Human Arrhythmia Research Articles

Comparative expression analysis of Shox2-deficient embryonic stem cell-derived sinoatrial node-like cells

The homeodomain transcription factor Shox2 controls the development and function of the native cardiac pacemaker, the sinoatrial node (SAN). Moreover, SHOX2 mutations have been associated with cardiac arrhythmias in humans. For detailed examination of Shox2-dependent developmental mechanisms in SAN cells, we established a murine embryonic stem cell (ESC)-based model using Shox2 as a molecular tool. Shox2+/+ and Shox2−/− ESC clones were isolated and differentiated according to five different protocols in order to evaluate the most efficient enrichment of SAN-like cells. Expression analysis of cell subtype-specific marker genes revealed most efficient enrichment after CD166-based cell sorting. Comparative cardiac expression profiles of Shox2+/+ and Shox2−/− ESCs were examined by nCounter technology. Among other genes, we identified Nppb as a novel putative Shox2 target during differentiation in ESCs. Differential expression of Nppb could be confirmed in heart tissue of Shox2−/− embryos. Taken together, we established an ESC-based cardiac differentiation model and successfully purified Shox2+/+ and Shox2−/− SAN-like cells. This now provides an excellent basis for the investigation of molecular mechanisms under physiological and pathophysiological conditions for evaluating novel therapeutic approaches.

Denervation of the extrinsic cardiac sympathetic nervous system as a treatment modality for arrhythmia.

Denervation of the extrinsic cardiac sympathetic nervous system is a method of altering the autonomic tone experienced by the heart and vasculature. It has been studied and employed as a therapy for cardiac disease for decades. Currently, there is a high level of interest in using cardiac denervation for treatment of arrhythmias. This review describes the anatomy and physiology of the cardiac autonomic nervous system followed by a discussion of the mechanistic studies which provide a basis for the therapeutic use of sympathetic denervation. The clinical research supporting its use in human arrhythmias is then appraised, covering the standard indications, such as long QT syndrome, as well as future possibilities. Last, a detailed account of the methods for performing surgical cardiac denervation and percutaneous stellate ganglion anesthetic block is provided, including the complications of each procedure. An understanding of the anatomy and physiology of the cardiac autonomic nervous system along with the techniques of surgical denervation and percutaneous anesthetic block will allow the clinician to effectively discuss and implement these therapies.

Application of computer models on atrial fibrillation research.

Atrial fibrillation (AF) is a multifactorial and multiscale disease, where electrical, structural, anatomical and genetic factors contribute to the emergence of a complex and dynamic macroscopic phenotype. The incomplete understanding of AF mechanisms and the diversity and multiplicity of factors promoting the arrhythmia in humans have hampered the development of effective therapeutic approaches. Computer models and simulation of atrial arrhythmias may represent a unique ally to investigate AF mechanisms and direct therapeutic strategies, being able to merge and provide interpretation for multiscale data from cellular to entire organ scale levels. This review presents a broad overview of the principal modeling approaches applied in AF research to model atrial activity and ventricular response. The description of methodological aspects is followed by representative contributions of modeling to the dissection of AF mechanisms at both the atrial and atrioventricular level. A specific focus is given to controversial themes in AF research, such as calcium dynamics, fibrosis, multiple wavelets versus rotors propagation patterns, and AF heritability. Following modeling mechanistic insights, the review showcases modeling contributions in the domain of AF management and therapy, including the development of antiarrhythmic agents for rate and rhythm control, the optimization of ablation strategy, and the validation of mapping techniques and signal processing tools for the investigation of AF. A summary of current challenges and future developments necessary to improve the model capability at different scales and to transfer modeling results into clinical practice is finally presented.

Sirtuin 1 regulates cardiac electrical activity by deacetylating the cardiac sodium channel.

The voltage-gated cardiac Na+ channel (Nav1.5), encoded by the SCN5A gene, conducts the inward depolarizing cardiac Na+ current (INa) and is vital for normal cardiac electrical activity. Inherited loss-of-function mutations in SCN5A lead to defects in the generation and conduction of the cardiac electrical impulse and are associated with various arrhythmia phenotypes. Here we show that sirtuin 1 deacetylase (Sirt1) deacetylates Nav1.5 at lysine 1479 (K1479) and stimulates INa via lysine-deacetylation-mediated trafficking of Nav1.5 to the plasma membrane. Cardiac Sirt1 deficiency in mice induces hyperacetylation of K1479 in Nav1.5, decreases expression of Nav1.5 on the cardiomyocyte membrane, reduces INa and leads to cardiac conduction abnormalities and premature death owing to arrhythmia. The arrhythmic phenotype of cardiac-Sirt1-deficient mice recapitulated human cardiac arrhythmias resulting from loss of function of Nav1.5. Increased Sirt1 activity or expression results in decreased lysine acetylation of Nav1.5, which promotes the trafficking of Nav1.5 to the plasma membrane and stimulation of INa. As compared to wild-type Nav1.5, Nav1.5 with K1479 mutated to a nonacetylatable residue increases peak INa and is not regulated by Sirt1, whereas Nav1.5 with K1479 mutated to mimic acetylation decreases INa. Nav1.5 is hyperacetylated on K1479 in the hearts of patients with cardiomyopathy and clinical conduction disease. Thus, Sirt1, by deacetylating Nav1.5, plays an essential part in the regulation of INa and cardiac electrical activity.

Suppression of Arrhythmia by Enhancing Mitochondrial Calcium Uptake in Experimental Models of Catecholaminergic Ventricular Tachycardia

We have recently identified a critical role of mitochondria to shape intracellular Ca2+ signals and to regulate cardiac rhythmicity. Activation of mitochondrial Ca2+ uptake by efsevin, an agonist of the voltage-dependent anion channel 2 in the outer mitochondrial membrane, restored rhythmic cardiac contractions in a zebrafish cardiac arrhythmia model. Here we investigated the potential of pharmacological activation of mitochondrial Ca2+ uptake as a novel pharmacological strategy for human cardiac arrhythmia in a translational approach. To this aim we first used a murine model of ryanodine receptor 2 (RyR2)-mediated catecholaminergic polymorphic ventricular tachycardia (CPVT). In freshly isolated cardiomyocytes of RyR2R4496C/WT mice, harboring the human RyR2R4496C mutation associated with CPVT, efsevin restricted diastolic Ca2+ sparks and prevented the formation of propagating Ca2+ waves and spontaneous, diastolic action potentials. This anti-arrhythmic effect was abolished in the presence of mitochondrial Ca2+ uniporter (MCU) blocker Ru360 , but could be reproduced with the MCU activator kaempferol, demonstrating an immediate role of mitochondrial Ca2+ uptake for the anti-arrhythmic effect of efsevin. In RyR2R4496C/WT mice both mitochondrial Ca2+ uptake enhancers (MiCUps), efsevin and kaempferol, significantly reduced episodes of ventricular tachycardia after catecholaminergic stimulation by a bolus injection of epinephrine and caffeine in vivo while baseline ECG was unaffected. Finally, we used stem cell-derived cardiomyocytes from a CPVT patient to show efficacy of MiCUps in a human model. Both MiCUps abolished arrhythmogenic events in human CPVT cardiomyocytes. Our results demonstrate that enhancement of mitochondrial Ca2+ uptake by MiCUps is a promising pharmacological strategy for treatment and prevention of Ca2+-triggered arrhythmias, such as CPVT.

Early Cardioversion in Atrial Fibrillation: Earlier Is Better, but Not Always and (Maybe) Not Immediately.

Atrial fibrillation (AF) is the most common cardiac arrhythmia in humans. One of its important features is the tendency to become more persistent over time, even in the absence of underlying progressive heart disease. Conversion and maintenance of sinus rhythm by pharmacological or electrical methods become increasingly difficult the longer the arrhythmia persists. Electrical, mechanical, structural, and autonomic remodeling processes have been implicated in the mechanisms of AF initiation, perpetuation, and progression. Prevention or reversal of these remodeling processes can halt the progression of the disease. Cardioversion is a powerful tool and rhythm control is a widely used strategy in the management of AF. However, important questions remain unanswered regarding not only if, but also when to perform cardioversion. There are observations from past trials and clinical situations that support attempting to restore sinus rhythm, but further prospective randomized clinical trials are needed. Optimal timing of cardioversion remains somewhat uncertain, but it appears to be some time after the first few hours and before the first few months: the earlier, the better, but not always, and maybe not immediately, and it has to be tailored to the clinical situation and its many variables. This review is intended to summarize the evidence supporting early intervention for the prevention of remodeling in patients with AF.

HERG Channels: From Antitargets to Novel Targets for Cancer Therapy

In this issue of Clinical Cancer Research, evidence is provided on how to avoid cardiotoxicity when targeting hERG K+ channel for cancer therapy. hERG regulates different aspects of neoplastic progression. Although its blockade has effective anticancer effects in experimental models, it may lead to fatal arrhythmias in humans. Clin Cancer Res; 23(1); 3-5. ©2016 AACRSee related article by Pointer et al., p. 73.

Species-Dependent Mechanisms of Cardiac Arrhythmia: A Cellular Focus.

Although ventricular arrhythmia remains a leading cause of morbidity and mortality, available antiarrhythmic drugs have limited efficacy. Disappointing progress in the development of novel, clinically relevant antiarrhythmic agents may partly be attributed to discrepancies between humans and animal models used in preclinical testing. However, such differences are at present difficult to predict, requiring improved understanding of arrhythmia mechanisms across species. To this end, we presently review interspecies similarities and differences in fundamental cardiomyocyte electrophysiology and current understanding of the mechanisms underlying the generation of afterdepolarizations and reentry. We specifically highlight patent shortcomings in small rodents to reproduce cellular and tissue-level arrhythmia substrate believed to be critical in human ventricle. Despite greater ease of translation from larger animal models, discrepancies remain and interpretation can be complicated by incomplete knowledge of human ventricular physiology due to low availability of explanted tissue. We therefore point to the benefits of mathematical modeling as a translational bridge to understanding and treating human arrhythmia.

Scn2b Deletion in Mice Results in Ventricular and Atrial Arrhythmias.

Mutations in SCN2B, encoding voltage-gated sodium channel β2-subunits, are associated with human cardiac arrhythmias, including atrial fibrillation and Brugada syndrome. Because of this, we propose that β2-subunits play critical roles in the establishment or maintenance of normal cardiac electric activity in vivo. To understand the pathophysiological roles of β2 in the heart, we investigated the cardiac phenotype of Scn2b null mice. We observed reduced sodium and potassium current densities in ventricular myocytes, as well as conduction slowing in the right ventricular outflow tract region. Functional reentry, resulting from the interplay between slowed conduction, prolonged repolarization, and increased incidence of premature ventricular complexes, was found to underlie the mechanism of spontaneous polymorphic ventricular tachycardia. Scn5a transcript levels were similar in Scn2b null and wild-type ventricles, as were levels of Nav1.5 protein, suggesting that similar to the previous work in neurons, the major function of β2-subunits in the ventricle is to chaperone voltage-gated sodium channel α-subunits to the plasma membrane. Interestingly, Scn2b deletion resulted in region-specific effects in the heart. Scn2b null atria had normal levels of sodium current density compared with wild type. Scn2b null hearts were more susceptible to atrial fibrillation, had increased levels of fibrosis, and higher repolarization dispersion than wild-type littermates. Genetic deletion of Scn2b in mice results in ventricular and atrial arrhythmias, consistent with reported SCN2B mutations in human patients.

Transcriptional regulation of the sodium channel gene (SCN5A) by GATA4 in human heart

Aberrant expression of the sodium channel gene (SCN5A) has been proposed to disrupt cardiac action potential and cause human cardiac arrhythmias, but the mechanisms of SCN5A gene regulation and dysregulation still remain largely unexplored. To gain insight into the transcriptional regulatory networks of SCN5A, we surveyed the promoter and first intronic regions of the SCN5A gene, predicting the presence of several binding sites for GATA transcription factors (TFs). Consistent with this prediction, chromatin immunoprecipitation (ChIP) and sequential ChIP (Re-ChIP) assays show co-occupancy of cardiac GATA TFs GATA4 and GATA5 on promoter and intron 1 SCN5A regions in fresh-frozen human left ventricle samples. Gene reporter experiments show GATA4 and GATA5 synergism in the activation of the SCN5A promoter, and its dependence on predicted GATA binding sites. GATA4 and GATA6 mRNAs are robustly expressed in fresh-frozen human left ventricle samples as measured by highly sensitive droplet digital PCR (ddPCR). GATA5 mRNA is marginally but still clearly detected in the same samples. Importantly, GATA4 mRNA levels are strongly and positively correlated with SCN5A transcript levels in the human heart. Together, our findings uncover a novel mechanism of GATA TFs in the regulation of the SCN5A gene in human heart tissue. Our studies suggest that GATA5 but especially GATA4 are main contributors to SCN5A gene expression, thus providing a new paradigm of SCN5A expression regulation that may shed new light into the understanding of cardiac disease.

Evaluation of a novel high-resolution mapping system for catheter ablation of ventricular arrhythmias

The mapping of ventricular arrhythmias in humans using a minibasket 64-electrode catheter paired with a novel automatic mapping system (Rhythmia) has not been evaluated. The purpose of this study was to evaluate the safety and efficacy of mapping ventricular arrhythmias and clinical outcomes after ablation using this system. Electroanatomic maps for ventricular arrhythmias were obtained during 20 consecutive procedures in 19 patients (12 with ventricular tachycardia [VT] and 2 with ventricular ectopy [VE]). High-density maps were acquired using automatic beat acceptance and automatic system annotation of electrograms. Forty-seven electroanatomic maps (including 3 right ventricular and 9 epicardial maps) were obtained. Left ventricular endocardial mapping by transseptal (n = 13) and/or transaortic (n = 11) access was safe with no complications related to the minibasket catheter. VT substrate maps (n = 14; median 10,184 points) consistently demonstrated late potentials with high resolution. VT activation maps (n = 25; median 6401 points) obtained by automatic annotation included 7 complete maps (covering ≥90% of the tachycardia cycle length) in 5 patients in whom the entire VT circuit was accurately visualized. VE timing maps (n = 8) successfully localized the origin of VEs in all, with all accepted beats consistent with clinical VEs. Over a median follow-up of 10 months, no arrhythmia recurrence was noted in 75% after VT ablation and 86% after VE ablation. In this first human experience for ventricular arrhythmias using this system, ultra-high-density maps were created rapidly and safely, with a reliable automatic annotation of VT and consistent recording of abnormal electrograms. Medium-term outcomes after ablation were encouraging. Further larger studies are needed to validate these findings.

Methodology Considerations in Phase Mapping of Human Cardiac Arrhythmias.

Phase analysis of cardiac arrhythmias, particularly atrial fibrillation, has gained interest because of the ability to detect organized stable drivers (rotors) and target them for therapy. However, the lack of methodology details in publications on the topic has resulted in ongoing debate over the phase mapping technique. By comparing phase maps and activation maps, we examined advantages and limitations of phase mapping. Seven subjects were enrolled. We generated phase maps and activation maps from electrocardiographic imaging-reconstructed epicardial unipolar electrograms. For ventricular signals, phase was computed with (1) pseudoempirical mode decomposition detrending and (2) a novel Moving Average (MVG) detrending approach. For atrial fibrillation signals, MVG was modified to incorporate dynamic cycle length (DCL) changes (MVG-DCL). Phase maps were visually analyzed to study phase singularity points and rotors. Results show that phase is sensitive to cycle length choice, a limitation that was addressed by the MVG-DCL algorithm. MVG-DCL was optimal for atrial fibrillation analysis. Phase maps helped to highlight high-curvature wavefronts and rotors. However, for some activation patterns, phase generated nonrotational singularity points and false rotors. Phase mapping computes singularity points and visually highlights rotors. As such, it can help to provide a clearer picture of the spatiotemporal activation characteristics during atrial fibrillation. However, it is advisable to incorporate electrogram characteristics and the time-domain activation sequence in the analysis, to prevent misinterpretation and false rotor detection. Therefore, for mapping complex arrhythmias, a combined time-domain activation and phase mapping with variable cycle length seems to be the most reliable method.

Ultra Stable Dual Functionalized Gold Nanoparticles for the Effective Colorimetric Detection of Clenbuterol

Clenbuterol, a medium cumulative drug also appears as residues in animal would cause dizziness, fatigue, arrhythmia, and metabolic disorders in humans if eaten.1 Illegal abuse of clenbuterol was not stopped eventhough it has been banned from adding to animal feeds in many countries.2 Clenbuterol incident at Shanghai Co. (March 2011) caused great concern worldwide.3 Therefore, recently many sensor probes have been developed for the detection of clenbuterol. Among them, gold nanoparticles based detection was found to be promising.4, 5 However, to the best of our knowledge, dual functionalized AuNPs have not been reported for the detection of clenbuterol. Henceforth, we tend to develop ultra stable dual functionalized AuNPs for the effective detection of Clenbuterol (CLB). These newly ultra stable dual functionalized PE-Glu-AuNPs were developed by functionalization of AuNPs with Glutamic acid (Glu) and poly ethyleneimine (PE)via NaBH4 reduction method. We have elucidated the stability of PE-Glu-AuNPs over a period of six months. Further, the colorimetric assay of PE-Glu-AuNPs to CLB revealed that, its selective sensing ability starts from 300 nM visualized through naked eyes at pH 5. Additionally, the sub nanomolar detection of CLB has been estimated from linear fittings and standard deviation of UV-vis titration. The reaction mechanism was further verified through TEM images and dynamic light scattering (DLS) data. By DLS analysis, the initial size of PE-Glu-AuNPs was calculated to be 12.8 ± 8.6 nm, which later aggregated as 84.8 ± 52.3 nm during the detection of CLB-(C). It has also been well demonstrated that the probe does not evidence the selectivity towards any other interferences (A-Alamine; B-Phenylalanine; D-Nacl; E-CaCl2; F-Threonine; G-Cysteine; H-Glycine; I-Glucose and J-Urea). Moreover, the aggregation induced detection of CLB was well confirmed by TEM studies. References Brambilla, G.; Cenci, T.; Franconi, F.; Galarini, R.; Macrì, A.; Rondoni, F.; Strozzi, M.; Loizzo, A. Toxicol. Lett. 2000, 114, 47−53.Wu, K. S.; Guo, L.; Xua, W.; Xu, H. Y.; Aguilar, Z. P.; Xu, G. M.; Lai, W. H.; Xiong, H.; Wan, Y. Q. Talanta 2014, 129, 431−437. Yan, H.; Xu, D.; Meng, H.; Shi, L.; Li, L. Qual. Assur. Saf. Crops Foods 2015, 7, 27−35. Zhang, X. F.; Zhao, H.; Xue, Y.; Wu, Zh. J.; Zhang, Y.; He, Y. J.; Li, X. J.; Yuan, Zh. B. Biosens. Bioelectron. 2012, 34, 112−117. Kang, J.; Zhang, Y.; Li, X.; Miao, L.; Wu, A. ACS Appl. Mater. Interfaces 2016, 8, 1−5 Figure 1

Robotic magnetic navigation for ablation of human arrhythmias

Radiofrequency treatment represents the first choice of treatment for arrhythmias, in particular complex arrhythmias and especially atrial fibrillation, due to the greater benefit/risk ratio compared to antiarrhythmic drugs. However, complex arrhythmias such as atrial fibrillation require long procedures with additional risks such as X-ray exposure or serious complications such as tamponade. Given this context, the treatment of arrhythmias using robotic magnetic navigation entails a technique well suited to complex arrhythmias on account of its efficacy, reliability, significant reduction in X-ray exposure for both patient and operator, as well as a very low risk of perforation. As ongoing developments will likely improve results and procedure times, this technology will become one of the most modern technologies for treating arrhythmias. Based on the literature, this review summarizes the advantages and limitations of robotic magnetic navigation for ablation of human arrhythmias.

KCNN2 polymorphisms and cardiac tachyarrhythmias.

Potassium calcium-activated channel subfamily N member 2 (KCNN2) encodes an integral membrane protein that forms small-conductance calcium-activated potassium (SK) channels. Recent studies in animal models show that SK channels are important in atrial and ventricular repolarization and arrhythmogenesis. However, the importance of SK channels in human arrhythmia remains unclear. The purpose of the present study was to test the association between genetic polymorphism of the SK2 channel and the occurrence of cardiac tachyarrhythmias in humans. We enrolled 327 Han Chinese, including 72 with clinically significant ventricular tachyarrhythmias (VTa) who had a history of aborted sudden cardiac death (SCD) or unexplained syncope, 98 with a history of atrial fibrillation (AF), and 144 normal controls. We genotyped 12 representative tag single nucleotide polymorphisms (SNPs) across a 141-kb genetic region containing the KCNN2 gene; these captured the full haplotype information. The rs13184658 and rs10076582 variants of KCNN2 were associated with VTa in both the additive and dominant models (odds ratio [OR] 2.89, 95% confidence interval [CI] = 1.505-5.545, P = 0.001; and OR 2.55, 95% CI = 1.428-4.566, P = 0.002, respectively). After adjustment for potential risk factors, the association remained significant. The population attributable risks of these 2 variants of VTa were 17.3% and 10.6%, respectively. One variant (rs13184658) showed weak but significant association with AF in a dominant model (OR 1.91, CI = 1.025-3.570], P = 0.042). There was a significant association between the KCNN2 variants and clinically significant VTa. These findings suggest an association between KCNN2 and VTa; it also appears that KCNN2 variants may be adjunctive markers for risk stratification in patients susceptible to SCD.

The Future of Arrhythmias and Electrophysiology.

Cardiac arrhythmias are a major cause of morbidity and mortality in heart disease, and are the likely cause of more than a quarter of a million deaths annually in the U.S. alone. While our current approach to reducing suffering and death from arrhythmias relies largely on diagnosis and therapy aimed at early recognition and treatment, the benefits of prevention could be even more substantial. As we consider how the approach to arrhythmias may evolve in the coming decades, it is useful to consider the great breadth of presentations and scenarios that are encountered. Human arrhythmias occur in two scenarios. First, and particularly early in life, they arise as a consequence of inborn (congenital) constellations of anatomy and electrophysiology that change over time. There may be no structural cardiac abnormality (e.g., long QT syndrome), a minimal structural abnormality (e.g., an accessory pathway causing WPW) or a severe structural abnormality (e.g., endocardial cushion defect with heart block). Secondly, arrhythmias emerge later in life as a consequence of acquired disease (e.g., ventricular tachycardia late after myocardial infarction) or aging (atrial fibrillation). Genetic susceptibilities influence all aspects of the pathophysiology. As approaches to prevention and therapy evolve, genetic influences will receive increasing attention.

Pulmonary vein isolation in the treatment of atrial fibrillation

Pulmonary vein isolation in the treatment of atrial fibrillation Saurabh Kumar, Gregory F Michaud Cardiac Arrhythmia Service, Cardiovascular Division, Brigham and Women's Hospital, Boston, MA, USA Abstract: Atrial fibrillation (AF) is the commonest arrhythmia in humans and is associated with marked reduction in quality of life and an elevated thromboembolic risk. Paroxysmal, persistent, and permanent forms of AF have been recognized. Whilst antiarrhythmic drugs are considered as first-line therapy, the role of catheter ablation is increasing due to its superior efficacy in terms of quality of life and reduction in AF burden. The central paradigm for catheter ablation of AF is that triggers for AF are located near and within the pulmonary veins (PVs), and electrical isolation of the PVs from the left atrium forms the cornerstone of most catheter ablation strategies. Whilst paroxysmal form is generally trigger dependent, persistent and permanent forms are associated with variable interaction between triggers and "substrate" comprised of atrial and PV electrical and structural remodeling. Nevertheless, isolation of the PVs still forms a critical component of catheter ablation strategies, regardless of AF type. Procedural efficacy, however, is limited by PV conduction recovery. This is likely due to deficiencies in ablation tools or limitations of intraprocedural assessment of lesion efficacy. Careful attention to surrogates of tissue heating, such as impedance decrease and electrogram morphology changes, along with advances in catheter technology like contact force catheters may improve rates of durable PV isolation and single-procedural success. This review discusses the mechanism of paroxysmal AF with particular focus on the role of the PVs in AF initiation and PV isolation in the management of AF. Keywords: contact force, lesion transmurality, radiofrequency catheter ablation, paroxysmal atrial fibrillation, electrophysiology, AF

Dysfunction of the β2-spectrin-based pathway in human heart failure.

β2-Spectrin is critical for integrating membrane and cytoskeletal domains in excitable and nonexcitable cells. The role of β2-spectrin for vertebrate function is illustrated by dysfunction of β2-spectrin-based pathways in disease. Recently, defects in β2-spectrin association with protein partner ankyrin-B were identified in congenital forms of human arrhythmia. However, the role of β2-spectrin in common forms of acquired heart failure and arrhythmia is unknown. We report that β2-spectrin protein levels are significantly altered in human cardiovascular disease as well as in large and small animal cardiovascular disease models. Specifically, β2-spectrin levels were decreased in atrial samples of patients with atrial fibrillation compared with tissue from patients in sinus rhythm. Furthermore, compared with left ventricular samples from nonfailing hearts, β2-spectrin levels were significantly decreased in left ventricle of ischemic- and nonischemic heart failure patients. Left ventricle samples of canine and murine heart failure models confirm reduced β2-spectrin protein levels. Mechanistically, we identify that β2-spectrin levels are tightly regulated by posttranslational mechanisms, namely Ca(2+)- and calpain-dependent proteases. Furthermore, consistent with this data, we observed Ca(2+)- and calpain-dependent loss of β2-spectrin downstream effector proteins, including ankyrin-B in heart. In summary, our findings illustrate that β2-spectrin and downstream molecules are regulated in multiple forms of cardiovascular disease via Ca(2+)- and calpain-dependent proteolysis.

Safety pharmacology studies using EFP and impedance

IntroductionWhile extracellular field potential (EFP) recordings using multi-electrode arrays (MEAs) are a well-established technique for monitoring changes in cardiac and neuronal function, impedance is a relatively unexploited technology. The combination of EFP, impedance and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) has important implications for safety pharmacology as functional information about contraction and field potentials can be gleaned from human cardiomyocytes in a beating monolayer. The main objectives of this study were to demonstrate, using a range of different compounds, that drug effects on contraction and electrophysiology can be detected using a beating monolayer of hiPSC-CMs on the CardioExcyte 96. MethodshiPSC-CMs were grown as a monolayer on NSP-96 plates for the CardioExcyte 96 (Nanion Technologies) and recordings were made in combined EFP and impedance mode at physiological temperature. The effect of the hERG blockers, E4031 and dofetilide, hERG trafficking inhibitor, pentamidine, β-adrenergic receptor agonist, isoproterenol, and calcium channel blocker, nifedipine, was tested on the EFP and impedance signals. ResultsCombined impedance and EFP measurements were made from hiPSC-CMs using the CardioExcyte 96 (Nanion Technologies). E4031 and dofetilide, known to cause arrhythmia and Torsades de Pointes (TdP) in humans, decreased beat rate in impedance and EFP modes. Early afterdepolarization (EAD)-like events, an in vitro marker of TdP, could also be detected using this system. Isoproterenol and nifedipine caused an increase in beat rate. A long-term study (over 30h) of pentamidine, a hERG trafficking inhibitor, showed a concentration and time-dependent effect of pentamidine. DiscussionIn the light of the new Comprehensive in Vitro Proarrhythmia Assay (CiPA) initiative to improve guidelines and standardize assays and protocols, the use of EFP and impedance measurements from hiPSCs may become critical in determining the proarrhythmic risk of potential drug candidates. The combination of EFP offering information about cardiac electrophysiology, and impedance, providing information about contractility from the same area of a synchronously beating monolayer of human cardiomyocytes in a 96-well plate format has important implications for future cardiac safety testing.

A Computational Framework for Personalized Blood Flow Analysis in the Human Left Atrium

Atrial fibrillation (AF), the most common human arrhythmia, is a marker of an increased risk of embolic stroke. However, recent studies suggest that AF may not be mechanistically responsible for the stroke events. An alternative explanation for the mechanism of intracardiac thrombosis and stroke in patients with AF is structural remodeling of the left atrium (LA). Nevertheless, a mechanistic link between LA structural remodeling and intracardiac thrombosis is unclear, because there is no clinically feasible methodology to evaluate the complex relationship between these two phenomena in individual patients. Computational fluid dynamics (CFD) is a powerful tool that could potentially link LA structural remodeling and intracardiac thrombosis in individual patients by evaluating the patient-specific LA blood flow characteristics. However, the lack of knowledge of the material and mechanical properties of the heart wall in specific patients makes it challenging to solve the complexity of fluid-structure interaction. In this study, our aim was to develop a clinically feasible methodology to perform personalized blood flow analysis within the heart. We propose an alternative computational approach to perform personalized blood flow analysis by providing the three-dimensional LA endocardial surface motion estimated from patient-specific cardiac CT images. In two patients (case 1 and 2), a four-dimensional displacement vector field was estimated using nonrigid registration. The LA blood outflow across the mitral valve (MV) was calculated from the LV volume, and the flow field within the LA was derived from the incompressible Navier-Stokes equation. The CFD results successfully captured characteristic features of LA blood flow observed clinically by transesophageal echocardiogram. The LA global flow characteristics and vortex structures also agreed well with previous reports. The time course of LAA emptying was similar in both cases, despite the substantial difference in the LA structure and function. We conclude that our CT-based, personalized LA blood flow analysis is a clinically feasible methodology that can be used to improve our understanding of the mechanism of intracardiac thrombosis and stroke in individual patients with LA structural remodeling.