Molecular remodeling of Cx43, but not structural remodeling, promotes arrhythmias in an arrhythmogenic canine model of nonischemic heart failure.

BACKGROUND: Both gap junctional remodeling and interstitial fibrosis have been linked to impaired electrical conduction velocity (CV) and fatal ventricular arrhythmias in nonischemic heart failure (HF). However, the arrhythmogenic role of the ventricular gap junctional Cx43 in nonischemic HF remains in debate. Here, we assessed this in a newly developed arrhythmogenic canine model of nonischemic HF. METHODS AND RESULTS: Nonischemic HF was induced in canines by combined aortic valve insufficiency and aortic constriction. Left ventricular (LV) myocardium from HF dogs showed similar pathological changes to that of humans. HF dogs had reduced LV function, widened QRS complexes, and spontaneous nonsustained ventricular tachycardia. CV was measured in intact LV epicardium with high-density grid mapping. Total (Cx43-T) and nonphosphorylated Cx43 (Cx43-NP) and histological interstitial fibrosis were assessed from these mapped LV tissues. Longitudinal CV, which was slowed in HF (49 ± 1 vs. 65 ± 2 cm/s in Ctl), was positively correlated with reduced total junctional Cx43 and negatively correlated with markedly increased junctional Cx43-NP (2-fold) in HF. Cx43 dephosphorylation in HF was associated with enhanced colocalization of PP2A at the level of Cx43. Unchanged action potential upstroke and transverse CV were associated with unaltered Cx43 lateralization and interstitial fibrosis in the nonischemic HF canine LV. CONCLUSION: Our unique arrhythmogenic canine model of HF resembles human nonischemic HF (prior to the end stage). Cx43 remodeling occurs prior to the structural remodeling (with lack of fibrosis) in HF and it is crucial in slowed CV and ventricular arrhythmia development. Our findings suggest that altered Cx43 alone is arrhythmogenic and modulation of Cx43 has the anti-arrhythmic therapeutic potential for HF patients.

Flexor tendon repair with a knotless, bidirectional barbed suture: an in vivo biomechanical analysis.

PURPOSE: To compare and analyze biomechanical properties and histological characteristics of flexor tendons either repaired by a 4-strand modified Kessler technique or using barbed suture with a knotless repair technique in an in vivo model. METHODS: A total of 25 chickens underwent surgical transection of the flexor digitorum profundus tendon followed by either a 4-strand Kessler repair or a knotless repair with barbed suture. Chickens were randomly assigned to 1 of 3 groups with various postoperative times to death. Harvested tendons were subjected to biomechanical testing or histologic analysis. RESULTS: Harvested tendons revealed failures in 25% of knotless repairs (8 of 32) and 8% of 4-strand Kessler repairs (2 of 24). Biomechanical testing revealed no significant difference in tensile strength between 4-strand Kessler and barbed repairs; however, this lack of difference may be attributed to lower statistical power. We noted a trend toward a gradual decrease in strength over time for barbed repairs, whereas we noticed the opposite for the 4-strand Kessler repairs. Mode of failure during testing differed between repair types. The barbed repairs tended toward suture breakage as opposed to 4-strand Kessler repairs, which demonstrated suture pullout. Histological analysis identified no difference in the degree of inflammation or fibrosis; however, there was a vigorous foreign body reaction around the 4-strand Kessler repair and no such response around the barbed repairs. CONCLUSIONS: In this model, knotless barbed repairs trended toward higher in vivo failure rates and biomechanical inferiority under physiologic conditions, with each repair technique differing in mode of failure and respective histologic reaction. We are unable to recommend the use of knotless barbed repair over the 4-strand modified Kessler technique. CLINICAL RELEVANCE: For the repair techniques tested, surgeons should prefer standard Kessler repairs over the described knotless technique with barbed suture.

IND-directed safety and biodistribution study of intravenously injected cetuximab-IRDye800 in cynomolgus macaques.

PURPOSE: The use of receptor-targeted antibodies conjugated to fluorophores is actively being explored for real-time imaging of disease states; however, the toxicity of the bioconjugate has not been assessed in non-human primates. PROCEDURES: To this end, the in vivo toxicity and pharmacokinetics of IRDye800 conjugated to cetuximab (cetuximab-IRDye800; 21 mg/kg; equivalent to 250 mg/m(2) human dose) were assessed in male cynomolgus monkeys over 15 days following intravenous injection and compared with an unlabeled cetuximab-dosed control group. RESULTS: Cetuximab-IRDye800 was well tolerated. There were no infusion reactions, adverse clinical signs, mortality, weight loss, or clinical histopathology findings. The plasma half-life for the cetuximab-IRDye800 and cetuximab groups was equivalent (2.5 days). The total recovered cetuximab-IRDye800 in all tissues at study termination was estimated to be 12 % of the total dose. Both cetuximab-IRDye800 and cetuximab groups showed increased QTc after dosing. The QTc for the cetuximab-dosed group returned to baseline by day 15, while the QTc of the cetuximab-IRDye800 remained elevated compared to baseline. CONCLUSION: IRDye800 in low molar ratios does not significantly impact cetuximab half-life or result in organ toxicity. These studies support careful cardiac monitoring (ECG) for human studies using fluorescent dyes.

Imaging cardiac activation sequence during ventricular tachycardia in a canine model of nonischemic heart failure.

Noninvasive cardiac activation imaging of ventricular tachycardia (VT) is important in the clinical diagnosis and treatment of arrhythmias in heart failure (HF) patients. This study investigated the ability of the three-dimensional cardiac electrical imaging (3DCEI) technique for characterizing the activation patterns of spontaneously occurring and norepinephrine (NE)-induced VTs in a newly developed arrhythmogenic canine model of nonischemic HF. HF was induced by aortic insufficiency followed by aortic constriction in three canines. Up to 128 body-surface ECGs were measured simultaneously with bipolar recordings from up to 232 intramural sites in a closed-chest condition. Data analysis was performed on the spontaneously occurring VTs (n=4) and the NE-induced nonsustained VTs (n=8) in HF canines. Both spontaneously occurring and NE-induced nonsustained VTs initiated by a focal mechanism primarily from the subendocardium, but occasionally from the subepicardium of left ventricle. Most focal initiation sites were located at apex, right ventricular outflow tract, and left lateral wall. The NE-induced VTs were longer, more rapid, and had more focal sites than the spontaneously occurring VTs. Good correlation was obtained between imaged activation sequence and direct measurements (averaged correlation coefficient of ∼0.70 over 135 VT beats). The reconstructed initiation sites were ∼10 mm from measured initiation sites, suggesting good localization in such a large animal model with cardiac size similar to a human. Both spontaneously occurring and NE-induced nonsustained VTs had focal initiation in this canine model of nonischemic HF. 3DCEI is feasible to image the activation sequence and help define arrhythmia mechanism of nonischemic HF-associated VTs.

Morning surge of ventricular arrhythmias in a new arrhythmogenic canine model of chronic heart failure is associated with attenuation of time-of-day dependence of heart rate and autonomic adaptation, and reduced cardiac chaos.

Patients with chronic heart failure (CHF) exhibit a morning surge in ventricular arrhythmias, but the underlying cause remains unknown. The aim of this study was to determine if heart rate dynamics, autonomic input (assessed by heart rate variability (HRV)) and nonlinear dynamics as well as their abnormal time-of-day-dependent oscillations in a newly developed arrhythmogenic canine heart failure model are associated with a morning surge in ventricular arrhythmias. CHF was induced in dogs by aortic insufficiency & aortic constriction, and assessed by echocardiography. Holter monitoring was performed to study time-of-day-dependent variation in ventricular arrhythmias (PVCs, VT), traditional HRV measures, and nonlinear dynamics (including detrended fluctuations analysis α1 and α2 (DFAα1 & DFAα2), correlation dimension (CD), and Shannon entropy (SE)) at baseline, as well as 240 days (240 d) and 720 days (720 d) following CHF induction. LV fractional shortening was decreased at both 240 d and 720 d. Both PVCs and VT increased with CHF duration and showed a morning rise (2.5-fold & 1.8-fold increase at 6 AM-noon vs midnight-6 AM) during CHF. The morning rise in HR at baseline was significantly attenuated by 52% with development of CHF (at both 240 d & 720 d). Morning rise in the ratio of low frequency to high frequency (LF/HF) HRV at baseline was markedly attenuated with CHF. DFAα1, DFAα2, CD and SE all decreased with CHF by 31, 17, 34 and 7%, respectively. Time-of-day-dependent variations in LF/HF, CD, DFA α1 and SE, observed at baseline, were lost during CHF. Thus in this new arrhythmogenic canine CHF model, attenuated morning HR rise, blunted autonomic oscillation, decreased cardiac chaos and complexity of heart rate, as well as aberrant time-of-day-dependent variations in many of these parameters were associated with a morning surge of ventricular arrhythmias.

Chymase mediates injury and mitochondrial damage in cardiomyocytes during acute ischemia/reperfusion in the dog.

Cardiac ischemia and reperfusion (I/R) injury occurs because the acute increase in oxidative/inflammatory stress during reperfusion culminates in the death of cardiomyocytes. Currently, there is no drug utilized clinically that attenuates I/R injury in patients. Previous studies have demonstrated degranulation of mast cell contents into the interstitium after I/R. Using a dog model of I/R, we tested the role of chymase, a mast cell protease, in cardiomyocyte injury using a specific oral chymase inhibitor (CI). 15 adult mongrel dogs had left anterior descending artery occlusion for 60 min and reperfusion for 100 minutes. 9 dogs received vehicle and 6 were pretreated with a specific CI. In vivo cardiac microdialysis demonstrated a 3-fold increase in interstitial fluid chymase activity in I/R region that was significantly decreased by CI. CI pretreatment significantly attenuated loss of laminin, focal adhesion complex disruption, and release of troponin I into the circulation. Microarray analysis identified an I/R induced 17-fold increase in nuclear receptor subfamily 4A1 (NR4A1) and significantly decreased by CI. NR4A1 normally resides in the nucleus but can induce cell death on migration to the cytoplasm. I/R caused significant increase in NR4A1 protein expression and cytoplasmic translocation, and mitochondrial degradation, which were decreased by CI. Immunohistochemistry also revealed a high concentration of chymase within cardiomyocytes after I/R. In vitro, chymase added to culture HL-1 cardiomyocytes entered the cytoplasm and nucleus in a dynamin-dependent fashion, and promoted cytoplasmic translocation of NR4A1 protein. shRNA knockdown of NR4A1 on pre-treatment of HL-1 cells with CI significantly decreased chymase-induced cell death and mitochondrial damage. These results suggest that the beneficial effects of an orally active CI during I/R are mediated in the cardiac interstitium as well as within the cardiomyocyte due to a heretofore-unrecognized chymase entry into cardiomyocytes.

Intestinal epithelial apoptosis initiates gut mucosal injury during extracorporeal membrane oxygenation in the newborn piglet.

Neonates and young infants exposed to extracorporeal circulation during extracorporeal membrane oxygenation (ECMO) and cardiopulmonary bypass are at risk of developing a systemic inflammatory response syndrome with multi-organ dysfunction. We used a piglet model of ECMO to investigate the hypothesis that epithelial apoptosis is an early event that precedes villous damage during ECMO-related bowel injury. Healthy 3-week-old piglets were subjected to ECMO for up to 8 h. Epithelial apoptosis was measured in histopathological analysis, nuclear imaging, and terminal deoxynucleotidyl transferase dUTP nick end labeling. Plasma intestinal fatty acid-binding protein (I-FABP) levels were measured by enzyme immunoassay. Intestinal mast cells were isolated by fluorescence-assisted cell sorting. Cleaved caspase-8, caspase-9, phospho-p38 MAPK, and fas ligand expression were investigated by immunohistochemistry, western blots, and reverse transcriptase-quantitative PCR. Piglet ECMO was associated with increased gut epithelial apoptosis. Extensive apoptotic changes were noted on villus tips and in scattered crypt cells after 2 h of ECMO. After 8 h, the villi were denuded and apoptotic changes were evident in a majority of crypt cells. Increased circulating I-FABP levels, a marker of gut epithelial injury, showed that epithelial injury occurred during ECMO. We detected increased cleaved caspase-8, but not cleaved caspase-9, in epithelial cells indicating that the extrinsic apoptotic pathway was active. ECMO was associated with increased fas ligand expression in intestinal mast cells, which was induced through activation of the p38 mitogen-activated protein kinase. We conclude that epithelial apoptosis is an early event that initiates gut mucosal injury in a piglet model of ECMO.

Noninvasive cardiac activation imaging of ventricular arrhythmias during drug-induced QT prolongation in the rabbit heart.

BACKGROUND: Imaging myocardial activation from noninvasive body surface potentials promises to aid in both cardiovascular research and clinical medicine. OBJECTIVE: To investigate the ability of a noninvasive 3-dimensional cardiac electrical imaging technique for characterizing the activation patterns of dynamically changing ventricular arrhythmias during drug-induced QT prolongation in rabbits. METHODS: Simultaneous body surface potential mapping and 3-dimensional intracardiac mapping were performed in a closed-chest condition in 8 rabbits. Data analysis was performed on premature ventricular complexes, couplets, and torsades de pointes (TdP) induced during intravenous administration of clofilium and phenylephrine with combinations of various infusion rates. RESULTS: The drug infusion led to a significant increase in the QT interval (from 175 ± 7 to 274 ± 31 ms) and rate-corrected QT interval (from 183 ± 5 to 262 ± 21 ms) during the first dose cycle. All the ectopic beats initiated by a focal activation pattern. The initial beat of TdPs arose at the focal site, whereas the subsequent beats were due to focal activity from different sites or 2 competing focal sites. The imaged results captured the dynamic shift of activation patterns and were in good correlation with the simultaneous measurements, with a correlation coefficient of 0.65 ± 0.02 averaged over 111 ectopic beats. Sites of initial activation were localized to be ~5 mm from the directly measured initiation sites. CONCLUSIONS: The 3-dimensional cardiac electrical imaging technique could localize the origin of activation and image activation sequence of TdP during QT prolongation induced by clofilium and phenylephrine in rabbits. It offers the potential to noninvasively investigate the proarrhythmic effects of drug infusion and assess the mechanisms of arrhythmias on a beat-to-beat basis.

Evaluation of acute cardiac and chest wall damage after shocks with a subcutaneous implantable cardioverter defibrillator in Swine.

BACKGROUND: A subcutaneous implantable cardioverter defibrillator (S-ICD) could ease placement and reduce complications of transvenous ICDs, but requires more energy than transvenous ICDs. Therefore we assessed cardiac and chest wall damage caused by the maximum energy shocks delivered by both types of clinical devices. METHODS: During sinus rhythm, anesthetized pigs (38 ± 6 kg) received an S-ICD (n = 4) and five 80-Joule (J) shocks, or a transvenous ICD (control, n = 4) and five 35-J shocks. An inactive S-ICD electrode was implanted into the same control pigs to study implant trauma. All animals survived 24 hours. Troponin I and creatine kinase muscle isoenzyme (CK-MM) were measured as indicators of myocardial and skeletal muscle injury. Histopathological injury of heart, lungs, and chest wall was assessed using semiquantitative scoring. RESULTS: Troponin I was significantly elevated at 4 hours and 24 hours (22.6 ± 16.3 ng/mL and 3.1 ± 1.3 ng/mL; baseline 0.07 ± 0.09 ng/mL) in control pigs but not in S-ICD pigs (0.12 ± 0.11 ng/mL and 0.13 ± 0.13 ng/mL; baseline 0.06 ± 0.03 ng/mL). CK-MM was significantly elevated in S-ICD pigs after shocks (6,544 ± 1,496 U/L and 9,705 ± 6,240 U/L; baseline 704 ± 398 U/L) but not in controls. Electrocardiogram changes occurred postshock in controls but not in S-ICD pigs. The myocardium and lungs were histologically normal in both groups. Subcutaneous injury was greater in S-ICD compared to controls. CONCLUSION: Although CK-MM suggested more skeletal muscle injury in S-ICD pigs, significant cardiac, lung, and chest wall histopathological changes were not detected in either group. Troponin I data indicate significantly less cardiac injury from 80-J S-ICD shocks than 35-J transvenous shocks.

Ascending-ramp biphasic waveform has a lower defibrillation threshold and releases less troponin I than a truncated exponential biphasic waveform.

BACKGROUND: We tested the hypothesis that the shape of the shock waveform affects not only the defibrillation threshold but also the amount of cardiac damage. METHODS AND RESULTS: Defibrillation thresholds were determined for 11 waveforms-3 ascending-ramp waveforms, 3 descending-ramp waveforms, 3 rectilinear first-phase biphasic waveforms, a Gurvich waveform, and a truncated exponential biphasic waveform-in 6 pigs with electrodes in the right ventricular apex and superior vena cava. The ascending, descending, and rectilinear waveforms had 4-, 8-, and 16-millisecond first phases and a 3.5-millisecond rectilinear second phase that was half the voltage of the first phase. The exponential biphasic waveform had a 60% first-phase and a 50% second-phase tilt. In a second study, we attempted to defibrillate after 10 seconds of ventricular fibrillation with a single ≈30-J shock (6 pigs successfully defibrillated with 8-millisecond ascending, 8-millisecond rectilinear, and truncated exponential biphasic waveforms). Troponin I blood levels were determined before and 2 to 10 hours after the shock. The lowest-energy defibrillation threshold was for the 8-milliseconds ascending ramp (14.6±7.3 J [mean±SD]), which was significantly less than for the truncated exponential (19.6±6.3 J). Six hours after shock, troponin I was significantly less for the ascending-ramp waveform (0.80±0.54 ng/mL) than for the truncated exponential (1.92±0.47 ng/mL) or the rectilinear waveform (1.17±0.45 ng/mL). CONCLUSIONS: The ascending ramp has a significantly lower defibrillation threshold and at ≈30 J causes 58% less troponin I release than the truncated exponential biphasic shock. Therefore, the shock waveform affects both the defibrillation threshold and the amount of cardiac damage.

Left ventricular remodeling in preclinical experimental mitral regurgitation of dogs.

Dogs with experimental mitral regurgitation (MR) provide insights into the left ventricular remodeling in preclinical MR. The early preclinical left ventricular (LV) changes after mitral regurgitation represent progressive dysfunctional remodeling, in that no compensatory response returns the functional stroke volume (SV) to normal even as total SV increases. The gradual disease progression leads to mitral annulus stretch and enlargement of the regurgitant orifice, further increasing the regurgitant volume. Remodeling with loss of collagen weave and extracellular matrix (ECM) is accompanied by stretching and hypertrophy of the cross-sectional area and length of the cardiomyocyte. Isolated ventricular cardiomyocytes demonstrate dysfunction based on decreased cell shortening and reduced intracellular calcium transients before chamber enlargement or decreases in contractility in the whole heart can be clinically appreciated. The genetic response to increased end-diastolic pressure is down-regulation of genes associated with support of the collagen and ECM and up-regulation of genes associated with matrix remodeling. Experiments have not demonstrated any beneficial effects on remodeling from treatments that decrease afterload via blocking the renin-angiotensin system (RAS). Beta-1 receptor blockade and chymase inhibition have altered the progression of the LV remodeling and have supported cardiomyocyte function. The geometry of the LV during the remodeling provides insight into the importance of regional differences in responses to wall stress.

Noninvasive reconstruction of the three-dimensional ventricular activation sequence during pacing and ventricular tachycardia in the canine heart.

Single-beat imaging of myocardial activation promises to aid in both cardiovascular research and clinical medicine. In the present study we validate a three-dimensional (3D) cardiac electrical imaging (3DCEI) technique with the aid of simultaneous 3D intracardiac mapping to assess its capability to localize endocardial and epicardial initiation sites and image global activation sequences during pacing and ventricular tachycardia (VT) in the canine heart. Body surface potentials were measured simultaneously with bipolar electrical recordings in a closed-chest condition in healthy canines. Computed tomography images were obtained after the mapping study to construct realistic geometry models. Data analysis was performed on paced rhythms and VTs induced by norepinephrine (NE). The noninvasively reconstructed activation sequence was in good agreement with the simultaneous measurements from 3D cardiac mapping with a correlation coefficient of 0.74 ± 0.06, a relative error of 0.29 ± 0.05, and a root mean square error of 9 ± 3 ms averaged over 460 paced beats and 96 ectopic beats including premature ventricular complexes, couplets, and nonsustained monomorphic VTs and polymorphic VTs. Endocardial and epicardial origins of paced beats were successfully predicted in 72% and 86% of cases, respectively, during left ventricular pacing. The NE-induced ectopic beats initiated in the subendocardium by a focal mechanism. Sites of initial activation were estimated to be ∼7 mm from the measured initiation sites for both the paced beats and ectopic beats. For the polymorphic VTs, beat-to-beat dynamic shifts of initiation site and activation pattern were characterized by the reconstruction. The present results suggest that 3DCEI can noninvasively image the 3D activation sequence and localize the origin of activation of paced beats and NE-induced VTs in the canine heart with good accuracy. This 3DCEI technique offers the potential to aid interventional therapeutic procedures for treating ventricular arrhythmias arising from epicardial or endocardial sites and to noninvasively assess the mechanisms of these arrhythmias.

Noninvasive imaging of three-dimensional cardiac activation sequence during pacing and ventricular tachycardia.

BACKGROUND: Imaging cardiac excitation within ventricular myocardium is important in the treatment of cardiac arrhythmias and might help improve our understanding of arrhythmia mechanisms. OBJECTIVE: This study sought to rigorously assess the imaging performance of a 3-dimensional (3D) cardiac electrical imaging (3DCEI) technique with the aid of 3D intracardiac mapping from up to 216 intramural sites during paced rhythm and norepinephrine (NE)-induced ventricular tachycardia (VT) in the rabbit heart. METHODS: Body surface potentials and intramural bipolar electrical recordings were simultaneously measured in a closed-chest condition in 13 healthy rabbits. Single-site pacing and dual-site pacing were performed from ventricular walls and septum. VTs and premature ventricular complexes (PVCs) were induced by intravenous NE. Computed tomography images were obtained to construct geometry models. RESULTS: The noninvasively imaged activation sequence correlated well with invasively measured counterpart, with a correlation coefficient of 0.72 ± 0.04, and a relative error of 0.30 ± 0.02 averaged over 520 paced beats as well as 73 NE-induced PVCs and VT beats. All PVCs and VT beats initiated in the subendocardium by a nonreentrant mechanism. The averaged distance from the imaged site of initial activation to the pacing site or site of arrhythmias determined from intracardiac mapping was ∼5 mm. For dual-site pacing, the double origins were identified when they were located at contralateral sides of ventricles or at the lateral wall and the apex. CONCLUSION: 3DCEI can noninvasively delineate important features of focal or multifocal ventricular excitation. It offers the potential to aid in localizing the origins and imaging activation sequences of ventricular arrhythmias, and to provide noninvasive assessment of the underlying arrhythmia mechanisms.

Vernakalant selectively prolongs atrial refractoriness with no effect on ventricular refractoriness or defibrillation threshold in pigs.

Vernakalant is a novel antiarrhythmic agent that has demonstrated clinical efficacy for the treatment of atrial fibrillation. Vernakalant blocks, to various degrees, cardiac sodium and potassium channels with a pattern that suggests atrial selectivity. We hypothesized, therefore, that vernakalant would affect atrial more than ventricular effective refractory period (ERP) and have little or no effect on ventricular defibrillation threshold (DFT). Atrial and ventricular ERP and ventricular DFT were determined before and after treatment with vernakalant or vehicle in 23 anesthetized male mixed-breed pigs. Vernakalant was infused at a rate designed to achieve stable plasma levels similar to those in human clinical trials. Atrial and ventricular ERP were determined by endocardial extrastimuli delivered to the right atria or right ventricle. Defibrillation was achieved using external biphasic shocks delivered through adhesive defibrillation patches placed on the thorax after 10 seconds of electrically induced ventricular fibrillation. The DFT was estimated using the Dixon "up-and-down" method. Vernakalant significantly increased atrial ERP compared with vehicle controls (34 ± 8 versus 9 ± 7 msec, respectively) without significantly affecting ventricular ERP or DFT. This is consistent with atrial selective actions and supports the conclusion that vernakalant does not alter the efficacy of electrical defibrillation.

Noninvasive reconstruction of the three-dimensional ventricular activation sequence during pacing and ventricular tachycardia in the rabbit heart.

Ventricular arrhythmias represent one of leading causes for sudden cardiac death, a significant problem in public health. Noninvasive imaging of cardiac electric activities associated with ventricular arrhythmias plays an important role in better our understanding of the mechanisms and optimizing the treatment options. The present study aims to rigorously validate a novel three-dimensional (3-D) cardiac electrical imaging (3-DCEI) technique with the aid of 3-D intra-cardiac mapping during paced rhythm and ventricular tachycardia (VT) in the rabbit heart. Body surface potentials and intramural bipolar electrical recordings were simultaneously measured in a closed-chest condition in thirteen healthy rabbits. Single-site pacing and dual-site pacing were performed from ventricular walls and septum. VTs and premature ventricular complexes (PVCs) were induced by intravenous norepinephrine (NE). The non-invasively imaged activation sequence correlated well with invasively measured counterparts, with a correlation coefficient of 0.72 and a relative error of 0.30 averaged over all paced beats and NE-induced PVCs and VT beats. The averaged distance from imaged site of initial activation to measured site determined from intra-cardiac mapping was ∼5mm. These promising results suggest that 3-DCEI is feasible to non-invasively localize the origins and image activation sequence of focal ventricular arrhythmias.

Chymase inhibition prevents fibronectin and myofibrillar loss and improves cardiomyocyte function and LV torsion angle in dogs with isolated mitral regurgitation.

BACKGROUND: The left ventricular (LV) dilatation of isolated mitral regurgitation (MR) is associated with an increase in chymase and a decrease in interstitial collagen and extracellular matrix. In addition to profibrotic effects, chymase has significant antifibrotic actions because it activates matrix metalloproteinases and kallikrein and degrades fibronectin. Thus, we hypothesize that chymase inhibitor (CI) will attenuate extracellular matrix loss and LV remodeling in MR. METHODS AND RESULTS: We studied dogs with 4 months of untreated MR (MR; n=9) or MR treated with CI (MR+CI; n=8). Cine MRI demonstrated a >40% increase in LV end-diastolic volume in both groups, consistent with a failure of CI to improve a 25% decrease in interstitial collagen in MR. However, LV cardiomyocyte fractional shortening was decreased in MR versus normal dogs (3.71±0.24% versus 4.81±0.31%; P<0.05) and normalized in MR+CI dogs (4.85±0.44%). MRI with tissue tagging demonstrated an increase in LV torsion angle in MR+CI versus MR dogs. CI normalized the significant decrease in fibronectin and FAK phosphorylation and prevented cardiomyocyte myofibrillar degeneration in MR dogs. In addition, total titin and its stiffer isoform were increased in the LV epicardium and paralleled the changes in fibronectin and FAK phosphorylation in MR+CI dogs. CONCLUSIONS: These results suggest that chymase disrupts cell surface-fibronectin connections and FAK phosphorylation that can adversely affect cardiomyocyte myofibrillar structure and function. The greater effect of CI on epicardial versus endocardial titin and noncollagen cell surface proteins may be responsible for the increase in torsion angle in chronic MR.

Mast cell stabilization decreases cardiomyocyte and LV function in dogs with isolated mitral regurgitation.

BACKGROUND: Mast cells are increased in isolated mitral regurgitation (MR) in the dog and may mediate extracellular matrix loss and left ventricular (LV) dilatation. We tested the hypothesis that mast cell stabilization would attenuate LV remodeling and improve function in the MR dog. METHODS AND RESULTS: MR was induced in adult dogs randomized to no treatment (MR, n = 5) or to the mast cell stabilizer, ketotifen (MR + MCS, n = 4) for 4 months. LV hemodynamics were obtained at baseline and after 4 months of MR and magnetic resonance imaging (MRI) was performed at sacrifice. MRI-derived, serial, short-axis LV end-diastolic (ED) and end-systolic (ES) volumes, LVED volume/mass ratio, and LV 3-dimensional radius/wall thickness were increased in MR and MR + MCS dogs compared with normal dogs (n = 6) (P < .05). Interstitial collagen was decreased by 30% in both MR and MR + MCS versus normal dogs (P < .05). LV contractility by LV maximum time-varying elastance was significantly depressed in MR and MR + MCS dogs. Furthermore, cardiomyocyte fractional shortening was decreased in MR versus normal dogs and further depressed in MR + MCS dogs (P < .05). In vitro administration of ketotifen to normal cardiomyocytes also significantly decreased fractional shortening and calcium transients. CONCLUSIONS: Chronic mast cell stabilization did not attenuate eccentric LV remodeling or collagen loss in MR. However, MCS therapy had a detrimental effect on LV function because of a direct negative inotropic effect on cardiomyocyte function.

Extracorporeal membrane oxygenation causes loss of intestinal epithelial barrier in the newborn piglet.

Extracorporeal membrane oxygenation (ECMO) is an important life-support system used in neonates and young children with intractable cardiorespiratory failure. In this study, we used our porcine neonatal model of venoarterial ECMO to investigate whether ECMO causes gut barrier dysfunction. We subjected 3-wk-old previously healthy piglets to venoarterial ECMO for up to 8 h and evaluated gut mucosal permeability, bacterial translocation, plasma levels of bacterial products, and ultrastructural changes in gut epithelium. We also measured plasma lipopolysaccharide (LPS) levels in a small cohort of human neonates receiving ECMO. In our porcine model, ECMO caused a rapid increase in gut mucosal permeability within the first 2 h of treatment, leading to a 6- to 10-fold rise in circulating bacterial products. These changes in barrier function were associated with cytoskeletal condensation in epithelial cells, which was explained by phosphorylation of a myosin II regulatory light chain. In support of these findings, we also detected elevated plasma LPS levels in human neonates receiving ECMO, indicating a similar loss of gut barrier function in these infants. On the basis of these data, we conclude that ECMO is an independent cause of gut barrier dysfunction and bacterial translocation may be an important contributor to ECMO-related inflammation.

Comparison of low-energy versus high-energy biphasic defibrillation shocks following prolonged ventricular fibrillation.

INTRODUCTION: Since the initial development of the defibrillator, there has been concern that, while delivery of a large electric shock would stop fibrillation, it would also cause damage to the heart. This concern has been raised again with the development of the biphasic defibrillator. OBJECTIVE: To compare defibrillation efficacy, postshock cardiac function, and troponin I levels following 150-J and 360-J shocks. METHODS: Nineteen swine were anesthetized with isoflurane and instrumented with pressure catheters in the left ventricle, aorta, and right atrium. The animals were fibrillated for 6 minutes, followed by defibrillation with either low-energy (n = 8) or high-energy (n = 11) shocks. After defibrillation, chest compressions were initiated and continued until return of spontaneous circulation (ROSC). Epinephrine, 0.01 mg/kg every 3 minutes, was given for arterial blood pressure < 50 mmHg. Hemodynamic parameters were recorded for four hours. Transthoracic echocardiography was performed and troponin I levels were measured at baseline and four hours following ventricular fibrillation (VF). RESULTS: Survival rates at four hours were not different between the two groups (low-energy, 5 of 8; high-energy, 7 of 11). Results for arterial blood pressure, positive dP/dt (first derivative of pressure measured over time, a measure of left ventricular contractility), and negative dP/dt at the time of lowest arterial blood pressure (ABP) following ROSC were not different between the two groups (p = not significant [NS]), but were lower than at baseline. All hemodynamic measures returned to baseline by four hours. Ejection fractions, stroke volumes, and cardiac outputs were not different between the two groups at four hours. Troponin I levels at four hours were not different between the two groups (12 +/- 11 ng/mL versus 21 +/- 26 ng/mL, p = NS) but were higher at four hours than at baseline (19 +/- 19 ng/mL versus 0.8 +/- 0.5 ng/mL, p < 0.05, groups combined). CONCLUSION: Biphasic 360-J shocks do not cause more cardiac damage than biphasic 150-J shocks in this animal model of prolonged VF and resuscitation.

Plasma concentrations of inflammatory cytokines rise rapidly during ECMO-related SIRS due to the release of preformed stores in the intestine.

Extracorporeal membrane oxygenation (ECMO) is a life-saving support system used in neonates and young children with severe cardiorespiratory failure. Although ECMO has reduced mortality in these critically ill patients, almost all patients treated with ECMO develop a systemic inflammatory response syndrome (SIRS) characterized by a 'cytokine storm', leukocyte activation, and multisystem organ dysfunction. We used a neonatal porcine model of ECMO to investigate whether rising plasma concentrations of inflammatory cytokines during ECMO reflect de novo synthesis of these mediators in inflamed tissues, and therefore, can be used to assess the severity of ECMO-related SIRS. Previously healthy piglets (3-week-old) were subjected to venoarterial ECMO for up to 8 h. SIRS was assessed by histopathological analysis, measurement of neutrophil activation (flow cytometry), plasma cytokine concentrations (enzyme immunoassays), and tissue expression of inflammatory genes (PCR/western blots). Mast cell degranulation was investigated by measurement of plasma tryptase activity. Porcine neonatal ECMO was associated with systemic inflammatory changes similar to those seen in human neonates. Tumor necrosis factor-alpha (TNF-alpha) and interleukin-8 (IL-8) concentrations rose rapidly during the first 2 h of ECMO, faster than the tissue expression of these cytokines. ECMO was associated with increased plasma mast cell tryptase activity, indicating that increased plasma concentrations of inflammatory cytokines during ECMO may result from mast cell degranulation and associated release of preformed cytokines stored in mast cells. TNF-alpha and IL-8 concentrations rose faster in plasma than in the peripheral tissues during ECMO, indicating that rising plasma levels of these cytokines immediately after the initiation of ECMO may not reflect increasing tissue synthesis of these cytokines. Mobilization of preformed cellular stores of inflammatory cytokines such as in mucosal mast cells may have an important pathophysiological role in ECMO-related SIRS.