Marina Cerrone

Left Cardiac Sympathetic Denervation in the Management of High-Risk Patients Affected by the Long-QT Syndrome

Background—The management of long-QT syndrome (LQTS) patients who continue to have cardiac events (CEs) despite β-blockers is complex. We assessed the long-term efficacy of left cardiac sympathetic denervation (LCSD) in a group of high-risk patients. Methods and Results—We identified 147 LQTS patients who underwent LCSD. Their QT interval was very prolonged (QTc, 543±65 ms); 99% were symptomatic; 48% had a cardiac arrest; and 75% of those treated with β-blockers remained symptomatic. The average follow-up periods between first CE and LCSD and post-LCSD were 4.6 and 7.8 years, respectively. After LCSD, 46% remained asymptomatic. Syncope occurred in 31%, aborted cardiac arrest in 16%, and sudden death in 7%. The mean yearly number of CEs per patient dropped by 91% (P <0.001). Among 74 patients with only syncope before LCSD, all types of CEs decreased significantly as in the entire group, and a post-LCSD QTc <500 ms predicted very low risk. The percentage of patients with >5 CEs declined from 55% to 8% (P <0.001). In 5 patients with preoperative implantable defibrillator and multiple discharges, the post-LCSD count of shocks decreased by 95% (P =0.02) from a median number of 25 to 0 per patient. Among 51 genotyped patients, LCSD appeared more effective in LQT1 and LQT3 patients. Conclusions—LCSD is associated with a significant reduction in the incidence of aborted cardiac arrest and syncope in high-risk LQTS patients when compared with pre-LCSD events. However, LCSD is not entirely effective in preventing cardiac events including sudden cardiac death during long-term follow-up. LCSD should be considered in patients with recurrent syncope despite β-blockade and in patients who experience arrhythmia storms with an implanted defibrillator.

Bidirectional Ventricular Tachycardia and Fibrillation Elicited in a Knock-In Mouse Model Carrier of a Mutation in the Cardiac Ryanodine Receptor

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited disease characterized by adrenergically mediated polymorphic ventricular tachycardia leading to syncope and sudden cardiac death. The autosomal dominant form of CPVT is caused by mutations in the RyR2 gene encoding the cardiac isoform of the ryanodine receptor. In vitro functional characterization of mutant RyR2 channels showed altered behavior on adrenergic stimulation and caffeine administration with enhanced calcium release from the sarcoplasmic reticulum. As of today no experimental evidence is available to demonstrate that RyR2 mutations can reproduce the arrhythmias observed in CPVT patients. We developed a conditional knock-in mouse model carrier of the R4496C mutation, the mouse equivalent to the R4497C mutations identified in CPVT families, to evaluate if the animals would develop a CPVT phenotype and if beta blockers would prevent arrhythmias. Twenty-six mice (12 wild-type (WT) and 14RyRR4496C) underwent exercise stress testing followed by epinephrine administration: none of the WT developed ventricular tachycardia (VT) versus 5/14 RyRR4496C mice (P=0.02). Twenty-one mice (8 WT, 8 RyRR4496C, and 5 RyRR4496C pretreated with beta-blockers) received epinephrine and caffeine: 4/8 (50%) RyRR4496C mice but none of the WT developed VT (P=0.02); 4/5 RyRR4496C mice pretreated with propranolol developed VT (P=0.56 nonsignificant versus RyRR4496C mice). These data provide the first experimental demonstration that the R4496C RyR2 mutation predisposes the murine heart to VT and VF in response caffeine and/or adrenergic stimulation. Furthermore, the results show that analogous to what is observed in patients, beta adrenergic stimulation seems ineffective in preventing life-threatening arrhythmias.

Missense Mutations in Plakophilin-2 Cause Sodium Current Deficit and Associate With a Brugada Syndrome Phenotype

Background— Brugada syndrome (BrS) primarily associates with the loss of sodium channel function. Previous studies showed features consistent with sodium current (INa) deficit in patients carrying desmosomal mutations, diagnosed with arrhythmogenic cardiomyopathy (or arrhythmogenic right ventricular cardiomyopathy). Experimental models showed correlation between the loss of expression of desmosomal protein plakophilin-2 (PKP2) and reduced INa. We hypothesized that PKP2 variants that reduce INa could yield a BrS phenotype, even without overt structural features characteristic of arrhythmogenic right ventricular cardiomyopathy. Methods and Results— We searched for PKP2 variants in the genomic DNA of 200 patients with a BrS diagnosis, no signs of arrhythmogenic cardiomyopathy, and no mutations in BrS-related genes SCN5A, CACNa1c, GPD1L, and MOG1. We identified 5 cases of single amino acid substitutions. Mutations were tested in HL-1–derived cells endogenously expressing NaV1.5 but made deficient in PKP2 (PKP2-KD). Loss of PKP2 caused decreased INa and NaV1.5 at the site of cell contact. These deficits were restored by the transfection of wild-type PKP2, but not of BrS-related PKP2 mutants. Human induced pluripotent stem cell cardiomyocytes from a patient with a PKP2 deficit showed drastically reduced INa. The deficit was restored by transfection of wild type, but not BrS-related PKP2. Super-resolution microscopy in murine PKP2-deficient cardiomyocytes related INa deficiency to the reduced number of channels at the intercalated disc and increased separation of microtubules from the cell end. Conclusions— This is the first systematic retrospective analysis of a patient group to define the coexistence of sodium channelopathy and genetic PKP2 variations. PKP2 mutations may be a molecular substrate leading to the diagnosis of BrS.

Catecholaminergic polymorphic ventricular tachycardia: A paradigm to understand mechanisms of arrhythmias associated to impaired Ca2+ regulation

In the 8 years since the discovery of the genetic bases of catecholaminergic polymorphic ventricular tachycardia (CPVT), we have witnessed a remarkable improvement of knowledge on arrhythmogenic mechanisms involving disruption of cardiac Ca(2+) homeostasis. Studies on the consequences of RyR2 and CASQ2 mutations in cellular systems and mouse models have shed new light on pathways that are also implicated in arrhythmias occurring in highly prevalent diseases, such as heart failure. This research track has also led to the identification of therapeutic targets of potential clinical impact to abate the burden of sudden death in CPVT. Here, we review the current knowledge on the pathophysiology of CPVT also highlighting the existing controversies and possible future development.

KCNJ2 mutation in short QT syndrome 3 results in atrial fibrillation and ventricular proarrhythmia

We describe a mutation (E299V) in KCNJ2, the gene that encodes the strong inward rectifier K+ channel protein (Kir2.1), in an 11-y-old boy. The unique short QT syndrome type-3 phenotype is associated with an extremely abbreviated QT interval (200 ms) on ECG and paroxysmal atrial fibrillation. Genetic screening identified an A896T substitution in a highly conserved region of KCNJ2 that resulted in a de novo mutation E299V. Whole-cell patch-clamp experiments showed that E299V presents an abnormally large outward IK1 at potentials above −55 mV (P < 0.001 versus wild type) due to a lack of inward rectification. Coexpression of wild-type and mutant channels to mimic the heterozygous condition still resulted in a large outward current. Coimmunoprecipitation and kinetic analysis showed that E299V and wild-type isoforms may heteromerize and that their interaction impairs function. The homomeric assembly of E299V mutant proteins actually results in gain of function. Computer simulations of ventricular excitation and propagation using both the homozygous and heterozygous conditions at three different levels of integration (single cell, 2D, and 3D) accurately reproduced the electrocardiographic phenotype of the proband, including an exceedingly short QT interval with merging of the QRS and the T wave, absence of ST segment, and peaked T waves. Numerical experiments predict that, in addition to the short QT interval, absence of inward rectification in the E299V mutation should result in atrial fibrillation. In addition, as predicted by simulations using a geometrically accurate three-dimensional ventricular model that included the His–Purkinje network, a slight reduction in ventricular excitability via 20% reduction of the sodium current should increase vulnerability to life-threatening ventricular tachyarrhythmia.

Arrhythmogenic cardiomyopathy and Brugada syndrome: Diseases of the connexome

This review summarizes data in support of the notion that the cardiac intercalated disc is the host of a protein interacting network, called “the connexome”, where molecules classically defined as belonging to one particular structure (e.g., desmosomes, gap junctions, sodium channel complex) actually interact with others, and together, control excitability, electrical coupling and intercellular adhesion in the heart. The concept of the connexome is then translated into the understanding of the mechanisms leading to two inherited arrhythmia diseases: arrhythmogenic cardiomyopathy, and Brugada syndrome. The cross‐over points in these two diseases are addressed to then suggest that, though separate identifiable clinical entities, they represent “bookends” of a spectrum of manifestations that vary depending on the effect that a particular mutation has on the connexome as a whole.

Genetically engineered SCN5A mutant pig hearts exhibit conduction defects and arrhythmias

SCN5A encodes the α subunit of the major cardiac sodium channel Na(V)1.5. Mutations in SCN5A are associated with conduction disease and ventricular fibrillation (VF); however, the mechanisms that link loss of sodium channel function to arrhythmic instability remain unresolved. Here, we generated a large-animal model of a human cardiac sodium channelopathy in pigs, which have cardiac structure and function similar to humans, to better define the arrhythmic substrate. We introduced a nonsense mutation originally identified in a child with Brugada syndrome into the orthologous position (E558X) in the pig SCN5A gene. SCN5A(E558X/+) pigs exhibited conduction abnormalities in the absence of cardiac structural defects. Sudden cardiac death was not observed in young pigs; however, Langendorff-perfused SCN5A(E558X/+) hearts had an increased propensity for pacing-induced or spontaneous VF initiated by short-coupled ventricular premature beats. Optical mapping during VF showed that activity often began as an organized focal source or broad wavefront on the right ventricular (RV) free wall. Together, the results from this study demonstrate that the SCN5A(E558X/+) pig model accurately phenocopies many aspects of human cardiac sodium channelopathy, including conduction slowing and increased susceptibility to ventricular arrhythmias.

Multilevel analyses of SCN5A mutations in arrhythmogenic right ventricular dysplasia/cardiomyopathy suggest non-canonical mechanisms for disease pathogenesis.

Aims Arrhythmogenic Right Ventricular Dysplasia/Cardiomyopathy (ARVD/C) is often associated with desmosomal mutations. Recent studies suggest an interaction between the desmosome and sodium channel protein Nav1.5. We aimed to determine the prevalence and biophysical properties of mutations in SCN5A (the gene encoding Nav1.5) in ARVD/C. Methods and results We performed whole-exome sequencing in six ARVD/C patients (33% male, 38.2 ± 12.1 years) without a desmosomal mutation. We found a rare missense variant (p.Arg1898His; R1898H) in SCN5A in one patient. We generated induced pluripotent stem cell-derived cardiomyocytes (hIPSC-CMs) from the patient’s peripheral blood mononuclear cells. The variant was then corrected (R1898R) using Clustered Regularly Interspaced Short Palindromic Repeats/Cas9 technology, allowing us to study the impact of the R1898H substitution in the same cellular background. Whole-cell patch clamping revealed a 36% reduction in peak sodium current (P = 0.002); super-resolution fluorescence microscopy showed reduced abundance of NaV1.5 (P = 0.005) and N-Cadherin (P = 0.026) clusters at the intercalated disc. Subsequently, we sequenced SCN5A in an additional 281 ARVD/C patients (60% male, 34.8 ± 13.7 years, 52% desmosomal mutation-carriers). Five (1.8%) subjects harboured a putatively pathogenic SCN5A variant (p.Tyr416Cys, p.Leu729del, p.Arg1623Ter, p.Ser1787Asn, and p.Val2016Met). SCN5A variants were associated with prolonged QRS duration (119 ± 15 vs. 94 ± 14 ms, P < 0.01) and all SCN5A variant carriers had major structural abnormalities on cardiac imaging. Conclusions Almost 2% of ARVD/C patients harbour rare SCN5A variants. For one of these variants, we demonstrated reduced sodium current, Nav1.5 and N-Cadherin clusters at junctional sites. This suggests that Nav1.5 is in a functional complex with adhesion molecules, and reveals potential non-canonical mechanisms by which Nav1.5 dysfunction causes cardiomyopathy.

Genetics of ion-channel disorders.

Purpose of review In this article, we summarize the main features of the most common inherited channelopathies, focusing on the findings that advanced the field in the last few years. Recent findings The progress in genetics prompted the discovery of several new genes associated with ion-channel disorders, elucidating new molecular pathways and new arrhythmogenic mechanisms. The diffusion and availability of genetic screening gave a new relevance to the application of genetics not only for diagnosis, but also for risk assessment and therapeutic decisions. As a consequence, the present challenge in the field is represented by the need to use genetic data to develop personalized clinical approaches. Summary Over a few years, the field of inherited arrhythmogenic diseases has rapidly expanded, thus reshaping clinical management for these conditions. It is now clear that to handle these patients a specialized expertise is needed, able to translate the discoveries derived from basic science studies into the clinical care of the patients.

A Clinical Approach to Inherited Arrhythmias

In the past decade, the discovery that cases of ventricular arrhythmias and sudden cardiac death (SCD) in young individuals potentially could be caused by an unrecognized genetic substrate has defined a new subset of cardiac conditions: inherited arrhythmogenic diseases (IADs).1 Although rare in clinical practice, these diseases are more common than previously thought. They represent a challenge for the arrhythmia specialist in terms of diagnosis and clinical management. The correct diagnosis of IADs, as well as the use and interpretation of the results of genetic testing, are not straightforward and require a specific expertise such as that provided by specialized and dedicated centers. Here we will review the general issues arising from the correct interpretation of genetic testing and its indications. We also will focus on the clinical management and use of genetic information in different inherited arrhythmias. The term “channelopathies” defines a group of inherited arrhythmic syndromes caused by mutations on genes encoding for ion channel proteins and proteins that regulate ion channels.1 These mutations disrupt the balance of currents in the cardiac action potential, favoring the onset of life-threatening arrhythmias in the absence of structural heart defects. The long QT syndrome (LQTS) is a heritable channelopathy characterized by an exceedingly prolonged cardiac repolarization that may trigger ventricular arrhythmias and SCD.2 LQTS is one of the first channelopathies in which clinical and genetic features have been discovered, and a large series of patients have been collected and followed over the years.2–4 Thanks to these studies, LQTS management takes into account a patient’s genetic background and has become a paradigm for the use of genetic information applied to the clinical practice.3,4 Long QT syndrome can manifest with syncope and cardiac arrest. These commonly are triggered by adrenergic stress; however, roughly 10% …