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Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): - Grant from the Spanish Ministerio de Economía y Competitividad - FPI Severo Ochoa Predoctoral grant Introduction Catecholaminergic polymorphic ventricular tachycardia-2 (CPVT2) is an arrhythmogenic inherited autosomal recessive disorder caused by mutations in the gene encoding cardiac calsequestrin (CASQ2). The homozygous mouse model CASQ2R33Q/R33Q closely mimics the clinical phenotype of patients that carried this mutation, which causes a substantial imbalance in the homeostasis of intracellular calcium, resulting in polymorphic ventricular tachycardia through triggered activity. CASQ2-R33Q and triadin (TRDN) levels are markedly reduced with no changes in the mRNA expression levels. So far, the mechanisms underlying the decrease of these proteins are not completely understood. Deciphering these mechanisms will help us to develop new or personalized therapeutic options for CPVT patients. Purpose The purpose of this study was to identify the molecular mechanisms regulating the decrease of the couplon proteins in the CASQ2R33Q/R33Q mice to find new therapeutic options. Methods Mass spectrometry proteomics and Western blot analysis were performed to explore the role of the main cell proteolytic systems – the proteasome, autophagy and calpain. Experiments were performed in adult mice and in isolated neonatal cardiomyocytes treated with inhibitors of these systems. To explore the role of calcium homeostasis in the degradation of these proteins, mice were treated with a RYR2 inhibitor. Results CASQ2-R33Q and TRDN proteins showed a shorter half-life in CASQ2R33Q/R33Q mice. Mass spectrometry proteomics revealed an increase in proteins that participate in Endoplasmic reticulum stress (ER stress) and changes in calcium release and uptake regulators. Western blot analysis of myocardial tissue from adult mice demonstrated an alteration in the activity of the proteasome, the autophagy flux and the calpain enzyme. Experiments in isolated neonatal cardiomyocytes showed that CASQ2-R33Q, but not TRDN, degradation can be blocked with proteasome, ERAD (ER associated degradation) or autophagy inhibitors. In contrast, both CASQ2-R33Q and TRDN degradation were prevented when the calpain enzyme is inhibited. Inhibition of RYR2 in vivo induced an increase in CASQ2-R33Q and TRDN protein levels. Conclusions We propose that changes in calcium homeostasis lead to the activation of some of the main cell proteolytic systems – proteasome and calpain. CASQ2-R33Q is degraded by the proteasome through the ERAD pathway that is activated as a consequence of ER stress. Calpain inhibition is the only mechanism that recovers both CASQ2-R33Q and TRDN protein levels. In the near future, in vivo ECGs and in vitro calcium transient experiments will determine their therapeutic potential for the treatment of recessive CPVT. [ABSTRACT FROM AUTHOR]

Informatics for Integrating Biology and the Bedside (i2b2) is an initiative funded by the NIH that aims at building an informatics infrastructure to support biomedical research. The University of Pavia has recently integrated i2b2 infrastructure with a registry of inherited arrhythmogenic diseases. Within this project, the authors created a novel i2b2 cell, named R Engine Cell, which allows the communication between i2b2 and the R statistical software. As survival analyses are routinely performed by cardiology researchers, the authors have first concentrated on making Kaplan-Meier analyses available within the i2b2 web interface. To this aim, the authors developed a web-client plug-in to select the patient set on which to perform the analysis and to display the results in a graphical, intuitive way. R Engine Cell has been designed to easily support the integration of other R-based statistical analyses into i2b2. [ABSTRACT FROM AUTHOR]

Cardiac Diseases Caused by SCN5A Mutations. A prerequisite for a normal cardiac function is a proper generation and propagation of electrical impulses. Contraction of the heart is obtained through a delicate matched transmission of the electrical impulses. A pivotal element of the impulse propagation is the depolarizing sodium current, responsible for the initial depolarization of the cardiomyocytes. Recent research has shown that mutations in the SCN5A gene, encoding the cardiac sodium channel Nav1.5, are associated with both rare forms of ventricular arrhythmia, as well as the most frequent form of arrhythmia, atrial fibrillation (AF). In this comprehensive review, we describe the functional role of Nav1.5 and its associated proteins in propagation and depolarization both in a normal- and in a pathophysiological setting. Furthermore, several of the arrhythmogenic diseases, such as long-QT syndrome, Brugada syndrome, and AF, reported to be associated with mutations in SCN5A, are thoroughly described. (J Cardiovasc Electrophysiol, Vol. 21, pp. 107-115, January 2010) [ABSTRACT FROM AUTHOR]

Family studies, and more recent molecular genetic investigations, indicate that the Wolff-Parkinson-White (WPW) syndrome and associated preexcitation disorders can have a substantial genetic component. Because preexcitation disorders are sometimes inherited as single gene disorders, key mechanistic insights can be gained that are expected to be relevant also to the more common multifactorial forms of these traits. Potentially, such insights will inform the future management of these conditions. Where WPW is inherited as a familial trait, with or without associated cardiac defects or a systemic syndrome, there are clinical genetic ramifications that are already of practical importance. [ABSTRACT FROM AUTHOR]

Context: Data on the efficacy of beta-blockers in the 3 most common genetic long QT syndrome (LQTS) loci are limited.Objective: To describe and assess outcome in a large systematically genotyped population of beta-blocker-treated LQTS patients.Design, Setting, and Patients: Consecutive LQTS-genotyped patients (n = 335) in Italy treated with beta-blockers for an average of 5 years.Main Outcome Measures: Cardiac events (syncope, ventricular tachycardia/torsades de pointes, cardiac arrest, and sudden cardiac death) while patients received beta-blocker therapy according to genotype.Results: Cardiac events among patients receiving beta-blocker therapy occurred in 19 of 187 (10%) LQT1 patients, 27 of 120 (23%) LQT2 patients, and 9 of 28 (32%) LQT3 patients (P<.001). The risk of cardiac events was higher among LQT2 (adjusted relative risk, 2.81; 95% confidence interval [CI], 1.50-5.27; P =.001) and LQT3 (adjusted relative risk, 4.00; 95% CI, 2.45-8.03; P<.001) patients than among LQT1 patients, suggesting inadequate protection from beta-blocker therapy. Other important predictors of risk were a QT interval corrected for heart rate that was more than 500 ms in patients receiving therapy (adjusted relative risk, 2.01; 95% CI, 1.16-3.51; P =.01) and occurrence of a first cardiac event before the age of 7 years (adjusted RR, 4.34; 95% CI, 2.35-8.03; P<.001).Conclusion: Among patients with genetic LQTS treated with beta-blockers, there is a high rate of cardiac events, particularly among patients with LQT2 and LQT3 genotypes. [ABSTRACT FROM AUTHOR]

Purpose: Spontaneous sarcoplasmic reticulum (SR) Ca2+ release through defective cardiac ryanodine receptor (RyR2) promotes contractile dysfunction and myocardial remodeling in response to pressure overload. We tested the hypothesis that RyR2 stabilizer K201 attenuates the SR Ca2+ leak and ameliorates pressure overload-induced cardiac remodeling in mice carrying RyR2 gain-of-function mutation.Methods: RyR2R4496C+/- knock-in mice (R4496C) and their wild-type (WT) littermates underwent surgery without (Sham) or with transaortic banding (TAB). Changes in Ca2+ spark frequency (as a measure for the SR Ca2+ leak) were monitored by means of confocal Ca2+ imaging (Fluo-4/AM, linescan mode) in isolated ventricular cardiomyocytes (1 week post-surgery) under control conditions and after the pre-incubation (≥ 1-hour) with 300 nmol/L K201 or 1 mmol/L dantrolene (positive control). K201 (12 mg/kg/day) or DMSO (vehicle) was administered in R4496C-TAB mice for 4 weeks via osmotic minipumps. K201 plasma concentration was determined by using high performance liquid chromatography. Structural dimensions and function of the left ventricle (LV) were assessed by transthoracic echocardiography 1 and 3 weeks after surgery. Data are shown as mean±S.E.M.Results: K201 markedly reduced the increase in Ca2+ spark frequency at matched SR Ca2+ load in R4496C-TAB (83.9±8.6 pL-1s-1 vs. 332.7±42.9 pL-1s-1) and WT-TAB cells (82.2±6.5 pL-1s-1 vs. 190.1±27 pL-1s-1) as well as R4496C-Sham cells (72.1±7.5 pL-1s-1 vs. 173.6±20.9 pL-1s-1, all P<0.05 versus control, respectively). The RyR2 stabilizing effect was more pronounced in the presence of K201 than dantrolene. Chronic infusion of K201 resulted in a mean K201 plasma concentration of 163±25 nmol/L. In R4496C-TAB mice, K201 prevented the decline in fractional shortening (1 week: 24.5±2.6% vs. 17.9±1.8%; 3 weeks: 21.3±2.6% vs. 15.2±2.4%, both P<0.05 versus vehicle-treated R4496C-TAB, respectively), restored an increase in the relative wall thickness (1 week: 0.44±0.02 mm vs. 0.40±0.01 mm; 3 weeks: 0.46±0.02 mm vs. 0.36±0.03 mm) and largely ameliorated LV dilatation (LV end-systolic diameter, 1 week: 3.10±0.15 mm vs. 3.75±0.26 mm; 3 weeks: 3.35±0.19 mm vs.4.23±0.31 mm, all P<0.05 versus vehicle-treated R4496C-TAB, respectively). K201 treatment reduced mortality over 6 weeks after TAB: 83% of K201-treated R4496C-TAB mice were alive vs. 31% treated with the vehicle (P<0.05).Conclusions: Pharmacological stabilization of SR Ca2+ release by K201 ameliorates the development of heart failure, preserves systolic function and improves survival in R4496C-TAB mice. [ABSTRACT FROM PUBLISHER]