Proliferation and Maturation: Janus and the Art of Cardiac Tissue Engineering.

During cardiac development and morphogenesis, cardiac progenitor cells differentiate into cardiomyocytes that expand in number and size to generate the fully formed heart. Much is known about the factors that regulate initial differentiation of cardiomyocytes, and there is ongoing research to identify how these fetal and immature cardiomyocytes develop into fully functioning, mature cells. Accumulating evidence indicates that maturation limits proliferation and conversely proliferation occurs rarely in cardiomyocytes of the adult myocardium. We term this oppositional interplay the proliferation-maturation dichotomy. Here we review the factors that are involved in this interplay and discuss how a better understanding of the proliferation-maturation dichotomy could advance the utility of human induced pluripotent stem cell-derived cardiomyocytes for modeling in 3-dimensional engineered cardiac tissues to obtain truly adult-level function.

Atherosclerotic Calcifications Have a Local Effect on the Peel Behavior of Human Aortic Media.

Aortic dissections, characterized by the propagation of a tear through the layers of the vessel wall, are critical, life-threatening events. Aortic calcifications are a common comorbidity in both acute and chronic dissections, yet their impact on dissection mechanics remains unclear. Using micro-computed tomography (CT) imaging, peel testing, and finite element modeling, this study examines the interplay between atherosclerotic calcifications and dissection mechanics. Samples cut from cadaveric human thoracic aortas were micro-CT imaged and subsequently peel-tested to map peel tension curves to the location of aortic calcifications. Empirical mode decomposition separated peel tension curves into high and low-frequency components, with high-frequency effects corresponding to interlamellar bonding mechanics and low-frequency effects to peel tension fluctuations. Finally, we used an idealized finite element model to examine how stiff calcifications affect aortic failure mechanics. Results showed that atherosclerosis influences dissection behavior on multiple length scales. Experimentally, atherosclerotic samples exhibited higher peel tensions and greater variance in the axial direction. The variation was driven by increased amplitudes of low-frequency tension fluctuations in diseased samples, indicating that more catastrophic propagations occur near calcifications. The simulations corroborated this finding, suggesting that the low-frequency changes resulted from the presence of a stiff calcification in the vessel wall. There were also modifications to the high-frequency peel mechanics, a response likely attributable to alterations in the microstructure and interlamellar bonding within the media. Considered collectively, these findings demonstrate that dissection mechanics are modified in aortic media nearby and adjacent to aortic calcifications.

A Data-Driven Preprocessing Framework for Atrial Fibrillation Intracardiac Electrocardiogram Analysis.

Atrial Fibrillation (AF) is the most common cardiac arrhythmia. Signal-processing approaches are widely used for the analysis of intracardiac electrograms (iEGMs), which are collected during catheter ablation from patients with AF. In order to identify possible targets for ablation therapy, dominant frequency (DF) is widely used and incorporated in electroanatomical mapping systems. Recently, a more robust measure, multiscale frequency (MSF), for iEGM data analysis was adopted and validated. However, before completing any iEGM analysis, a suitable bandpass (BP) filter must be applied to remove noise. Currently, no clear guidelines for BP filter characteristics exist. The lower bound of the BP filter is usually set to 3-5 Hz, while the upper bound (BP¯th) of the BP filter varies from 15 Hz to 50 Hz according to many researchers. This large range of BP¯th subsequently affects the efficiency of further analysis. In this paper, we aimed to develop a data-driven preprocessing framework for iEGM analysis, and validate it based on DF and MSF techniques. To achieve this goal, we optimized the BP¯th using a data-driven approach (DBSCAN clustering) and demonstrated the effects of different BP¯th on subsequent DF and MSF analysis of clinically recorded iEGMs from patients with AF. Our results demonstrated that our preprocessing framework with BP¯th = 15 Hz has the best performance in terms of the highest Dunn index. We further demonstrated that the removal of noisy and contact-loss leads is necessary for performing correct data iEGMs data analysis.

In Situ Expansion, Differentiation, and Electromechanical Coupling of Human Cardiac Muscle in a 3D Bioprinted, Chambered Organoid.

RATIONALE: One goal of cardiac tissue engineering is the generation of a living, human pump in vitro that could replace animal models and eventually serve as an in vivo therapeutic. Models that replicate the geometrically complex structure of the heart, harboring chambers and large vessels with soft biomaterials, can be achieved using 3-dimensional bioprinting. Yet, inclusion of contiguous, living muscle to support pump function has not been achieved. This is largely due to the challenge of attaining high densities of cardiomyocytes-a notoriously nonproliferative cell type. An alternative strategy is to print with human induced pluripotent stem cells, which can proliferate to high densities and fill tissue spaces, and subsequently differentiate them into cardiomyocytes in situ. OBJECTIVE: To develop a bioink capable of promoting human induced pluripotent stem cell proliferation and cardiomyocyte differentiation to 3-dimensionally print electromechanically functional, chambered organoids composed of contiguous cardiac muscle. METHODS AND RESULTS: We optimized a photo-crosslinkable formulation of native ECM (extracellular matrix) proteins and used this bioink to 3-dimensionally print human induced pluripotent stem cell-laden structures with 2 chambers and a vessel inlet and outlet. After human induced pluripotent stem cells proliferated to a sufficient density, we differentiated the cells within the structure and demonstrated function of the resultant human chambered muscle pump. Human chambered muscle pumps demonstrated macroscale beating and continuous action potential propagation with responsiveness to drugs and pacing. The connected chambers allowed for perfusion and enabled replication of pressure/volume relationships fundamental to the study of heart function and remodeling with health and disease. CONCLUSIONS: This advance represents a critical step toward generating macroscale tissues, akin to aggregate-based organoids, but with the critical advantage of harboring geometric structures essential to the pump function of cardiac muscle. Looking forward, human chambered organoids of this type might also serve as a test bed for cardiac medical devices and eventually lead to therapeutic tissue grafting.

A Bionic Testbed for Cardiac Ablation Tools.

Bionic-engineered tissues have been proposed for testing the performance of cardiovascular medical devices and predicting clinical outcomes ex vivo. Progress has been made in the development of compliant electronics that are capable of monitoring treatment parameters and being coupled to engineered tissues; however, the scale of most engineered tissues is too small to accommodate the size of clinical-grade medical devices. Here, we show substantial progress toward bionic tissues for evaluating cardiac ablation tools by generating a centimeter-scale human cardiac disk and coupling it to a hydrogel-based soft-pressure sensor. The cardiac tissue with contiguous electromechanical function was made possible by our recently established method to 3D bioprint human pluripotent stem cells in an extracellular matrix-based bioink that allows for in situ cell expansion prior to cardiac differentiation. The pressure sensor described here utilized electrical impedance tomography to enable the real-time spatiotemporal mapping of pressure distribution. A cryoablation tip catheter was applied to the composite bionic tissues with varied pressure. We found a close correlation between the cell response to ablation and the applied pressure. Under some conditions, cardiomyocytes could survive in the ablated region with more rounded morphology compared to the unablated controls, and connectivity was disrupted. This is the first known functional characterization of living human cardiomyocytes following an ablation procedure that suggests several mechanisms by which arrhythmia might redevelop following an ablation. Thus, bionic-engineered testbeds of this type can be indicators of tissue health and function and provide unique insight into human cell responses to ablative interventions.

Novel mapping techniques for rotor core detection using simulated intracardiac electrograms.

BACKGROUND: Catheter ablation is associated with limited success rates in patients with persistent atrial fibrillation (AF). Currently, existing mapping systems fail to identify critical target sites for ablation. Recently, we proposed and validated several techniques (multiscale frequency [MSF], Shannon entropy [SE], kurtosis [Kt], and multiscale entropy [MSE]) to identify pivot point of rotors using ex-vivo optical mapping animal experiments. However, the performance of these techniques is unclear for the clinically recorded intracardiac electrograms (EGMs), due to the different nature of the signals. OBJECTIVE: This study aims to evaluate the performance of MSF, MSE, SE, and Kt techniques to identify the pivot point of the rotor using unipolar and bipolar EGMs obtained from numerical simulations. METHODS: Stationary and meandering rotors were simulated in a 2D human atria. The performances of new approaches were quantified by comparing the "true" core of the rotor with the core identified by the techniques. Also, the performances of all techniques were evaluated in the presence of noise, scar, and for the case of the multielectrode multispline and grid catheters. RESULTS: Our results demonstrate that all the approaches are able to accurately identify the pivot point of both stationary and meandering rotors from both unipolar and bipolar EGMs. The presence of noise and scar tissue did not significantly affect the performance of the techniques. Finally, the core of the rotors was correctly identified for the case of multielectrode multispline and grid catheter simulations. CONCLUSION: The core of rotors can be successfully identified from EGMs using novel techniques; thus, providing motivation for future clinical implementations.

Interplay between ephaptic coupling and complex geometry of border zone during acute myocardial ischemia: Effect on arrhythmogeneity.

Acute myocardial ischemia is an imbalance between myocardial blood supply and demand, which is caused by the cessation of blood flow within the heart resulting from an obstruction in one of the major coronary arteries. A severe blockage may result in a region of nonperfused tissue known as ischemic core (IC). As a result, a border zone (BZ) between perfused and nonperfused regions is created due to differences in blood and oxygen supplies. Recent experimental findings reveal a complex "finger-like" geometry in BZ; however, its effect on arrhythmogenicity is not clear. Ephaptic coupling, which relies on the intercalated disk between cell ends, has been suggested to play an active role in mediating intercellular electrical communication when gap junctions are impaired. In this paper, we explored the interplay between ephaptic coupling and the geometry of BZ on action potential propagation across the ischemic region. Our study shows that ephaptic coupling can greatly suppress the occurrence of a conduction block, which points to its beneficial effect. The beneficial effect of ephaptic coupling is more evident in BZ with the "finger-like" geometry. In addition, the complex geometry of BZ, i.e., more frequent, deeper, and wider "fingers," promotes the conduction through the ischemic region. In contrast, the larger size of IC impedes the cardiac conduction across the ischemic region. Our results also show that ephaptic coupling promotes the impact of the complex geometry of BZ on signal propagation; however, it inhibits the impact of IC size.

Global vs local control of cardiac alternans in a 1D numerical model of human ventricular tissue.

Cardiac alternans is a proarrhythmic state in which the action potential duration (APD) of cardiac myocytes alternate between long and short values and often occurs under conditions of rapid pacing of cardiac tissue. In the ventricles, alternans is especially dangerous due to the life-threatening risk of developing arrhythmias, such as ventricular fibrillation. Alternans can be formed in periodically paced tissue as a result of pacing itself. Recently, it has been demonstrated that this pacing-induced alternans can be prevented by performing constant diastolic interval (DI) pacing, in which DI is independent of APD. However, constant DI pacing is difficult to implement in experimental settings since it requires the real-time measurement of APD. A more practical way was proposed based on electrocardiograms (ECGs), which give an indirect measure of the global DI relaxation period through the TR interval assessment. Previously, we demonstrated that constant TR pacing prevented alternans formation in isolated Langendorff-perfused rabbit hearts. However, the efficacy of "local" constant DI pacing vs "global" constant TR pacing in preventing alternans formation has never been investigated. Thus, the purpose of this study was to implement an ECG-based constant TR pacing in a 1D numerical model of human ventricular tissue and to compare the dynamical behavior of cardiac tissue with that resulted from a constant DI pacing. The results showed that both constant TR and constant DI pacing prevented the onset of alternans until lower basic cycle length when compared to periodic pacing. For longer cable lengths, constant TR pacing was shown to exhibit greater control on alternans than constant DI pacing.

Towards the Development of Nonlinear Approaches to Discriminate AF from NSR Using a Single-Lead ECG.

Paroxysmal atrial fibrillation (Paro. AF) is challenging to identify at the right moment. This disease is often undiagnosed using currently existing methods. Nonlinear analysis is gaining importance due to its capability to provide more insight into complex heart dynamics. The aim of this study is to use several recently developed nonlinear techniques to discriminate persistent AF (Pers. AF) from normal sinus rhythm (NSR), and more importantly, Paro. AF from NSR, using short-term single-lead electrocardiogram (ECG) signals. Specifically, we adapted and modified the time-delayed embedding method to minimize incorrect embedding parameter selection and further support to reconstruct proper phase plots of NSR and AF heart dynamics, from MIT-BIH databases. We also examine information-based methods, such as multiscale entropy (MSE) and kurtosis (Kt) for the same purposes. Our results demonstrate that embedding parameter time delay ( τ ), as well as MSE and Kt values can be successfully used to discriminate between Pers. AF and NSR. Moreover, we demonstrate that τ and Kt can successfully discriminate Paro. AF from NSR. Our results suggest that nonlinear time-delayed embedding method and information-based methods provide robust discriminating features to distinguish both Pers. AF and Paro. AF from NSR, thus offering effective treatment before suffering chaotic Pers. AF.

The dual effect of ephaptic coupling on cardiac conduction with heterogeneous expression of connexin 43.

Decreased and heterogeneous expression of connexin 43 (Cx43) are common features in animal heart failure models. Ephpatic coupling, which relies on the presence of junctional cleft space between the ends of adjacent cells, has been suggested to play a more active role in mediating intercellular electrical communication when gap junctions are reduced. To better understand the interplay of Cx43 expression and ephaptic coupling on cardiac conduction during heart failure, we performed numerical simulations on our model when Cx43 expression is reduced and heterogeneous. Under severely reduced Cx43 expression, we identified three new phenomena in the presence of ephaptic coupling: alternating conduction, in which ephaptic and gap junction-mediated mechanisms alternate; instability of planar fronts; and small amplitude action potential (SAP), which has a smaller potential amplitude than the normal action potential. In the presence of heterogeneous Cx43 expression, ephaptic coupling can either prevent or promote conduction block (CB) depending on the Cx43 knockout (Cx43KO) content. When Cx43KO content is relatively high, ephaptic coupling reduces the probabilities of CB. However, ephaptic coupling promotes CB when Cx43KO and wild type cells are mixed in roughly equal proportion, which can be attributed to an increase in current-to-load mismatch.

The role of short term memory and conduction velocity restitution in alternans formation.

Alternans is the periodic beat-to-beat short-long alternation in action potential duration (APD), which is considered to be a precursor of ventricular arrhythmias and sudden cardiac death. In extended cardiac tissue, electrical alternans can be either spatially concordant (SCA, all cells oscillate in phase) or spatially discordant (SDA, cells in different regions oscillate out of phase). SDA gives rise to an increase in the spatial dispersion of repolarization, which is thought to be proarrhythmic. In this paper, we investigated the effect of two aspects of short term memory (STM) (α, τ) and their interplay with conduction velocity (CV) restitution on alternans formation using numerical simulations of a mapping model with two beats of memory. Here, α quantifies the dependence of APD restitution on pacing history and τ characterizes APD accommodation, which is an exponential change of APD over time once basic cycle length (BCL) changes. Our main findings are as follows: In both single cell and spatially coupled homogeneous cable, the interplay between α and τ affects the dynamical behaviors of the system. For the case of large APD accommodation (τ ≥ 290 ms), increase in α leads to suppression of alternans. However, if APD accommodation is small (τ ≤ 250 ms), increase in α leads to appearance of additional alternans region. On the other hand, the slope of CV restitution does not change the regions of alternans in the cable. However, steep CV restitution leads to more complicated dynamical behaviors of the system. Specifically, SDA instead of SCA are observed. In addition, for steep CV restitution and sufficiently large τ, we observed formations of type II conduction block (CB2), transition from type I conduction block (CB1) to CB2, and unstable nodes.

Heart rate variability and alternans formation in the heart: The role of feedback in cardiac dynamics.

A beat-to-beat alternation in the action potential duration (APD) of myocytes, i.e. alternans, is believed to be a direct precursor of ventricular fibrillation in the whole heart. A common approach for the prediction of alternans is to construct the restitution curve, which is the nonlinear functional relationship between the APD and the preceding diastolic interval (DI). It was proposed that alternans appears when the magnitude of the slope of the restitution curve exceeds one, known as the restitution hypothesis. However, this restitution hypothesis was derived under the assumption of periodic stimulation, when there is a dependence of the DI on the immediate preceding APD (i.e. feedback). However, under physiological conditions, the heart rate exhibits substantial variations in time, known as heart rate variability (HRV), which introduces deviations from periodic stimulation in the system. In this manuscript, we investigated the role of HRV on alternans formation in isolated cardiac myocytes using numerical simulations of an ionic model of the cardiac action potential. We used this model with two different pacing protocols: a periodic pacing protocol with feedback and a protocol without feedback. We show that when HRV is incorporated in the periodic pacing protocol, it facilitated alternans formation in the isolated cell, but did not significantly change the magnitude of alternans. On the other hand, in the case of the pacing protocol without feedback, alternans formation was prevented, even in the presence of HRV.

Image-Decomposition-Enhanced Deep Learning for Detection of Rotor Cores in Cardiac Fibrillation.

OBJECTIVE: Rotors, regions of spiral wave reentry in cardiac tissues, are considered as the drivers of atrial fibrillation (AF), the most common arrhythmia. Whereas physics-based approaches have been widely deployed to detect the rotors, in-depth knowledge in cardiac physiology and electrogram interpretation skills are typically needed. The recent leap forward in smart sensing, data acquisition, and Artificial Intelligence (AI) has offered an unprecedented opportunity to transform diagnosis and treatment in cardiac ailment, including AF. This study aims to develop an image-decomposition-enhanced deep learning framework for automatic identification of rotor cores on both simulation and optical mapping data. METHODS: We adopt the Ensemble Empirical Mode Decomposition algorithm (EEMD) to decompose the original image, and the most representative component is then fed into a You-Only-Look-Once (YOLO) object-detection architecture for rotor detection. Simulation data from a bi-domain simulation model and optical mapping acquired from isolated rabbit hearts are used for training and validation. RESULTS: This integrated EEMD-YOLO model achieves high accuracy on both simulation and optical mapping data (precision: 97.2%, 96.8%, recall: 93.8%, 92.2%, and F1 score: 95.5%, 94.4%, respectively). CONCLUSION: The proposed EEMD-YOLO yields comparable accuracy in rotor detection with the gold standard in literature.

Uncoupling the mitochondria facilitates alternans formation in the isolated rabbit heart.

Alternans of action potential duration (APD) and intracellular calcium ([Ca²âº]i) transients in the whole heart are thought to be markers of increased propensity to ventricular fibrillation during ischemia-reperfusion injuries. During ischemia, ATP production is affected and the mitochondria become uncoupled, which may affect alternans formation in the heart. The aim of our study was to investigate the role of mitochondria on the formation of APD and [Ca²âº]i alternans in the isolated rabbit heart. We performed dual voltage and [Ca²âº]i optical mapping of isolated rabbit hearts under control conditions, global no-flow ischemia (n = 6), and after treatment with 50 nM of the mitochondrial uncoupler FCCP (n = 6). We investigated the formation of alternans of APD, [Ca²âº]i amplitude (CaA), and [Ca²âº]i duration (CaD) under different rates of pacing. We found that treatment with FCCP leads to the early occurrence of APD, CaD, and CaA alternans; an increase of intraventricular APD but not CaD heterogeneity; and significant reduction in conduction velocity compared with that of control. Furthermore, we demonstrated that FCCP and global ischemia have similar effects on the prolongation of [Ca²âº]i transients, whereas ischemia induces a significantly larger reduction of APD compared with that in FCCP treatment. In conclusion, our results demonstrate that uncoupling of mitochondria leads to an earlier occurrence of alternans in the heart. Thus, in conditions of mitochondrial stress, as seen during myocardial ischemia, uncoupled mitochondria may be responsible for the formation of both APD and [Ca²âº]i alternans in the heart, which in turn creates a substrate for ventricular arrhythmias.

Spatiotemporal evolution and prediction of [Ca(2+) ]i and APD alternans in isolated rabbit hearts.

INTRODUCTION: Action potential duration (APD) alternans can be accompanied by alternans in intracellular calcium transients ([Ca(2+) ]i ), leading to electromechanical alternans. Electromechanical alternans is considered a substrate for ventricular fibrillation. Although some techniques have been developed to predict APD alternans, the onset of [Ca(2+) ]i alternans has never been predicted. METHODS AND RESULTS: Simultaneous mapping of voltage and calcium was performed in 8 Langendorff-perfused rabbit hearts. APD, [Ca(2+) ]i amplitude (CaA) and duration (CaD) alternans were induced using a perturbed downsweep protocol. Local onset of alternans (B(onset) ) was defined as the cycle length (BCL) at which at least 10% of the RV exhibited alternans. We observed that the local onset of CaA alternans always occurred first, followed by APD and then CaD alternans. We constructed APD, CaD, and CaA restitution portraits for 2 regions of the heart defined at B(onset) : the 1:1alt region, which developed alternans, and the 1:1 region, which did not. Our results also show that the slopes S12 Max and SDyn were higher in 1:1alt region (SDyn  = 0.99 ± 0.04 vs 0.73 ± 0.06; S12 Max  = 0.95 ± 0.13 vs 0.65 ± 0.1, P < 0.05) prior to onset of CaD alternans, while S12 and S12 Max were significantly higher in the 1:1alt region (S12  = 0.59 ± 0.19 vs 0.19 ± 0.02; S12 Max  = 1.09 ± 0.1 vs 0.61 ± 0.08, P < 0.05) prior to onset of CaA alternans. CONCLUSION: We successfully applied the restitution portrait technique to the prediction of [Ca(2+) ]i (both CaA and CaD) alternans. The slopes of the APD/CaD/CaA restitution portrait are definitive indicators of APD, CaD, and CaA alternans.

Complexity analysis of electrical activity during endocardial and epicardial biventricular mapping of ventricular fibrillation.

BACKGROUND: Ventricular fibrillation (VF) is a lethal cardiac arrhythmia that is a significant cause of sudden cardiac death. Comprehensive studies of spatiotemporal characteristics of VF in situ are difficult to perform with current mapping systems and catheter technology. OBJECTIVE: The goal of this study was to develop a computational approach to characterize VF using a commercially available technology in a large animal model. Prior data suggests that characterization of spatiotemporal organization of electrical activity during VF can be used to provide better mechanistic understanding and potential ablation targets to modify VF and its substrate. We therefore evaluated intracardiac electrograms during biventricular mapping of the endocardium (ENDO) and epicardium (EPI) in acute canine studies. METHODS: To develop thresholds for organized and disorganized activity, a linear discriminant analysis (LDA)-based approach was performed to the known organized and disorganized activities recorded in ex vivo Langendorff-perfused rat and rabbit hearts using optical mapping experiments. Several frequency- and time-domain approaches were used as individual and paired features to identify the optimal thresholds for the LDA approach. Subsequently, VF was sequentially mapped in 4 canine hearts, using the CARTO mapping system with a multipolar mapping catheter in the ENDO left and right ventricles and EPI to capture the progression of VF at 3 discrete post-induction time intervals: VF period 1 (just after induction of VF to 15 min), VF period 2 (15 to 30 min), and VF period 3 (30 to 45 min). The developed LDA model, cycle lengths (CL), and regularity indices (RI) were applied to all recorded intracardiac electrograms to quantify the spatiotemporal organization of VF in canine hearts. RESULTS: We demonstrated the presence of organized activity in the EPI as VF progresses, in contrary to the ENDO, where the activity stays disorganized. The shortest CL always occurred in the ENDO, especially the RV, indicating a faster VF activity. The highest RI was found in the EPI in all hearts for all VF stages, indicating spatiotemporal consistency of RR intervals. CONCLUSION: We identified electrical organization and spatiotemporal differences throughout VF in canine hearts from induction to asystole. Notably, the RV ENDO is characterized by a high level of disorganization and faster VF frequency. In contrast, EPI has a high spatiotemporal organization of VF and consistently long RR intervals.

Interventricular heterogeneity as a substrate for arrhythmogenesis of decoupled mitochondria during ischemia in the whole heart.

Myocardial ischemia results in metabolic changes, which collapse the mitochondrial network, that increase the vulnerability of the heart to ventricular fibrillation (VF). It has been demonstrated at the single cell level that uncoupling the mitochondria using carbonyl cyanide p-(tri-fluoromethoxy)phenyl-hydrazone (FCCP) at low concentrations can be cardioprotective. The aim of our study was to investigate the effect of FCCP on arrhythmogenesis during ischemia in the whole rabbit heart. We performed optical mapping of isolated rabbit hearts (n = 33) during control and 20 min of global ischemia and reperfusion, both with and without pretreatment with the mitochondrial uncoupler FCCP at 100, 50, or 30 nM. No hearts went into VF during ischemia under the control condition, with or without the electromechanical uncoupler blebbistatin. We found that pretreatment with 100 (n = 4) and 50 (n = 6) nM FCCP, with or without blebbistatin, leads to VF during ischemia in all hearts, whereas pretreatment with 30 nM of FCCP led to three out of eight hearts going into VF during ischemia. We demonstrated that 30 nM of FCCP significantly increased interventricular (but not intraventricular) action potential duration and conduction velocity heterogeneity in the heart during ischemia, thus providing the substrate for VF. We showed that wavebreaks during VF occurred between the right and left ventricular junction. We also demonstrated that no VF occurred in the heart pretreated with 10 µM glibenclamide, which is known to abolish interventricular heterogeneity. Our results indicate that low concentrations of FCCP, although cardioprotective at the single cell level, are arrhythmogenic at the whole heart level. This is due to the fact that FCCP induces different electrophysiological changes to the right and left ventricle, thus increasing interventricular heterogeneity and providing the substrate for VF.

Visualizing the complex 3D geometry of the perfusion border zone in isolated rabbit heart.

Myocardial infarction, caused by a major blockage of a coronary artery, creates a border zone (BZ) between perfused and nonperfused tissue, which is believed to be the origin of fatal cardiac arrhythmias. We used a combination of optical clearing and polarization-sensitive optical coherence tomography to visualize a three-dimensional organization of the BZ in isolated rabbit hearts (n=5) at the microscopic level with a high spatial resolution. We found that the BZ has a complex three-dimensional structure with nonperfused areas penetrating into perfused tissue with finger-like projections. These "fingers" may play an important role in the initiation and maintenance of ventricular arrhythmias.

Mechanisms of arrhythmia termination during acute myocardial ischemia: Role of ephaptic coupling and complex geometry of border zone.

Myocardial ischemia occurs when blood flow to the heart is reduced, preventing the heart muscle from receiving enough oxygen required for survival. Several anatomical and electrophysiological changes occur at the ischemic core (IC) and border zone (BZ) during myocardial ischemia, for example, gap junctional remodeling, changes in ionic channel kinetics and electrophysiologic changes in cell excitability, which promote the development of cardiac arrhythmia. Ephaptic coupling (EpC), which is an electrical field effect developed in the shared cleft space between adjacent cells, has been suggested to rescue the conduction when gap junctions are impaired, such as myocardial ischemia. In this manuscript, we explored the impact of EpC, electrophysiological and anatomical components of myocardial ischemia on reentry termination during non-ischemic and ischemic condition. Our results indicated that EpC and BZ with complex geometry have opposite effects on the reentry termination. In particular, the presence of homogeneous EpC terminates reentry, whereas BZ with complex geometry alone facilitates reentry by producing wave break-up and alternating conduction block. The reentry is terminated in the presence of homogeneous or heterogeneous EpC despite the presence of complex geometry of the BZ, independent of the location of BZ. The inhibition of reentry can be attributed to a current-to-load mismatch. Our results points to an antiarrhythmic role of EpC and a pro-arrhythmic role of BZ with complex geometry.

Vitrification and Rewarming of Magnetic Nanoparticle-Loaded Rat Hearts.

To extend the preservation of donor hearts beyond the current 4-6 h, this paper explores heart cryopreservation by vitrification-cryogenic storage in a glass-like state. While organ vitrification is made possible by using cryoprotective agents (CPA) that inhibit ice during cooling, failure occurs during convective rewarming due to slow and non-uniform rewarming which causes ice crystallization and/or cracking. Here an alternative, "nanowarming", which uses silica-coated iron oxide nanoparticles (sIONPs) perfusion loaded through the vasculature is explored, that allows a radiofrequency coil to rewarm the organ quickly and uniformly to avoid convective failures. Nanowarming has been applied to cells and tissues, and a proof of principle study suggests it is possible in the heart, but proper physical and biological characterization especially in organs is still lacking. Here, using a rat heart model, controlled machine perfusion loading and unloading of CPA and sIONPs, cooling to a vitrified state, and fast and uniform nanowarming without crystallization or cracking is demonstrated. Further, nanowarmed hearts maintain histologic appearance and endothelial integrity superior to convective rewarming and indistinguishable from CPA load/unload control hearts while showing some promising organ-level (electrical) functional activity. This work demonstrates physically successful heart vitrification and nanowarming and that biological outcomes can be expected to improve by reducing or eliminating CPA toxicity during loading and unloading.