Tetsuo Sasano

Engraftment, differentiation, and functional benefits of autologous cardiosphere-derived cells in porcine ischemic cardiomyopathy

Background— Cardiosphere-derived cells (CDCs) isolated from human endomyocardial biopsies reduce infarct size and improve cardiac function in mice. Safety and efficacy testing in large animals is necessary for clinical translation. Methods and Results— Mesenchymal stem cells, which resemble CDCs in size and thrombogenicity, have been associated with infarction after intracoronary infusion. To maximize CDC engraftment while avoiding infarction, we optimized the infusion protocol in 19 healthy pigs. A modified cocktail of CDCs in calcium-free PBS, 100 U/mL of heparin, and 250 &mgr;g/mL of nitroglycerin eliminated infusion-related infarction. Subsequent infusion experiments in 17 pigs with postinfarct left ventricular dysfunction showed CDC doses ≥107 but <2.5×107 result in new myocardial tissue formation without infarction. In a pivotal randomized study, 7 infarcted pigs received 300 000 CDCs/kg (≈107 total) and 7 received placebo (vehicle alone). Cardiac magnetic resonance imaging 8 weeks later showed CDC treatment decreased relative infarct size (19.2% to 14.2% of left ventricle infarcted, P=0.01), whereas placebo did not (17.7% to 15.3%, P=0.22). End-diastolic volume increased in placebo, but not in CDC-treated animals. Hemodynamically, the rate of pressure change (dP/dt) maximum and dP/dt minimum were significantly better with CDC infusion. There was no difference between groups in the ability to induce ventricular tachycardia, nor was there any tumor or ectopic tissue formation. Conclusions— Intracoronary delivery of CDCs in a preclinical model of postinfarct left ventricular dysfunction results in formation of new cardiac tissue, reduces relative infarct size, attenuates adverse remodeling, and improves hemodynamics. The evidence of efficacy without obvious safety concerns at 8 weeks of follow-up motivates human studies in patients after myocardial infarction and in chronic ischemic cardiomyopathy.

Abnormal Sympathetic Innervation of Viable Myocardium and the Substrate of Ventricular Tachycardia After Myocardial Infarction

OBJECTIVES The aim of this study was to characterize the relationship between impaired sympathetic innervation and arrhythmia with noninvasive biologic imaging in an animal model of post-infarct ventricular tachycardia (VT). BACKGROUND Innervation might be abnormal in the normally perfused borderzone of myocardial infarction, contributing to myocardial catecholamine overexposure and arrhythmogenic risk. METHODS Myocardial infarction was induced by mid-left anterior descending coronary artery balloon occlusion in 11 pigs. Positron emission tomography (PET) of tissue perfusion and catecholamine uptake and storage was performed with [13N]-ammonia and [11C]-epinephrine 4 to 12 weeks later. Magnetic resonance imaging and invasive electrophysiology (electroanatomic mapping, basket catheter, VT inducibility) were performed within 1 week of PET. RESULTS When compared with a normal database of 9 healthy animals, reduced perfusion was observed in 37 +/- 7% of the left ventricle (LV). Epinephrine retention was reduced in 44 +/- 7% of LV, resulting in a perfusion/innervation mismatch of 7 +/- 4% LV. Sustained monomorphic VT was inducible in 7 of 11 animals. These animals showed a larger perfusion/innervation mismatch (10 +/- 4% vs. 4 +/- 2% LV for animals without VT; p = 0.02). Regionally, the degree of perfusion/innervation mismatch did not correlate with wall thickness or thickening but showed a significant correlation with reduced myocardial voltage (r = 0.93; p = 0.001) and with the site of earliest VT activation (chi-square 13.1; p < 0.001). CONCLUSIONS Noninvasive mapping of cardiac sympathetic nerve terminals reveals regionally impaired catecholamine uptake and storage in the normally perfused borderzone after experimental myocardial infarction. These areas might be useful to characterize the individual risk for ventricular arrhythmia.

Targeted Modification of Atrial Electrophysiology by Homogeneous Transmural Atrial Gene Transfer

Background—Safe and effective myocardial gene transfer remains elusive. Heterogeneous ventricular gene delivery has been achieved in small mammals but generally with methods not readily transferable to the clinic. Atrium-specific gene transfer has not yet been reported. We hypothesized that homogeneous atrial gene transfer could be achieved by direct application of adenoviral vectors to the epicardial surface, use of poloxamer gel to increase virus contact time, and mild trypsinization to increase virus penetration. Methods and Results—We “painted” recombinant adenovirus encoding the reporter gene Escherichia coli &bgr;-galactosidase directly onto porcine atria. Investigational variables included poloxamer use, trypsin concentration, and safety. Using the painting method, we modified the atrial phenotype with an adenovirus expressing HERG-G628S, a long-QT-syndrome mutant. Our results showed that application of virus with poloxamer alone resulted in diffuse epicardial gene transfer with negligible penetration into the myocardium. Dilute trypsin concentrations allowed complete transmural gene transfer. After trypsin exposure, echocardiographic left atrial diameter did not change. Left atrial function decreased on postoperative day 3 but returned to baseline by day 7. Tissue tensile strength was affected only in the 1% trypsin group. HERG-G628S gene transfer prolonged atrial action potential duration and refractory period without affecting ventricular electrophysiology. Conclusions—We show complete transmural atrial gene transfer by this novel painting method. Adaptation of the method could allow application to other tissue targets. Use with functional proteins in the atria could cure or even prevent diseases such as atrial fibrillation or sinus node dysfunction.

Connexin43 Gene Transfer Reduces Ventricular Tachycardia Susceptibility After Myocardial Infarction

OBJECTIVES The aim of this study was to evaluate the links between connexin43 (Cx43) expression, myocardial conduction velocity, and ventricular tachycardia in a model of healed myocardial infarction. BACKGROUND Post-infarction ventricular arrhythmias frequently cause sudden death. Impaired myocardial conduction has previously been linked to ventricular arrhythmias. Altered connexin expression is a potential source of conduction slowing identified in healed scar border tissues. The functional effect of increasing border-zone Cx43 has not been previously evaluated. METHODS Twenty-five Yorkshire pigs underwent anterior infarction by transient left anterior descending coronary artery occlusion, followed by weekly testing for arrhythmia inducibility. Twenty animals with reproducibly inducible sustained monomorphic ventricular tachycardia were randomized 2:1:1 to receive AdCx43, Adβgal, or no gene transfer. One week later, animals underwent follow-up electrophysiologic study and tissue assessment for several functional and molecular measures. RESULTS Animals receiving AdCx43 had less electrogram fractionation and faster conduction velocity in the anterior-septal border zone. Only 40% of AdCx43 animals remained inducible for ventricular tachycardia, while 100% of controls were inducible after gene transfer. AdCx43 animals had 2-fold higher Cx43 protein levels in the anterior-septal infarct border, with similar percents of phosphorylated and intercalated disk-localized Cx43 compared with controls. CONCLUSIONS These data mechanistically link Cx43 expression to slow conduction and arrhythmia susceptibility in the healed scar border zone. Targeted manipulation of Cx43 levels improved conduction velocity and reduced ventricular tachycardia susceptibility. Cx43 gene transfer represents a novel treatment strategy for post-infarction arrhythmias.

Integration of Infarct Size, Tissue Perfusion and Metabolism by Hybrid Cardiac PET–CT–Evaluation in a Porcine Model of Myocardial Infarction

Background—Hybrid positron emission tomography/computed tomography (PET-CT) allows for combination of PET perfusion/metabolism imaging with infarct detection by CT delayed contrast enhancement. We used this technique to obtain biomorphological insights into the interrelation between tissue damage, inflammation, and microvascular obstruction early after myocardial infarction. Methods and Results—A porcine model of left anterior descending coronary artery occlusion/reperfusion was studied. Seven animals underwent PET-CT within 3 days of infarction, and a control group of 3 animals was scanned at >4 weeks. Perfusion and glucose uptake were assessed by [13N]-ammonia/[18F]-deoxyglucose (FDG), and 64-slice CT delayed contrast enhancement was measured. In the acute infarct model, CT revealed a no-reflow phenomenon suggesting microvascular obstruction in 80% of all infarct segments. PET showed increased FDG uptake in 68% of the CT-defined infarct segments. Ex vivo staining and histology showed active inflammation in the acute infarct area as an explanation for increased glucose uptake. In chronic infarction, CT showed no microvascular obstruction and agreed well with matched perfusion/metabolism defects on PET. Conclusions—Perfusion/metabolism PET and delayed enhancement CT can be combined within a single hybrid PET-CT session. Increased regional FDG uptake in the acute infarct area is frequently observed. In contrast to the chronic infarct setting, this indicates tissue inflammation that is commonly associated with microvascular obstruction as identified by no reflow on CT. The consequences of these pathophysiological findings for subsequent ventricular remodeling should be explored in further studies.

Epileptic Seizure Prediction Based on Multivariate Statistical Process Control of Heart Rate Variability Features

Objective: The present study proposes a new epileptic seizure prediction method through integrating heart rate variability (HRV) analysis and an anomaly monitoring technique. Methods: Because excessive neuronal activities in the preictal period of epilepsy affect the autonomic nervous systems and autonomic nervous function affects HRV, it is assumed that a seizure can be predicted through monitoring HRV. In the proposed method, eight HRV features are monitored for predicting seizures by using multivariate statistical process control, which is a well-known anomaly monitoring method. Results: We applied the proposed method to the clinical data collected from 14 patients. In the collected data, 8 patients had a total of 11 awakening preictal episodes and the total length of interictal episodes was about 57 h. The application results of the proposed method demonstrated that seizures in ten out of eleven awakening preictal episodes could be predicted prior to the seizure onset, that is, its sensitivity was 91%, and its false positive rate was about 0.7 times per hour. Conclusion: This study proposed a new HRV-based epileptic seizure prediction method, and the possibility of realizing an HRV-based epileptic seizure prediction system was shown. Significance: The proposed method can be used in daily life, because the heart rate can be measured easily by using a wearable sensor.

Clinical Study Regarding the Anatomical Structures of the Right Atrial Isthmus Using Intra-Cardiac Echocardiography: Implication for Catheter Ablation of Common Atrial Flutter

Background: The construction of complete bi-directional block in the isthmus (ITH) between the tricuspid annulus and inferior vena cava by radiofrequency energy (RF) applications is sometimes hampered due to anatomical problems such as a thick isthmus or aneurysmal pouch in patients with common atrial flutter (AFL).Methods and Results: Fifteen patients were referred for RF ablation of AFL. The anatomical thickness of the right atrial ITH, diameter of the right atrium and thickness of the right atrial free wall were determined using intracardiac echocardiography (ICE), along with the endocardial electrogram recordings at the ITH. RF was applied at the ITH to create a transmural incision to treat the AFL. A significant parallel relationship between the maximum amplitude of the atrial electrogram and the thickness of the ITH, was observed. When the maximum amplitude of the atrial electrogram at the ITH exceeded 1.5 mV, the thickness at the ITH was approximately larger than 5 mm.Conclusions: Using ICE, the precise measurement of the anatomical structures in the heart, including the ITH, was feasible. From the amplitude of the atrial electrogram, a deduction of the thickness at the ITH was possible, which is indispensable information for the appropriate selection of the RF devices.

Ventricular tachycardia from the healed myocardial infarction scar: Validation of an animal model and utility of gene therapy

Life-threatening ventricular arrhythmias generally occur in the setting of structural heart disease. Current clinical options for patients at risk for these rhythm disturbances are limited. We developed a porcine model of inducible ventricular tachycardia originating in the border region of a healed myocardial infarction scar. After validating the model, we assessed gene transfer techniques, focusing on local modification of border zone tissues. We found that gene transfer of the dominant negative KCNH2-G628S mutation to the anteroseptal infarct border caused localized prolongation of effective refractory period in the target region and eliminated all ventricular arrhythmia inducibility. In this work, we characterize the animal model and review the gene transfer results.

Radiofrequency ablation of tachyarrhythmias in patients with Ebstein's anomaly

We performed radiofrequency catheter ablation in five patients associated with Ebsteins anomaly to cure their refractory tachyarrhythmias. The presenting arrhythmias were four cases of orthodromic circus movement tachycardia using accessory pathways as a requisite limb, including one case of a Mahaim fiber and one of atrial flutter of common variety. All accessory pathways, including the Mahaim fiber, were ablated by RF energy delivered through the catheter placed at the AV annulus rather than the displaced anatomical AV groove. Interestingly, the antegrade or retrograde conduction interval over these accessory pathways was relatively longer than that of usual accessory pathways, and the accessory pathway potential was fractionated in some cases. The location of the atrioventricular node was displaced from the usual position to the postero-inferior area of Kochs triangle in one case. The configuration of the flutter wave was larger than usual in height as well as in width. All tachyarrhythmias were cured by RF catheter ablation. In the case of RF catheter ablation for patients with Ebsteins anomaly, close attention is indispensable in order to accomplish it safely and successfully, because of the anatomical and functional differences peculiar to Ebsteins anomaly.

Novel method for earlier detection of phrenic nerve injury during cryoballoon applications for electrical isolation of pulmonary veins in patients with atrial fibrillation

BACKGROUND Diaphragmatic electrogram recording during cryoballoon ablation (CB-A) of atrial fibrillation is commonly used to predict phrenic nerve palsy (PNP). OBJECTIVE The purpose of this study was to investigate a novel method for predicting PNP at an earlier stage to prevent sustained PNP. METHODS A total of 197 patients undergoing CB-A were enrolled. We attempted to detect PNP using fluoroscopic images of diaphragmatic contractions and by monitoring diaphragmatic compound motor action potentials (CMAPs) provoked by superior vena cava (SVC) and left subclavian vein (LCV) pacing during CB-A for bilateral pulmonary veins (PVs). Pacing of the SVC and LCV was performed at 2 outputs, 1 exceeding the pacing threshold by 10% (MIN) and the other at maximum output (MAX). The time from freezing to the initiation of PNP, values of the CMAP amplitude, and severity of PNP were compared for the 2 outputs. RESULTS There was a significant difference in the time from freezing to initiation of PNP between MIN and MAX pacing (25.7 ± 5.7 vs 81.3 ± 7.4 seconds, P<.01). CMAP amplitudes also differed significantly (0.71 ± 0.39 vs 1.13 ± 0.42, P<.0001). SVC/LCV pacing with MIN output was able to detect PNP significantly earlier than MAX (27 ± 8 vs 91 ± 12 seconds, P<.01), and the time to PNP recovery was significantly shorter for the MIN output (20.2 ± 8.88 hours vs 4.8 ± 1.6 months, P<.001). CONCLUSION Pacing the SVC and LCV with lower output detect PNP significantly earlier than maximal output pacing and leads to recovery from PNP on the order of hours postprocedure rather than months.