings provide a rationale for reverse remodeling of the failing heart using direct mechanical compression.

Functional recovery of the native heart after cardiomyoplasty in sheep with heart failure: passive and dynamic effects of volume loading.

BACKGROUND: Dynamic cardiomyoplasty (d-CMP) encourages reverse remodeling and improved contractility and stroke work (SW) efficiency of the failing native heart. This contrasts with passive cardiomyoplasty (p-CMP), which provides "passive girdling." To further evaluate pump recovery we assessed native left ventricular performance (without assist) 6 months after dynamic and passive CMP in sheep with heart failure with acute volume loading. METHODS: Heart failure (left ventricular ejection fraction 26%+/-8%) induced by coronary microembolization was followed by CMP in 11 sheep. After 8 weeks of muscle "training," paced cardiac assist was undertaken in the d-CMP group (n = 6). Five sheep with heart failure served as controls. Six months later the pressure-volume relationship was derived before and after volume loading by colloid solution. Latissimus dorsi muscle pacing was previously ceased in the d-CMP group. RESULTS: Volume loading increased left ventricular end-diastolic volume and pressure in all groups. After volume loading in d-CMP, the SW and pressure-volume area were increased, and SW efficiency remained unchanged. In p-CMP neither variable changed, whereas in control heart failure SW efficiency decreased due to a rise in pressure-volume area with stable SW. CONCLUSIONS: Based on response to volume loading, the failing native heart after 6 months of d-CMP showed functional recovery from "active girdling," whereas p-CMP prevented functional deterioration through passive girdling. The failing control heart progressively deteriorated.

Glucose-insulin-potassium solution improves left ventricular mechanics in diabetes.

BACKGROUND: The mechanism by which glucose-insulin-potassium solutions enhance recovery of left ventricular function after myocardial ischemia in diabetic patients is not well understood. We evaluated the effect of glucose-insulin-potassium on ventriculoarterial coupling and left ventricular mechanics in a chronic ovine model of diabetes. METHODS: Diabetes was induced in 6 sheep with streptozotocin. After 6 months of diabetes, the response of the left ventricular pressure-volume relationship to 60 minutes of intravenous glucose-insulin-potassium solution (1,000 mL of 5% dextrose in water, 100 IU of regular insulin, 90 mmol of KCl at 1.5 mL x kg(-1) x h(-1)) was determined. RESULTS: Glucose-insulin-potassium solution increased end-systolic elastance 68% (p = 0.01) and improved ventriculoarterial coupling (1.7+/-0.3 to 1.0+/-0.1; p < 0.01). Potential energy decreased 35% (p = 0.01), and pressure-volume area decreased 20% (p = 0.01). However, stroke work did not change; therefore stroke work efficiency increased from 50.1%+/-3.5% to 60.2%+/-5.1% (p = 0.01). CONCLUSIONS: Glucose-insulin-potassium solution improves left ventricular contractility and ventriculoarterial coupling in diabetes. Left ventricular mechanics is improved by decreasing total mechanical work without significantly affecting stroke work, resulting in improved stroke work efficiency. Improved efficiency facilitates understanding of the enhanced tolerance to myocardial ischemia afforded by glucose-insulin-potassium solution.

Glucose-insulin-potassium solution improves left ventricular energetics in chronic ovine diabetes.

BACKGROUND: Therapeutic modulation of myocardial metabolism improves outcomes in diabetic patients following myocardial infarction and coronary artery surgery. However, the mechanism of this beneficial effect has not been fully elucidated. This study evaluated the effect of glucose-insulin-potassium solution (GIK) on left ventricular (LV) energetics and oxygen utilization efficiency in a chronic ovine model of diabetes. METHODS: Diabetes was induced in sheep with streptozotocin. Experiments were performed following 12 months untreated diabetes (n = 6) and in controls (n = 6). Open-chest anesthetized sheep were instrumented to determine the LV pressure-volume relationship, oxygen consumption, and free fatty acid uptake. Glucose-insulin-potassium was infused at 1.5 mL x kg(-1) x h(-1) for 60 minutes and assessment repeated. RESULTS: Glucose-insulin-potassium decreased LV free fatty acid uptake in control: 0.090 +/- 0.047 microg/beat/100 g to 0.024 +/- 0.022 microg/beat/100 g, p = 0.02 and diabetes: 0.33 +/- 0.32 microg/beat/100 g to 0.11 +/- 0.13 microg/beat/100 g, p = 0.04. Similarly, GIK decreased unloaded left ventricular oxygen consumption (LVVO(2)) in both control (0.42 +/- 0.05 to 0.37 +/- 0.13J/beat/100 g, p < 0.001) and diabetic sheep (0.40 +/- 0.24 to 0.23 +/- 0.23J/beat/100 g, p < 0.001). The slope of the LVVO(2)-pressure-volume area relation (contractile efficiency) was unchanged in either group. Glucose-insulin-potassium improved LV contractility 58% +/- 37% (p = 0.005) and stroke work efficiency 18% +/- 10% (p = 0.009) in diabetic animals but not controls. Therefore, oxygen utilization efficiency (stroke work-LVVO(2)) increased only in diabetic animals (16.6% +/- 4.8% to 26.9% +/- 3.6%, p = 0.002) following GIK. CONCLUSIONS: This study provides in vivo evidence that GIK improves LV energetics in diabetes. Oxygen utilization efficiency is improved as a result of improved stroke work efficiency and decreased unloaded LVVO(2). Improved efficiency of oxygen utilization provides a physiologic rationale for the beneficial effect of GIK in diabetic patients.

Direct compression of the failing heart reestablishes maximal mechanical efficiency.

BACKGROUND: In failing hearts, homeostatic mechanisms contrive to maximize stroke work and maintain normal arterial blood pressure at the expense of energetic efficiency. In contrast dobutamine reestablishes maximal mechanical efficiency by promoting energetically optimal loading conditions. However, dobutamine also wastefully increases nonmechanical oxygen consumption. We investigated whether direct mechanical cardiac compression would reestablish maximal mechanical efficiency without the oxygen-wasting effect. METHODS: The pressure-volume relationship and myocardial oxygen consumption were derived in sheep using left ventricular pressure and volume from manometer-tipped and conductance catheters, and coronary flow from Transonics flow probe. RESULTS: Propranolol hydrochloride and atropine sulfate were administered to reduce ejection fraction to 21% when ventricular elastance fell to 1.35 mm Hg/mL and mechanical efficiency to 79% of maximal. Low-pressure direct mechanical compression of the failing heart restored mechanical efficiency to 94% of maximal and realigned optimal left ventricular end-systolic pressure with operating left ventricular end-systolic pressure without altering nonmechanical oxygen consumption. CONCLUSIONS: We conclude that direct cardiac compression restores mechanical efficiency to normal maximum without wasting energy on additional nonmechanical activity.

HeartPatch implanted direct cardiac compression: effect on coronary flow and flow patterns in acute heart failure sheep.

A novel HeartPatch direct cardiac compression (DCC) device has been shown to effectively restore circulatory parameters in sheep with acute heart failure (HF). Its effect on the coronary circulation and myocardial perfusion, however, remains uncertain. The effect of DCC assist on coronary artery blood flow (CABF) and its patterns in acute HF sheep were examined in this study. Ten sheep (51 +/- 6 kg) were implanted with a heart patch on each of the left ventricular and right ventricular free walls 1 week before study. Stable HF [cardiac output (CO) at 51 +/- 8% of baseline] induced by intravenous esmolol resulted in CABF decreasing to 53 +/- 16% of baseline (p < 0.001). DCC device activation did not alter CABF (54 +/- 15% of baseline, N.S.) but was accompanied by increases in both peak antegrade and retrograde flow velocity (161 +/- 75%, p < 0.001 and 413 +/- 377%, p < 0.001). A shift in the proportion of flow occurring in diastole (%DF) also was observed: baseline, 81 +/- 9%; HF, 82 +/- 6%; DCC assist, 121 +/- 16% (p < 0.001). Despite significant changes in coronary artery flow pattern resulting from DCC of the failing heart, total antegrade coronary flow was maintained. These findings suggest that myocardial perfusion is not compromised by DCC.

Selective inhibition of the master regulator transcription factor Egr-1 with catalytic oligonucleotides reduces myocardial injury and improves left ventricular systolic function in a preclinical model of myocardial infarction.

BACKGROUND: Egr-1 is implicated in the pathogenesis of myocardial ischemia-reperfusion injury. The aim of this study was to ascertain the effectiveness of intracoronary delivery of DNAzyme targeting the transcription factor Egr-1 at reperfusion following experimental myocardial ischemia. METHODS AND RESULTS: Functional DNAzyme targeting Egr-1 or a size-matched scrambled control were delivered via the intracoronary route immediately on reperfusion after 60 minutes' balloon occlusion of the left anterior descending coronary artery in a pig model of myocardial I/R injury (n=7 per treatment group). Heart function and extent of myocardial infarction were determined following intervention by echocardiography and cardiac magnetic resonance imaging, respectively. Hearts were removed and examined for molecular and histological markers of inflammation and apoptosis. Administration of functional DNAzyme led to an overall decrease in the expression of inflammatory markers including intracellular adhesion molecule-1, tissue factor, and complement 3, with associated decreases in the extent of neutrophil infiltration, oxidative damage, and subsequent apoptosis within the infarct border zone. Functional significance was indicated by an increase in salvaged left ventricular myocardium (P=0.012), ejection fraction (P=0.002), and fractional area change (P=0.039) in the functional DNAzyme-treated group compared with the control. CONCLUSIONS: Egr-1 silencing through intracoronary delivery of a targeting DNAzyme at the time of reperfusion following acute myocardial ischemia decreases myocardial inflammation and apoptosis leading to improved cardiac function.

The Royal North Shore Hospital inquiry: an analysis of the recommendations and the implications for quality and safety in Australian public hospitals.

In October 2007, the New South Wales Parliament appointed a Joint Select Committee to inquire into the quality of patient care at Royal North Shore Hospital (RNSH). The inquiry was initiated in response to the publicity and complaints surrounding a patient who had a miscarriage in the toilets of the RNSH emergency department waiting area. The Committee held four public hearings and received 103 submissions. It handed down 45 recommendations in its report on 20 December 2007. There has been criticism from clinicians and others that the recommendations are too general and will not effect significant change for the severe systemic problems affecting the hospital. This article represents the view of some of the clinicians who work at RNSH, and who gave evidence at the inquiry, on the recommendations and some possible solutions for the health system in general.

Effect of direct cardiac compression on left ventricular axial dynamics in sheep.

The HeartPatch direct cardiac compression device consists of two separate, nonsurround patches placed on the left and right ventricular free walls. Although the device has been shown to effectively restore circulatory parameters in acute heart failure sheep, the impact of device inflation on left ventricular geometry is yet to be elucidated. This study used sonomicrometer crystal transducers to examine three orthogonal left ventricular dimensions under various cardiac states and assessed the feasibility of determining stroke volume from these dimensions. Seven sheep (weight, 51 +/- 5 kg) were implanted with six sonomicrometer crystals, and a heart patch was placed on each of the ventricles. The crystals were positioned to measure anterior-posterior, septal-lateral, and apex-base (long-axis) dimensions. Sheep were studied under both awake and anesthetized conditions. Septal-lateral shortening was increased with direct cardiac compression assist, whereas anterior-posterior and long-axis dimensions were either unchanged (awake) or decreased (anesthetized). Estimation of stroke volume, using the ellipsoid volume model, correlated well with stroke volume measured from an aortic flow probe; however, absolute stroke volumes were lacking in agreement.

Efficacy and mechanisms of biventricular and left/right direct cardiac compression in acute heart failure sheep.

Direct cardiac compression (DCC) with implanted heart patches has previously demonstrated efficacy of biventricular (BiV) support in acute heart failure (HF) sheep. We hypothesized that this was primarily due to a left ventricular (LV) effect. This study compared BiV, LV, and right ventricular (RV) assists in terms of hemodynamic and energetic response. Ten sheep underwent instrumentation and device implantation at least 1 week prior to study. HF (50% reduction in cardiac output) was maintained with intravenous esmolol infusion. BiV, LV, and RV assists were activated randomly with intervening stable HF periods. BiV assist was more effective than either LV or RV assist in restoring hemodynamic parameters; however, there was no difference in efficacy of LV and RV support. RV assist preserved left coronary flow patterns and chamber geometry compared to other assist conditions, but increased LV preload. These results suggest that LV and RV support each make a significant contribution to the efficacy of BiV assist, albeit through different mechanisms.

Regional cardiac dysfunction is associated with specific alterations in inflammatory cytokines and matrix metalloproteinases after acute myocardial infarction in sheep.

Cardiac remodeling following myocardial infarction (MI) is a maladaptive process, fundamental to the progression of ischemic heart failure. The extent of remodeling is influenced by mechanical stress, inflammatory response and activation of matrix metalloproteinases (MMPs). This study examined regional association between these parameters in response to acute MI. Coronary ligation was performed in ten sheep. Sonomicrometer transducers measured segmental length in the infarcted, border and non-infarcted region. Regional tissue samples obtained 3 h post MI from six sheep were analysed using RT-PCR, gelatin zymography and Western blot. Six sham-operated sheep served as controls.Region-specific dilation and reduced contraction was associated with corresponding alterations in matrix molecules.IL-6 and MMP-9 mRNA were increased in the infarcted and border regions compared to controls.MMP-2 and TIMP-1 mRNA increased in non-infarcted myocardium and both correlated positively with segmental shortening. IL-6 mRNA levels, in contrast, were negatively associated with segmental shortening. In summary, inflammatory cytokines and MMPs are altered early after MI in a region-specific manner, and these changes correspond to acute regional myocardial dysfunction. Therapies for LV remodeling from the time of reperfusion may benefit from further understanding this portfolio of acute alterations.

Postural effects when cycling in late pregnancy.

AIM: This study assessed if upright cycling is preferable to semi-recumbent cycling during pregnancy. METHOD: Healthy women with low risk singleton pregnancies were tested at 34-38 weeks gestation. They cycled for 12 min, either semi-recumbent (45 degrees, n = 27) or upright (n = 23), at 135-145 beats min(-1). RESULTS: When semi-recumbent, minute ventilation was greater (p<0.03) at rest and systolic blood pressure and pulse pressure were greater during exercise (p<0.05). Exercise maternal heart rate, oxygen consumption, oxygen consumption per kilogram, minute ventilation, cardiac output, stroke volume, mean and diastolic blood pressures and arterio-venous oxygen difference were posture-independent. All increased with exercise (p<0.01), except stroke volume when semi-recumbent (p>0.05). Small post-exercise fetal heart rate increases (by 8 beats min(-1), p<0.05) were similar in both postures (n = 11 in each sub-group), with no adverse changes. Fetal heart rate accelerations and uterine activity (n = 11 in each sub-group) were not influenced by posture or exercise. CONCLUSIONS: (1) Neither posture had a distinct advantage. (2) Both postures were safe for short duration cycling. (3) The same target maternal heart rates are suitable for both postures because they resulted in similar oxygen consumptions and fetal heart rates.

Remodeling of the chronic severely failing ischemic sheep heart after coronary microembolization: functional, energetic, structural, and cellular responses.

The mandatory use of pharmacotherapy in human heart failure (HF) impedes further study of natural history and remodeling mechanisms. We created a sheep model of chronic, severe, ischemic HF [left ventricular (LV) ejection fraction (LVEF) <35% stable over 4 wk] by selective coronary microembolization under general anesthesia and followed hemodynamic, energetic, neurohumoral, structural, and cellular responses over 6 mo. Thirty-eight sheep were induced into HF (58% success), with 23 sheep followed for 6 mo (21 sheep with sufficient data for analysis) after the LVEF stabilized (median of 3 embolizations). Early doubling of LV end-diastolic pressure persisted, as did increases in LV end-diastolic volume, LV wall stress, and LV wall thinning. Contractile impairment (LV end-systolic elastance, LV preload recruitable stroke work, and dobutamine-responsive contractile reserve) and diastolic dysfunction also remained stable. Cardiac mechanical energy efficiency did not recover. Plasma atrial natriuretic peptide levels remained elevated, but rises in plasma aldosterone and renin activity were transient. Collagen content increased 170%, the type I-to-III phenotype ratio doubled in the LV, but right ventricular collagen remained unaltered. Fas ligand cytokine levels correlated with expression of both caspase-3 and -2, suggesting a link in the apoptotic "death cascade." Caspase-3 activity also bore a close relationship to LV meridional wall stress calculated from echocardiographic and intraventricular pressure measurements. We concluded that the stability of chronic untreated severe ischemic HF depends on the recruitment of myocardial remodeling mechanisms that involve an interaction among hemodynamic load, contractile efficiency/energetics, neurohumoral activation, response of the extracellular matrix, wall stress, and the myocyte apoptotic pathway.