Radiofrequency ablation lesions in low-, intermediate-, and normal-voltage myocardium: an in vivo study in a porcine heart model.

AIMS: Contact force (CF) between radiofrequency (RF) ablation catheter and myocardium and ablation index (AI) correlates with RF lesion depth and width in normal-voltage (>1.5 mV) myocardium (NVM). We investigate the impact of CF on RF lesion depth and width in low (<0.5 mV) (LVM) and intermediate-voltage (0.5-1.5 mV) myocardium (IVM) following myocardial infarction. Correlation between RF lesion depth and width evaluated by native contrast magnetic resonance imaging (ncMRI) and gross anatomical evaluation was investigated. METHODS AND RESULTS: Twelve weeks after myocardial infarction, 10 pigs underwent electroanatomical mapping and endocardial RF ablations were deployed in NVM, IVM, and LVM myocardium. In vivo ncMRI was performed before the heart was excised and subjected to gross anatomical evaluation. Ninety (82%) RF lesions were evaluated. Radiofrequency lesion depth and width were smaller in IVM and LVM compared with NVM (P < 0.001). Radiofrequency lesion depth and width correlated with CF, AI, and impedance drop in NVM (CF and AI P < 0.001) and IVM (CF and AI depths P < 0.001; CF and AI widths P < 0.05). Native contrast magnetic resonance imaging evaluated RF lesion depth and width correlated with gross anatomical depth and width (NVM and IVM P < 0.001; LVM P < 0.05). CONCLUSIONS: Radiofrequency lesions deployed by similar duration, power and CF are smaller in IVM and LVM than in NVM. Radiofrequency lesion depth and width correlated with CF, AI, and impedance drop in NVM and IVM but not in LVM. Native contrast magnetic resonance imaging may be useful to assess RF lesion depth and width in NVM, IVM, and LVM.

Identification of the building blocks of ventricular septation in monitor lizards (Varanidae).

Among lizards, only monitor lizards (Varanidae) have a functionally divided cardiac ventricle. The division results from the combined function of three partial septa, which may be homologous to the ventricular septum of mammals and archosaurs. We show in developing monitors that two septa, the 'muscular ridge' and 'bulbuslamelle', express the evolutionarily conserved transcription factors Tbx5, Irx1 and Irx2, orthologues of which mark the mammalian ventricular septum. Compaction of embryonic trabeculae contributes to the formation of these septa. The septa are positioned, however, to the right of the atrioventricular junction and they do not participate in the separation of incoming atrial blood streams. That separation is accomplished by the 'vertical septum', which expresses Tbx3 and Tbx5 and orchestrates the formation of the electrical conduction axis embedded in the ventricular septum. These expression patterns are more pronounced in monitors than in other lizards, and are associated with a deep electrical activation near the vertical septum, in contrast to the primitive base-to-apex activation of other lizards. We conclude that evolutionarily conserved transcriptional programmes may underlie the formation of the ventricular septa of monitors.

Morpho-functional characterization of the systemic venous pole of the reptile heart.

Mammals evolved from reptile-like ancestors, and while the mammalian heart is driven by a distinct sinus node, a sinus node is not apparent in reptiles. We characterized the myocardial systemic venous pole, the sinus venosus, in reptiles to identify the dominant pacemaker and to assess whether the sinus venosus remodels and adopts an atrium-like phenotype as observed in mammals. Anolis lizards had an extensive sinus venosus of myocardium expressing Tbx18. A small sub-population of cells encircling the sinuatrial junction expressed Isl1, Bmp2, Tbx3, and Hcn4, homologues of genes marking the mammalian sinus node. Electrical mapping showed that hearts of Anolis lizards and Python snakes were driven from the sinuatrial junction. The electrical impulse was delayed between the sinus venosus and the right atrium, allowing the sinus venosus to contract and aid right atrial filling. In proximity of the systemic veins, the Anolis sinus venosus expressed markers of the atrial phenotype Nkx2-5 and Gja5. In conclusion, the reptile heart is driven by a pacemaker region with an expression signature similar to that of the immature sinus node of mammals. Unlike mammals, reptiles maintain a sinuatrial delay of the impulse, allowing the partly atrialized sinus venosus to function as a chamber.

Evaluation of cardiac electrophysiological properties in an experimental model of right ventricular hypertrophy and failure.

BACKGROUND: Malignant arrhythmias are a major cause of sudden cardiac death in adults with congenital heart disease. We developed a model to serially investigate electrophysiological properties in an animal model of right ventricular hypertrophy and failure. METHOD: We created models of compensated (cHF; n=11) and decompensated (dHF; n=11) right ventricular failure in Wistar rats by pulmonary trunk banding. Healthy controls underwent sham operation (Control; n=13). Surface electrocardiography was recorded from extremities, and inducibility of ventricular tachycardia was evaluated in vivo by programmed stimulation. Isolated right ventricular myocardium was analysed for mRNA expression of selected genes. RESULTS: Banding caused an increased mRNA expression of both connexin 43 and the voltage-gated sodium channel 1.5, as well as a prolongation of PQ, QRS and QTc intervals. Ventricular tachycardia was induced in the majority of banded animals compared with none in the healthy control group. No differences were found between the two degrees of failure in neither the electrophysiological parameters nor inducibility. CONCLUSIONS: The electrophysiological properties of rat hearts subjected to pulmonary trunk banding were significantly changed with increased susceptibility to ventricular tachycardia, but no differences were found between compensated and decompensated right ventricular failure. Furthermore, we demonstrate that in vivo electrophysiological evaluation is a sensitive method to characterise the cardiac electric phenotype in an experimental rat model.

Levosimendan Prevents Pressure-Overload-induced Right Ventricular Failure.

BACKGROUND: We investigated if chronic levosimendan treatment can prevent and revert pressure-overload-induced right ventricular hypertrophy and failure in rats. METHODS: Right ventricular hypertrophy and failure was induced in Wistar rats by pulmonary trunk banding (PTB). The PTB rats were treated with levosimendan (3 mg·kg·d) 3 days before surgery [n = 10, prevention (PREV)], 3 weeks after surgery [n = 10, reversal (REV)] or vehicle (n = 10, VEH). Sham-operated rats received vehicle (n = 16, SHAM). Right ventricular function was evaluated 7 weeks after surgery by echocardiography, magnetic resonance imaging, pressure-volume relations, gross anatomy, and histology. RESULTS: PTB induced right ventricular hypertrophy and compensated heart failure evident by reduced cardiac index (CI) without extra cardiac signs of heart failure. Levosimendan treatment prevented deterioration of right ventricular function measured by CI and right ventricular ejection fraction (RVEF) (CI: VEH vs. PREV 281 ± 17 vs. 362 ± 34 mL·min·kg, P ≤ 0.05, RVEF: VEH vs. PREV 57 ± 2% vs. 68 ± 3%, P ≤ 0.01) to values similar to SHAM (CI: 345 ± 21 mL·min·kg, RVEF: 71 ± 2%). RV contractility was improved in the REV group measured by preload recruitable stroke work (VEH vs. REV 39 ± 3 vs. 66 ± 10 mmHg P ≤ 0.05). CONCLUSIONS: Chronic treatment with levosimendan prevents the development of right ventricular failure and improves contractility in established pressure-overload-induced right ventricular failure.

Effects of bisoprolol and losartan treatment in the hypertrophic and failing right heart.

BACKGROUND: Sympathetic adrenergic stimulation and the renin-angiotensin-aldosterone system are highly elevated in right heart failure. We evaluated if treatment with the adrenergic receptor blocker bisoprolol or the angiotensin II receptor blocker losartan could prevent the progression of right ventricular (RV) hypertrophy and failure in rats after pulmonary trunk banding (PTB). METHODS AND RESULTS: Male Wistar rats were randomized to severe PTB with a 0.5-mm banding clip (PTB0.5, n = 29), moderate PTB with a 0.6-mm banding clip (PTB0.6, n = 28), or sham operation (SHAM, n = 13). The PTB0.5 and PTB0.6 rats were randomized to 6 weeks of 10 mg/kg/d bisoprolol treatment, 20 mg/kg/d losartan treatment, or vehicle treatment. The PTB caused hypertrophy, dilation, and reduced function of the RV in all rats subjected to the procedure. Rats subjected to the more severe banding developed decompensated RV failure with extracardiac manifestations. Treatment with bisoprolol slowed the heart rate, and treatment with losartan lowered mean arterial pressure, confirming adequate dosing, but none of the treatments improved RV function or arrested the progression of RV hypertrophy and failure compared with vehicle. CONCLUSIONS: In our PTB model of pressure overload-induced RV hypertrophy and failure, treatment with bisoprolol and losartan did not demonstrate any beneficial effects in compensated or decompensated RV failure.

Remote cardioprotection by transfer of coronary effluent from ischemic preconditioned rabbit heart preserves mitochondrial integrity and function via adenosine receptor activation.

BACKGROUND: Coronary effluent from an isolated perfused heart undergoing ischemic preconditioning can be transferred to precondition another naïve isolated heart. We investigated the effects of this effluent on mitochondrial integrity and function following a global infarct model of ischemia/reperfusion and the role of adenosine in this model of remote preconditioning. METHODS AND RESULTS: Coronary effluent from isolated perfused rabbit hearts was collected prior to (control effluent) and during three cycles of 5-min ischemia and 10-min reperfusion (IPC effluent). Adenosine concentration was significantly increased in IPC effluent (2.6 ± 1.1 µM) versus control effluent (0.21 ± 0.06 µM, P < 0.01). Infarct size (% necrotic LV mass) after 30-min global ischemia and 90-min reperfusion was significantly reduced in hearts preconditioned with IPC effluent (IPC(eff), 23 ± 7 %) and control effluent supplemented with 2.5 µM exogenous adenosine (C(eff)+ 2.5 µM ADO, 25 ± 10 %) when compared to control effluent perfused hearts (C(eff), 41 ± 8 %, P < 0.05). Compared to C(eff) mitochondria, IPC(eff) mitochondria had preserved complex I/State3 and complex IV/State 3 respiration and outer membrane integrity, and reduced cytochrome c release. In contrast, C(eff) + 2.5 µM ADO mitochondria had improved state 2 respiration and coupling to oxidative phosphorylation, reduced reactive oxygen species production and preserved outer membrane integrity. Administration of adenosine receptor blocker 8-(p-sulfophenyl)theophylline abolished the infarct limiting effect (46 ± 7 %) and the mitochondrial integrity and function preservation of IPC effluent. CONCLUSION: Remote cardioprotection by IPC effluent preserves mitochondrial integrity and function in an adenosine receptor dependent mechanism, and although infarct size reduction can be mimicked by adenosine, IPC effluent contains additional factor(s) contributing to modulation of the mitochondrial response to ischemia/reperfusion injury.

Acute effects of levosimendan in experimental models of right ventricular hypertrophy and failure.

Pulmonary arterial hypertension (PAH) is a fatal disease, and the ultimate cause of death is right ventricular (RV) failure. In this study, we investigated the acute hemodynamic effects of levosimendan in two rat models of RV hypertrophy and failure. Wistar rats were randomized to receive sham surgery (n = 8), pulmonary trunk banding (PTB; n = 8), or monocrotaline injection (MCT; n = 7). RV function was evaluated at baseline and after injection of placebo and two concentrations of levosimendan (12 and 60 µg/kg) using magnetic resonance imaging, echocardiography, and invasive pressure recordings. PTB and MCT injection caused hypertrophy, dilatation, and failure of the RV compared with sham surgery. Levosimendan increased RV end systolic pressure (sham surgery: 16.0% ± 3.8% [P = 0.0038]; MCT: 9.9% ± 3.1% [P = 0.018]; PTB: 24.5% ± 3.3% [P = 0.0001]; mean ± SEM) compared with placebo. Levosimendan markedly increased RV stroke volume (SV) in the MCT group (29.1% ± 8.3%; P = 0.012), did not change RV SV in the PTB group (0.4% ± 4.5%; P = 0.93), and decreased RV SV in the sham surgery group (-10.9% ± 3.7%; P = 0.020). Nitroprusside, which was used to mimic the systemic arterial vasodilator action of levosimendan, did not influence RV function. These data demonstrate that levosimendan acutely improves the failing right heart in a MCT model of PAH and that the mechanism involves a direct acute positive inotropic effect on the hypertrophic and failing RV of the rat.

The effects of cyclic guanylate cyclase stimulation on right ventricular hypertrophy and failure alone and in combination with phosphodiesterase-5 inhibition.

BACKGROUND: We investigated if soluble guanylate cyclase stimulation either alone or in combination with phosphodiesterase-5 (PDE5) inhibition could prevent pressure overload-induced right ventricular (RV) hypertrophy and failure. METHODS: The soluble guanylate cyclase stimulator BAY 41-2272 (BAY, 10 mg · kg⁻¹ · d⁻¹) either alone or in combination (BAY + SIL) with a PDE5 inhibitor sildenafil (SIL, 100 mg · kg⁻¹ · d⁻¹) was examined for prevention of RV hypertrophy and failure in Wistar rats (n = 73) operated by pulmonary trunk banding. RESULTS: All treatments failed to inhibit the development of RV hypertrophy and failure. In the BAY and BAY + SIL groups, there was an increased mortality. Mean arterial blood pressure was lowered and cardiac output increased in the BAY + SIL group. Systolic RV pressure was increased in the BAY and BAY + SIL groups possibly because of an inotropic response and/or increased venous return. CONCLUSIONS: Stimulation of soluble guanylate cyclase by BAY 41-2272 alone or in combination with sildenafil failed to prevent the development of RV hypertrophy and failure in rats subjected to pulmonary trunk banding. An increased mortality was observed in animals treated by BAY 41-2272 alone and in combination with sildenafil.

Iloprost improves ventricular function in the hypertrophic and functionally impaired right heart by direct stimulation.

Right heart function is an important predictor of morbidity and mortality in patients suffering from pulmonary arterial hypertension and congenital heart diseases. We investigated whether the prostacyclin analog iloprost has a direct inotropic effect in the pressure-overloaded hypertrophic and dysfunctional right ventricle (RV). Rats were randomized to monocrotaline injection (60 mg/kg; [Formula: see text]), pulmonary trunk banding (PTB; [Formula: see text]), or a sham operation ([Formula: see text]). RV function was evaluated with magnetic resonance imaging, echocardiography, and invasive pressure measurements at baseline, after intravenous administration of placebo, iloprost 10 ng/kg/min, or iloprost 100 ng/kg/min (Ilo100). Infusion of Ilo100 induced a [Formula: see text] ([Formula: see text]) increase in stroke volume in the sham group and a [Formula: see text] ([Formula: see text]) increase in the PTB group. RV [Formula: see text] was elevated by [Formula: see text] ([Formula: see text]) in the sham group and by [Formula: see text] ([Formula: see text]) in the PTB group. An elevation in cardiac output of [Formula: see text] ([Formula: see text]) and an [Formula: see text] ([Formula: see text]) increase in RV systolic pressure were found in the PTB group. Iloprost caused a decrease in mean arterial blood pressure (MAP) in all groups of animals. An equal reduction in MAP induced by the arterial vasodilator nitroprusside did not improve any of the measured parameters of RV function. We conclude that iloprost has inotropic properties directly improving ventricular function in the hypertrophic and dysfunctional right heart of the rat.

Acute effects of sildenafil and dobutamine in the hypertrophic and failing right heart in vivo.

Abstract The purpose of this study was to investigate whether acute intravenous administration of the phosphodiesterase type 5 (PDE-5) inhibitor sildenafil in a single clinically relevant dose improves the in vivo function of the hypertrophic and failing right ventricle (RV). Wistar rats ([Formula: see text]) were subjected to pulmonary trunk banding (PTB) causing RV hypertrophy and failure. Four weeks after surgery, they were randomized to receive an intravenous bolus dose of sildenafil (1 mg/kg; [Formula: see text]), vehicle ([Formula: see text]), or dobutamine (10 µg/kg; [Formula: see text]). Invasive RV pressures were recorded continuously, and transthoracic echocardiography was performed 1, 5, 15, 25, 35, 50, 70, and 90 minutes after injecting the bolus. Cardiac function was compared to baseline measurements to evaluate the in vivo effects of each specific treatment. The PTB procedure caused significant hypertrophy, cardiac fibrosis, and reduction in RV function evaluated by echocardiography (TAPSE) and invasive pressure measurements. Sildenafil did not improve the function of the hypertrophic failing right heart in vivo, measured by TAPSE, RV systolic pressure (RVsP), and dp/dtmax. Dobutamine improved RV function 1 minute after injection measured by TAPSE ([Formula: see text] vs. [Formula: see text] cm; [Formula: see text]), RVsP ([Formula: see text] vs. [Formula: see text] mmHg; [Formula: see text]), and dp/dtmax ([Formula: see text] vs. [Formula: see text] mmHg/s; [Formula: see text]). Acute administration of the PDE-5 inhibitor sildenafil in a single clinically relevant dose did not modulate the in vivo function of the hypertrophic failing right heart of the rat measured by echocardiography and invasive hemodynamics. In the same model, dobutamine acutely improved RV function.

Pulmonary gene silencing in transgenic EGFP mice using aerosolised chitosan/siRNA nanoparticles.

PURPOSE: This work describes the production and application of an aerosolised formulation of chitosan nanoparticles for improved pulmonary siRNA delivery and gene silencing in mice. METHODS: Aerosolised chitosan/siRNA nanoparticles were pneumatically formed using a nebulising catheter and sized by laser diffraction. In vitro silencing of aerosolised and non-aerosolised formulations was evaluated in an EGFP endogenous-expressing H1299 cell line by flow cytometry. Non-invasive intratracheal insertion of the catheter was used to study nanoparticle deposition by histological detection of Cy3-labeled siRNA and gene silencing in transgenic EGFP mouse lungs using a flow cytometric method. RESULTS: Flow cytometric analysis demonstrated minimal alteration in gene silencing efficiency before (68%) and after (62%) aerosolisation in EGFP-expressing H1299 cells. Intratracheal catheter administration in mice resulted in nanoparticle deposition throughout the entire lung in both alveoli and bronchiolar regions using low amounts of siRNA. Transgenic EGFP mice dosed with the aerosolised nanoparticle formulation showed significant EGFP gene silencing (68% reduction compared to mismatch group). CONCLUSIONS: This work provides a technology platform for effective pulmonary delivery and gene silencing of RNAi therapeutics with potential use in preclinical studies of respiratory disease treatment.

How the python heart separates pulmonary and systemic blood pressures and blood flows.

The multiple convergent evolution of high systemic blood pressure among terrestrial vertebrates has always been accompanied by lowered pulmonary pressure. In mammals, birds and crocodilians, this cardiac separation of pressures relies on the complete division of the right and left ventricles by a complete ventricular septum. However, the anatomy of the ventricle of most reptiles does not allow for complete anatomical division, but the hearts of pythons and varanid lizards can produce high systemic blood pressure while keeping the pulmonary blood pressure low. It is also known that these two groups of reptiles are characterised by low magnitudes of cardiac shunts. Little, however, is known about the mechanisms that allow for this pressure separation. Here we provide a description of cardiac structures and intracardiac events that have been revealed by ultrasonic measurements and angioscopy. Echocardiography revealed that the atrioventricular valves descend deep into the ventricle during ventricular filling and thereby greatly reduce the communication between the systemic (cavum arteriosum) and pulmonary (cavum pulmonale) ventricular chambers during diastole. Angioscopy and echocardiography showed how the two incomplete septa, the muscular ridge and the bulbuslamelle - ventricular structures common to all squamates - contract against each other in systole and provide functional division of the anatomically subdivided ventricle. Washout shunts are inevitable in the subdivided snake ventricle, but we show that the site of shunting, the cavum venosum, is very small throughout the cardiac cycle. It is concluded that the python ventricle is incapable of the pronounced and variable shunts of other snakes, because of its architecture and valvular mechanics.

Blockage of receptor for advanced glycation end products prevents development of cardiac dysfunction in db/db type 2 diabetic mice.

AIMS: Activation of the receptor for advanced glycation end products (RAGE) is associated with long-term complications in diabetes mellitus. In this study, we tested whether RAGE activation in the diabetic myocardium is implicated in the development of cardiac dysfunction. METHODS AND RESULTS: Using MRI and conductance catheter techniques, we evaluated cardiac function in a type 2 diabetic mouse model (db/db), and assessed the effect of blocking RAGE with a RAGE antibody. Gene expressions were evaluated in samples of myocardial tissue. Diabetic db/db mice demonstrated an accelerated age-dependent deterioration in cardiac function associated with altered expression of genes related to cardiac structure and function. Blockage of RAGE signalling prevented the reduction in systolic function (preload recruitable stroke work: 109.8 +/- 13.8 vs. 94.5 +/- 14.9 mmHg/microL, P = 0.04) and development of increased LV diastolic chamber stiffness (0.18 +/- 0.05 vs. 0.27 +/- 0.07 mmHg, P = 0.01). The cardiac expression of collagen (col1a1) was reduced by approximately 45% and the expression of myosin was switched from the foetal isoform (MHCbeta) to the adult isoform (MHCalpha). CONCLUSION: Activation of RAGE is a significant pathogenetic mechanism for the development of cardiac dysfunction in type 2 diabetes. The underlying mechanisms involve not only the passive biophysical properties of the myocardium but also myocyte function.

Cardiac expression of microsomal triglyceride transfer protein is increased in obesity and serves to attenuate cardiac triglyceride accumulation.

Obesity causes lipid accumulation in the heart and may lead to lipotoxic heart disease. Traditionally, the size of the cardiac triglyceride pool is thought to reflect the balance between uptake and beta-oxidation of fatty acids. However, triglycerides can also be exported from cardiomyocytes via secretion of apolipoproteinB-containing (apoB) lipoproteins. Lipoprotein formation depends on expression of microsomal triglyceride transfer protein (MTP); the mouse expresses two isoforms of MTP, A and B. Since many aspects of the link between obesity-induced cardiac disease and cardiac lipid metabolism remain unknown, we investigated how cardiac lipoprotein synthesis affects cardiac expression of triglyceride metabolism-controlling genes, insulin sensitivity, and function in obese mice. Heart-specific ablation of MTP-A in mice using Cre-loxP technology impaired upregulation of MTP expression in response to increased fatty acid availability during fasting and fat feeding. This resulted in cardiac triglyceride accumulation but unaffected cardiac insulin-stimulated glucose uptake. Long-term fat-feeding of male C57Bl/6 mice increased cardiac triglycerides, induced cardiac expression of triglyceride metabolism-controlling genes and attenuated heart function. Abolishing cardiac triglyceride accumulation in fat-fed mice by overexpression of an apoB transgene in the heart prevented the induction of triglyceride metabolism-controlling genes and improved heart function. The results suggest that in obesity, the physiological increase of cardiac MTP expression serves to attenuate cardiac triglyceride accumulation albeit without major effects on cardiac insulin sensitivity. Nevertheless, the data suggest that genetically increased lipoprotein secretion prevents development of obesity-induced lipotoxic heart disease.

Minimal engraftment of human CD34+ cells mobilized from healthy donors in the infarcted heart of athymic nude rats.

Cell-based regenerative therapy may be useful for treatment of acute myocardial infarction (AMI). Animal xenograft models are ideally suited for preclinical studies evaluating prospective treatment regimes, identifying candidate human cell populations, and gaining mechanistic insight. Here we address whether the athymic nude rat is suitable as a xenograft model for the study of human CD34+ mobilized peripheral blood stem cells (M-PBSCs) in the repair of AMI. We injected human donor cells into the infarct border of athymic nude rats with surgically induced AMI and evaluated engraftment and functional improvement. We found no human engraftment by immunofluorescence staining at 14 days after transplantation or functional improvement at days 2 and 14 compared to controls. The lack of long-term human engraftment was furthermore confirmed in a time series study analyzing animals at 0, 24, 48, 72, and 96 h after transplantation. Although we found fluorescent microbeads coinjected with human CD34+ M-PBSCs at all time points, the number of donor cells rapidly declined and became undetectable at 96 h. CD34+ M-PBSCs from the same donor used to treat athymic nude rat hearts engrafted the bone marrow of nonobese diabetic/severe combined immunodeficient mice 8-10 weeks after transplantation. In conclusion, human CD34+ M-PBSCs with confirmed hematopoietic engraftment potential rapidly disappeared from the site of injury following intramyocardial transplantation in the athymic nude rat AMI model.