Diagnosis, significance, and management of ventricular thrombi in patients referred for VT ablation.

INTRODUCTION: In patients with structural heart disease presenting with ventricular tachycardia (VT), detection of ventricular thrombi and subsequent management can be challenging. This study aimed to assess the value of multimodality imaging with cardiac magnetic resonance imaging (CMR), contrast-enhanced transthoracic echocardiography (TTE), and computed tomography (CT) for thrombus detection as well as a management algorithm geared towards anticoagulation and deferred ablation for patients referred for VT ablation. METHODS AND RESULTS: A total of 154 consecutive patients referred for VT ablation underwent preprocedural multimodality imaging with CMR, CT, and TTE. In 9 patients (6%) a new ventricular thrombus was detected and anticoagulation was initiated. Thrombi were detected by CMR in nine patients, by CT in seven patients, and by TTE in two patients. Five patients eventually underwent endocardial VT ablation procedures 6.0 ± 2.0 months after initiation of anticoagulation with one patient also requiring an epicardial approach. Two patients died while on anticoagulation, unrelated to ventricular arrhythmia. Four of five patients were rendered non-inducible and no testing was performed in 1/5 patients. Areas containing left ventricular thrombi were non-excitable with pacing. Six of thirty-two inducible VTs were mapped in close vicinity of ventricular thrombi. No clinical embolic events occurred during the ablation procedures. CONCLUSIONS: Ventricular thrombus was detected in 6% of consecutive patients with structural heart disease undergoing VT ablation. CMR was the most sensitive modality, while contrast-enhanced TTE failed to detect the majority of thrombi. Anticoagulation followed by ablation can be safely and successfully performed in patients with ventricular thrombi.

Wearables in Sports Cardiology.

The expanding array and adoption of consumer health wearables is creating a new dynamic to the patient-health-care provider relationship. Providers are increasingly tasked with integrating the biometric data collected from their patients into clinical care. Further, a growing body of evidence is supporting the provider-driven utility of wearables in the screening, diagnosis, and monitoring of cardiovascular disease. Here we highlight existing and emerging wearable health technologies and the potential applications for use within sports cardiology. We additionally highlight how wearables can advance the remote cardiovascular care of patients within the context of the COVID-19 pandemic. Finally, despite these promising advances, we acknowledge some of the significant challenges that remain before wearables can be routinely incorporated into clinical care.

Novel junctophilin-2 mutation A405S is associated with basal septal hypertrophy and diastolic dysfunction.

BACKGROUND: Hypertrophic cardiomyopathy (HCM), defined as asymmetric left ventricular hypertrophy, is a leading cause of cardiac death in the young. Perturbations in calcium (Ca2+) handling proteins have been implicated in the pathogenesis of HCM. JPH2-encoded junctophilin 2 is a major component of the junctional membrane complex, the subcellular microdomain involved in excitation-contraction coupling. We hypothesized that a novel JPH2 mutation identified in patients with HCM is causally linked to HCM, and alters intracellular Ca2+ signaling in a pro-hypertrophic manner. OBJECTIVES: To determine using a transgenic mouse model whether a JPH2 mutation found in a HCM patient is responsible for disease development. METHODS: Genetic interrogation of a large cohort of HCM cases was conducted for all coding exons of JPH2. Pseudo-knock-in (PKI) mice containing a novel JPH2 variant were subjected to echocardiography, cardiac MRI, hemodynamic analysis, and histology. RESULTS: A novel JPH2 mutation, A405S, was identified in a genotype-negative proband with significant basal septal hypertrophy. Although initially underappreciated by traditional echocardiographic imaging, PKI mice with this JPH2 mutation (residue A399S in mice) were found to exhibit similar basal hypertrophy using a newly developed echo imaging plane, and this was confirmed using cardiac MRI. Histological analysis demonstrated cardiomyocyte hypertrophy and disarray consistent with HCM. CONCLUSIONS: Variant A405S is a novel HCM-associated mutation in JPH2 found in a proband negative for mutations in the canonical HCM-associated genes. Studies in the analogous mouse model demonstrated for the first time a causal link between a JPH2 defect and HCM. Moreover, novel imaging approaches identified subvalvular septal hypertrophy, specific findings also reported in the human JPH2 mutation carrier.

Junctophilin-2 gene therapy rescues heart failure by normalizing RyR2-mediated Ca2+ release.

BACKGROUND: Junctophilin-2 (JPH2) is the primary structural protein for the coupling of transverse (T)-tubule associated cardiac L-type Ca channels and type-2 ryanodine receptors on the sarcoplasmic reticulum within junctional membrane complexes (JMCs) in cardiomyocytes. Effective signaling between these channels ensures adequate Ca-induced Ca release required for normal cardiac contractility. Disruption of JMC subcellular domains, a common feature of failing hearts, has been attributed to JPH2 downregulation. Here, we tested the hypothesis that adeno-associated virus type 9 (AAV9) mediated overexpression of JPH2 could halt the development of heart failure in a mouse model of transverse aortic constriction (TAC). METHODS AND RESULTS: Following TAC, a progressive decrease in ejection fraction was paralleled by a progressive decrease of cardiac JPH2 levels. AAV9-mediated expression of JPH2 rescued cardiac contractility in mice subjected to TAC. AAV9-JPH2 also preserved T-tubule structure. Moreover, the Ca2+ spark frequency was reduced and the Ca2+ transient amplitude was increased in AAV9-JPH2 mice following TAC, consistent with JPH2-mediated normalization of SR Ca2+ handling. CONCLUSIONS: This study demonstrates that AAV9-mediated JPH2 gene therapy maintained cardiac function in mice with early stage heart failure. Moreover, restoration of JPH2 levels prevented loss of T-tubules and suppressed abnormal SR Ca2+ leak associated with contractile failure following TAC. These findings suggest that targeting JPH2 might be an attractive therapeutic approach for treating pathological cardiac remodeling during heart failure.

Alterations in the interactome of serine/threonine protein phosphatase type-1 in atrial fibrillation patients.

BACKGROUND: Atrial fibrillation (AF) is the most common sustained cardiac arrhythmia, yet current pharmacological treatments are limited. Serine/threonine protein phosphatase type-1 (PP1), a major phosphatase in the heart, consists of a catalytic subunit (PP1c) and a large set of regulatory (R)-subunits that confer localization and substrate specificity to the holoenzyme. Previous studies suggest that PP1 is dysregulated in AF, but the mechanisms are unknown. OBJECTIVES: The purpose of this study was to test the hypothesis that PP1 is dysregulated in paroxysmal atrial fibrillation (PAF) at the level of its R-subunits. METHODS: Cardiac lysates were coimmunoprecipitated with anti-PP1c antibody followed by mass spectrometry-based, quantitative profiling of associated R-subunits. Subsequently, label-free quantification (LFQ) was used to evaluate altered R-subunit-PP1c interactions in PAF patients. R-subunits with altered binding to PP1c in PAF were further studied using bioinformatics, Western blotting (WB), immunocytochemistry, and coimmunoprecipitation. RESULTS: A total of 135 and 78 putative PP1c interactors were captured from mouse and human cardiac lysates, respectively, including many previously unreported interactors with conserved PP1c docking motifs. Increases in binding were found between PP1c and PPP1R7, cold-shock domain protein A (CSDA), and phosphodiesterase type-5A (PDE5A) in PAF patients, with CSDA and PDE5A being novel interactors validated by bioinformatics, immunocytochemistry, and coimmunoprecipitation. WB confirmed that these increases in binding cannot be ascribed to their changes in global protein expression alone. CONCLUSIONS: Subcellular heterogeneity in PP1 activity and downstream protein phosphorylation in AF may be attributed to alterations in PP1c-R-subunit interactions, which impair PP1 targeting to proteins involved in electrical and Ca(2+) remodeling. This represents a novel concept in AF pathogenesis and may provide more specific drug targets for treating AF.

The junctophilin family of proteins: from bench to bedside.

Excitable tissues rely on junctional membrane complexes to couple cell surface signals to intracellular channels. The junctophilins have emerged as a family of proteins critical in coordinating the maturation and maintenance of this cellular ultrastructure. Within skeletal and cardiac muscle, junctophilin 1 and junctophilin 2, respectively, couple sarcolemmal and intracellular calcium channels. In neuronal tissue, junctophilin 3 and junctophilin 4 may have an emerging role in coupling membrane neurotransmitter receptors and intracellular calcium channels. These important physiological roles are highlighted by the pathophysiology which results when these proteins are perturbed, and a growing body of literature has associated junctophilins with the pathogenesis of human disease.

Emerging roles of junctophilin-2 in the heart and implications for cardiac diseases.

Cardiomyocytes rely on a highly specialized subcellular architecture to maintain normal cardiac function. In a little over a decade, junctophilin-2 (JPH2) has become recognized as a cardiac structural protein critical in forming junctional membrane complexes (JMCs), which are subcellular domains essential for excitation-contraction coupling within the heart. While initial studies described the structure of JPH2 and its role in anchoring junctional sarcoplasmic reticulum and transverse-tubule (T-tubule) membrane invaginations, recent research has an expanded role of JPH2 in JMC structure and function. For example, JPH2 is necessary for the development of postnatal T-tubule in mammals. It is also critical for the maintenance of the complex JMC architecture and stabilization of local ion channels in mature cardiomyocytes. Loss of this function by mutations or down-regulation of protein expression has been linked to hypertrophic cardiomyopathy, arrhythmias, and progression of disease in failing hearts. In this review, we summarize current views on the roles of JPH2 within the heart and how JPH2 dysregulation may contribute to a variety of cardiac diseases.

Loss of microRNA-106b-25 cluster promotes atrial fibrillation by enhancing ryanodine receptor type-2 expression and calcium release.

BACKGROUND: Enhanced sarcoplasmic reticulum Ca(2+)-leak via ryanodine receptor type-2 (RyR2) contributes to the pathogenesis of atrial fibrillation (AF). Recent studies have shown that the level of RyR2 protein is elevated in atria of patients with paroxysmal AF, suggesting that microRNA-mediated post-transcriptional regulation of RyR2 might be an underlying mechanism. Bioinformatic analysis suggests that miR-106b and miR-93, members of the miR-106b-25 cluster, could bind to RyR2-3'-untranslated region and suppress its translation. Thus, we tested the hypothesis that loss of the miR-106b-25 cluster promotes AF via enhanced RyR2-mediated sarcoplasmic reticulum Ca(2+)-leak. METHODS AND RESULTS: Quantitative real-time polymerase chain reaction showed that the levels of mature miR-106b, miR-93, and miR-25 were lower in atria of patients with paroxysmal AF when compared with patients in sinus rhythm. In vitro assay showed that miR-93 reduced RyR2-3'-untranslated region luciferase activity. Total RyR2 protein in atrial tissue of miR-106b-25(-/-) mice was increased by 42% when compared with wild-type littermates but still maintained a normal subcellular distribution. Ca(2+)-spark frequency and total sarcoplasmic reticulum Ca(2+)-leak were increased in atrial myocytes of miR-106b-25(-/-) mice. Telemetry ECG recordings revealed that miR-106b-25(-/-) mice exhibited more frequent atrial ectopy and were also more susceptible to pacing-induced AF than wild-type littermates. Increased sarcoplasmic reticulum Ca(2+)-release and AF susceptibility in miR-106b-25(-/-) mice were abolished by the RyR2 blocker K201. CONCLUSIONS: These results suggest that miR-106b-25 cluster-mediated post-transcriptional regulation of RyR2 is a potential molecular mechanism involved in paroxysmal AF pathogenesis. As such, the miR-106b-25 cluster could be a novel gene-therapy target in AF associated with enhanced RyR2 expression.

Junctophilin-2 is necessary for T-tubule maturation during mouse heart development.

AIMS: Transverse tubules (TTs) provide the basic subcellular structures that facilitate excitation-contraction (EC) coupling, the essential process that underlies normal cardiac contractility. Previous studies have shown that TTs develop within the first few weeks of life in mammals but the molecular determinants of this development have remained elusive. This study aims to elucidate the role of junctophilin-2 (JPH2), a junctional membrane complex protein, in the maturation of TTs in cardiomyocytes. METHODS AND RESULTS: Using a novel cardiac-specific short-hairpin-RNA-mediated JPH2 knockdown mouse model (Mus musculus; αMHC-shJPH2), we assessed the effects of the loss of JPH2 on the maturation of the ventricular TT structure. Between embryonic day (E) 10.5 and postnatal day (P) 10, JPH2 mRNA and protein levels were reduced by >70% in αMHC-shJPH2 mice. At P8 and P10, knockdown of JPH2 significantly inhibited the maturation of TTs, while expression levels of other genes implicated in TT development remained mostly unchanged. At the same time, intracellular Ca(2+) handling was disrupted in ventricular myocytes from αMHC- shJPH2 mice, which developed heart failure by P10 marked by reduced ejection fraction, ventricular dilation, and premature death. In contrast, JPH2 transgenic mice exhibited accelerated TT maturation by P8. CONCLUSION: Our findings suggest that JPH2 is necessary for TT maturation during postnatal cardiac development in mice. In particular, JPH2 may be critical in anchoring the invaginating sarcolemma to the sarcoplasmic reticulum, thereby enabling the maturation of the TT network.

Mutation E169K in junctophilin-2 causes atrial fibrillation due to impaired RyR2 stabilization.

OBJECTIVES: This study sought to study the role of junctophilin-2 (JPH2) in atrial fibrillation (AF). BACKGROUND: JPH2 is believed to have an important role in sarcoplasmic reticulum (SR) Ca(2+) handling and modulation of ryanodine receptor Ca(2+) channels (RyR2). Whereas defective RyR2-mediated Ca(2+) release contributes to the pathogenesis of AF, nothing is known about the potential role of JPH2 in atrial arrhythmias. METHODS: Screening 203 unrelated hypertrophic cardiomyopathy patients uncovered a novel JPH2 missense mutation (E169K) in 2 patients with juvenile-onset paroxysmal AF (pAF). Pseudoknock-in (PKI) mouse models were generated to determine the molecular defects underlying the development of AF caused by this JPH2 mutation. RESULTS: PKI mice expressing E169K mutant JPH2 exhibited a higher incidence of inducible AF than wild type (WT)-PKI mice, whereas A399S-PKI mice expressing a hypertrophic cardiomyopathy-linked JPH2 mutation not associated with atrial arrhythmias were not significantly different from WT-PKI. E169K-PKI but not A399A-PKI atrial cardiomyocytes showed an increased incidence of abnormal SR Ca(2+) release events. These changes were attributed to reduced binding of E169K-JPH2 to RyR2. Atrial JPH2 levels in WT-JPH2 transgenic, nontransgenic, and JPH2 knockdown mice correlated negatively with the incidence of pacing-induced AF. Ca(2+) spark frequency in atrial myocytes and the open probability of single RyR2 channels from JPH2 knockdown mice was significantly reduced by a small JPH2-mimicking oligopeptide. Moreover, patients with pAF had reduced atrial JPH2 levels per RyR2 channel compared to sinus rhythm patients and an increased frequency of spontaneous Ca(2+) release events. CONCLUSIONS: Our data suggest a novel mechanism by which reduced JPH2-mediated stabilization of RyR2 due to loss-of-function mutation or reduced JPH2/RyR2 ratios can promote SR Ca(2+) leak and atrial arrhythmias, representing a potential novel therapeutic target for AF.

Pathogenesis of lethal cardiac arrhythmias in Mecp2 mutant mice: implication for therapy in Rett syndrome.

Rett syndrome is a neurodevelopmental disorder typically caused by mutations in methyl-CpG-binding protein 2 (MECP2) in which 26% of deaths are sudden and of unknown cause. To explore the hypothesis that these deaths may be due to cardiac dysfunction, we characterized the electrocardiograms in 379 people with Rett syndrome and found that 18.5% show prolongation of the corrected QT interval (QTc), an indication of a repolarization abnormality that can predispose to the development of an unstable fatal cardiac rhythm. Male mice lacking MeCP2 function, Mecp2(Null/Y), also have prolonged QTc and show increased susceptibility to induced ventricular tachycardia. Female heterozygous null mice, Mecp2(Null/+), show an age-dependent prolongation of QTc associated with ventricular tachycardia and cardiac-related death. Genetic deletion of MeCP2 function in only the nervous system was sufficient to cause long QTc and ventricular tachycardia, implicating neuronally mediated changes to cardiac electrical conduction as a potential cause of ventricular tachycardia in Rett syndrome. The standard therapy for prolonged QTc in Rett syndrome, ß-adrenergic receptor blockers, did not prevent ventricular tachycardia in Mecp2(Null/Y) mice. To determine whether an alternative therapy would be more appropriate, we characterized cardiomyocytes from Mecp2(Null/Y) mice and found increased persistent sodium current, which was normalized when cells were treated with the sodium channel-blocking anti-seizure drug phenytoin. Treatment with phenytoin reduced both QTc and sustained ventricular tachycardia in Mecp2(Null/Y) mice. These results demonstrate that cardiac abnormalities in Rett syndrome are secondary to abnormal nervous system control, which leads to increased persistent sodium current. Our findings suggest that treatment in people with Rett syndrome would be more effective if it targeted the increased persistent sodium current to prevent lethal cardiac arrhythmias.