Genetic background determines the severity of age-dependent cardiac structural abnormalities and arrhythmia susceptibility in Scn5a-1798insD mice.

AIMS: Patients with mutations in SCN5A encoding NaV1.5 often display variable severity of electrical and structural alterations, but the underlying mechanisms are not fully elucidated. We here investigate the combined modulatory effect of genetic background and age on disease severity in the Scn5a1798insD/+ mouse model. METHODS AND RESULTS: In vivo electrocardiogram and echocardiograms, ex vivo electrical and optical mapping, and histological analyses were performed in adult (2-7 months) and aged (8-28 months) wild-type (WT) and Scn5a1798insD/+ (mutant, MUT) mice from the FVB/N and 129P2 inbred strains. Atrio-ventricular (AV) conduction, ventricular conduction, and ventricular repolarization are modulated by strain, genotype, and age. An aging effect was present in MUT mice, with aged MUT mice of both strains showing prolonged QRS interval and right ventricular (RV) conduction slowing. 129P2-MUT mice were severely affected, with adult and aged 129P2-MUT mice displaying AV and ventricular conduction slowing, prolonged repolarization, and spontaneous arrhythmias. In addition, the 129P2 strain appeared particularly susceptible to age-dependent electrical, functional, and structural alterations including RV conduction slowing, reduced left ventricular (LV) ejection fraction, RV dilatation, and myocardial fibrosis as compared to FVB/N mice. Overall, aged 129P2-MUT mice displayed the most severe conduction defects, RV dilatation, and myocardial fibrosis, in addition to the highest frequency of spontaneous arrhythmia and inducible arrhythmias. CONCLUSION: Genetic background and age both modulate disease severity in Scn5a1798insD/+ mice and hence may explain, at least in part, the variable disease expressivity observed in patients with SCN5A mutations. Age- and genetic background-dependent development of cardiac structural alterations furthermore impacts arrhythmia risk. Our findings therefore emphasize the importance of continued assessment of cardiac structure and function in patients carrying SCN5A mutations.

A Possible Role of Tetrodotoxin-Sensitive Na+ Channels for Oxidation-Induced Late Na+ Currents in Cardiomyocytes.

An accumulation of reactive oxygen species (ROS) in cardiomyocytes can induce pro-arrhythmogenic late Na+ currents by removing the inactivation of voltage-gated Na+ channels including the tetrodotoxin (TTX)-resistant cardiac α-subunit Nav1.5 as well as TTX-sensitive α-subunits like Nav1.2 and Nav1.3. Here, we explored oxidant-induced late Na+ currents in mouse cardiomyocytes and human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) as well as in HEK 293 cells expressing Nav1.2, Nav1.3, or Nav1.5. Na+ currents in mouse cardiomyocytes and hiPSC-CMs treated with the oxidant chloramine T (ChT) developed a moderate reduction in peak current amplitudes accompanied by large late Na+ currents. While ChT induced a strong reduction in peak current amplitudes but only small persistent currents on Nav1.5, both Nav1.2 and Nav1.3 produced increased peak current amplitudes and large persistent currents following oxidation. TTX (300 nM) blocked ChT-induced late Na+ currents significantly stronger as compared to peak Na+ currents in both mouse cardiomyocytes and hiPSC-CMs. Similar differences between Nav1.2, Nav1.3, and Nav1.5 regarding ROS sensitivity were also evident when oxidation was induced with UVA-light (380 nm) or the cysteine-selective oxidant nitroxyl (HNO). To conclude, our data on TTX-sensitive Na+ channels expressed in cardiomyocytes may be relevant for the generation of late Na+ currents following oxidative stress.

Knockdown of SCN5A alters metabolic-associated genes and aggravates hypertrophy in the cardiomyoblast.

SCN5A mutations have been reported to cause various cardiomyopathies in humans. Most of the SCN5A mutations causes loss of function and thereby, alters the overall cellular function. Therefore, to understand the loss of SCN5A function in cardiomyocytes, we have knocked down the SCN5A gene (SCN5A-KD) in H9c2 cells and explored the cell phenotype and molecular behaviors in the presence and absence of isoproterenol (ISO), an adrenergic receptor agonist that induces cardiac hypertrophy. Expression of several genes related to hypertrophy, inflammation, fibrosis, and energy metabolism pathways were evaluated. It was found that the mRNA expression of hypertrophy-related gene, brain (B-type) natriuretic peptide (BNP) was significantly increased in SCN5A-KD cells as compared to 'control' H9c2 cells. There was a further increase in the mRNA expressions of BNP and ßMHC in SCN5A-KD cells after ISO treatment compared to their respective controls. Pro-inflammatory cytokine, tumor necrosis factor-alpha expression was significantly increased in 'SCN5A-KD' H9c2 cells. Further, metabolism-related genes like glucose transporter type 4, cluster of differentiation 36, peroxisome proliferator-activated receptor alpha, and peroxisome proliferator-activated receptor-gamma were significantly elevated in the SCN5A-KD cells as compared to the control cells. Upregulation of these metabolic genes is associated with increased ATP production. The study revealed that SCN5A knock-down causes alteration of gene expression related to cardiac hypertrophy, inflammation, and energy metabolism pathways, which may promote cardiac remodelling and cardiomyopathy.

Impacts of DCM-linked gating pore currents on the electrophysiological characteristics of hiPSC-CM monolayers.

BACKGROUND: Variants of the SCN5A gene, which encodes the NaV1.5 cardiac sodium channel, have been linked to arrhythmic disorders associated with dilated cardiomyopathy (DCM). However, the precise pathological mechanisms remain elusive. The present study aimed to elucidate the pathophysiological consequences of the DCM-linked Nav1.5/R219H variant, which is known to generate a gating pore current, using patient-specific human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) cultured in monolayers. METHODS: Ventricular- and atrial-like hiPSC-CM monolayers were generated from DCM patients carrying the R219H SCN5A variant as well as from healthy control individuals. CRISPR-corrected hiPSC-CMs served as isogenic controls. Simultaneous optical mapping of action potentials (APs) and calcium transients (CaTs) was employed to measure conduction velocities (CVs) and AP durations (APDs) and served as markers of electrical excitability. Calcium handling was evaluated by assessing CaT uptake (half-time to peak), recapture (tau of decay), and durations (TD50 and TD80). A multi-electrode array (MEA) analysis was conducted on hiPSC-CM monolayers to measure field potential (FP) parameters, including corrected Fridericia FP durations (FPDc). RESULTS: Our results revealed that CVs were significantly reduced by more than 50 % in both ventricular- and atrial-like hiPSC-CM monolayers carrying the R219H variant compared to the control group. APDs were also prolonged in the R219H group compared to the control and CRISPR-corrected groups. CaT uptake, reuptake, and duration were also markedly delayed in the R219H group compared to the control and CRISPR-corrected groups in both the ventricular- and the atrial-like hiPSC-CM monolayers. Lastly, the MEA data revealed a notably prolonged FPDc in the ventricular- and atrial-like hiPSC-CMs carrying the R219H variant compared to the control and isogenic control groups. CONCLUSIONS: These findings highlight the impact of the gating pore current on AP propagation and calcium homeostasis within a functional syncytium environment and offer valuable insights into the potential mechanisms underlying DCM pathophysiology.

CardioDPi: An explainable deep-learning model for identifying cardiotoxic chemicals targeting hERG, Cav1.2, and Nav1.5 channels.

The cardiotoxic effects of various pollutants have been a growing concern in environmental and material science. These effects encompass arrhythmias, myocardial injury, cardiac insufficiency, and pericardial inflammation. Compounds such as organic solvents and air pollutants disrupt the potassium, sodium, and calcium ion channels cardiac cell membranes, leading to the dysregulation of cardiac function. However, current cardiotoxicity models have disadvantages of incomplete data, ion channels, interpretability issues, and inability of toxic structure visualization. Herein, an interpretable deep-learning model known as CardioDPi was developed, which is capable of discriminating cardiotoxicity induced by the human Ether-à-go-go-related gene (hERG) channel, sodium channel (Na_v1.5), and calcium channel (Ca_v1.5) blockade. External validation yielded promising area under the ROC curve (AUC) values of 0.89, 0.89, and 0.94 for the hERG, Na_v1.5, and Ca_v1.5 channels, respectively. The CardioDPi can be freely accessed on the web server CardioDPipredictor (http://cardiodpi.sapredictor.cn/). Furthermore, the structural characteristics of cardiotoxic compounds were analyzed and structural alerts (SAs) can be extracted using the user-friendly CardioDPi-SAdetector web service (http://cardiosa.sapredictor.cn/). CardioDPi is a valuable tool for identifying cardiotoxic chemicals that are environmental and health risks. Moreover, the SA system provides essential insights for mode-of-action studies concerning cardiotoxic compounds.

Brugada Syndrome and Pulmonary Atresia with Intact Interventricular Septum: Fortuitous Finding or New Genetic Connection?

Brugada syndrome is a rare arrhythmogenic syndrome associated mainly with pathogenic variants in the SCN5A gene. Right ventricle outflow tract fibrosis has been reported in some cases of patients diagnosed with Brugada syndrome. Pulmonary atresia with an intact ventricular septum is characterized by the lack of a functional pulmonary valve, due to the underdevelopment of the right ventricle outflow tract. We report, for the first time, a 4-year-old boy with pulmonary atresia with an intact ventricular septum who harbored a pathogenic de novo variant in SCN5A, and the ajmaline test unmasked a type-1 Brugada pattern. We suggest that deleterious variants in the SCN5A gene could be implicated in pulmonary atresia with an intact ventricular septum embryogenesis, leading to overlapping phenotypes.

Pediatric and Familial Genetic Arrhythmia Syndromes: SCN5A-Related Disorders When It Is Not Long QT Type 3: Clinical Signs and Symptoms.

The following case series presents three different pediatric patients with SCN5A-related disease. In addition, family members are presented to demonstrate the variable penetrance that is commonly seen. Identifying features of this disease is important, because even in the very young, SCN5A disorders can cause lethal arrhythmias and sudden death.

Molecular Autopsy With Banked Cord Blood Reveals Brugada Syndrome in Past Sudden Death Case.

Molecular autopsy has recently been gaining attention as a means of postmortem diagnosis; however, it is usually performed using the victim's blood sample at the time of death. Here, we report the first case of a deceased infant with Brugada syndrome whose diagnosis was made with banked cord blood. A seemingly healthy 1-year-old male infant collapsed while having a fever; this collapse was witnessed by his mother. Despite cardiopulmonary resuscitation, he died of ventricular fibrillation. No abnormalities of cardiac structure were identified on autopsy. Genomic samples were not stored at the time because of a lack of suspicion for familial arrhythmia. Five years later, his sister showed Brugada electrocardiogram pattern while febrile from Kawasaki disease. Their father showed a spontaneous type 1 Brugada electrocardiogram pattern. A heterozygous SCN5A p.R893C variant was found by genetic testing in the proband's father and sister. Furthermore, the proband's genetic testing was performed using his banked cord blood, which identified the same variant. Family history of Brugada syndrome with an SCN5A-R893C variant and clinical evidence led to a postmortem diagnosis of Brugada syndrome in the proband. Identification of this variant in this case later contributed to verifying SCN5A-R893C as a pathogenic variant through data accumulation. Banked cord blood may prove useful for conducting molecular autopsies in previously undiagnosed cases of sudden death in which genomic samples were not stored.

Variable Penetrance and Expressivity of a Rare Pore Loss-of-Function Mutation (p.L889V) of Nav1.5 Channels in Three Spanish Families.

A novel rare mutation in the pore region of Nav1.5 channels (p.L889V) has been found in three unrelated Spanish families that produces quite diverse phenotypic manifestations (Brugada syndrome, conduction disease, dilated cardiomyopathy, sinus node dysfunction, etc.) with variable penetrance among families. We clinically characterized the carriers and recorded the Na+ current (INa) generated by p.L889V and native (WT) Nav1.5 channels, alone or in combination, to obtain further insight into the genotypic-phenotypic relationships in patients carrying SCN5A mutations and in the molecular determinants of the Nav1.5 channel function. The variant produced a strong dominant negative effect (DNE) since the peak INa generated by p.L889V channels expressed in Chinese hamster ovary cells, either alone (-69.4 ± 9.0 pA/pF) or in combination with WT (-62.2 ± 14.6 pA/pF), was significantly (n ≥ 17, p < 0.05) reduced compared to that generated by WT channels alone (-199.1 ± 44.1 pA/pF). The mutation shifted the voltage dependence of channel activation and inactivation to depolarized potentials, did not modify the density of the late component of INa, slightly decreased the peak window current, accelerated the recovery from fast and slow inactivation, and slowed the induction kinetics of slow inactivation, decreasing the fraction of channels entering this inactivated state. The membrane expression of p.L889V channels was low, and in silico molecular experiments demonstrated profound alterations in the disposition of the pore region of the mutated channels. Despite the mutation producing a marked DNE and reduction in the INa and being located in a critical domain of the channel, its penetrance and expressivity are quite variable among the carriers. Our results reinforce the argument that the incomplete penetrance and phenotypic variability of SCN5A loss-of-function mutations are the result of a combination of multiple factors, making it difficult to predict their expressivity in the carriers despite the combination of clinical, genetic, and functional studies.

Extracellular acidification reveals the antiarrhythmic properties of amiodarone related to late sodium current-induced atrial arrhythmia.

BACKGROUND: Amiodarone (AMIO) is an antiarrhythmic drug with the pKa in the physiological range. Here, we explored how mild extracellular pH (pHe) changes shape the interaction of AMIO with atrial tissue and impact its pharmacological properties in the classical model of sea anemone sodium channel neurotoxin type 2 (ATX) induced late sodium current (INa-Late) and arrhythmias. METHOD: Isolated atrial cardiomyocytes from male Wistar rats and human embryonic kidney cells expressing SCN5A Na+ channels were used for patch-clamp experiments. Isolated right atria (RA) and left atria (LA) tissue were used for bath organ experiments. RESULTS: A more acidophilic pHe caused negative inotropic effects on isolated RA and LA atrial tissue, without modification of the pharmacological properties of AMIO. A pHe of 7.0 changed the sodium current (INa) related components of the action potential (AP), which was enhanced in the presence of AMIO. ATXinduced arrhythmias in isolated RA and LA. Also, ATX prolonged the AP duration and enhanced repolarization dispersion in isolated cardiomyocytes in both pHe 7.4 and pHe 7.0. Pre-incubation of the isolated RA and LA and isolated atrial cardiomyocytes with AMIO prevented arrhythmias induced by ATX only at a pHe of 7.0. Moreover, AMIO was able to block INa-Late induced by ATX only at a pHe of 7.0. CONCLUSION: The pharmacological properties of AMIO concerning healthy rat atrial tissue are not dependent on pHe. However, the prevention of arrhythmias induced by INa-Late is pHe-dependent. The development of drugs analogous to AMIO with charge stabilization may help to create more effective drugs to treat arrhythmias related to the INa-Late.

Phosphorylation of cardiac sodium channel at Ser571 anticipates manifestations of the aging myopathy.

Diastolic dysfunction and delayed ventricular repolarization are typically observed in the elderly, but whether these defects are intimately associated with the progressive manifestation of the aging myopathy remains to be determined. In this regard, aging in experimental animals is coupled with increased late Na+ current (INa,L) in cardiomyocytes, raising the possibility that INa,L conditions the modality of electrical recovery and myocardial relaxation of the aged heart. For this purpose, aging male and female wild-type (WT) C57Bl/6 mice were studied together with genetically engineered mice with phosphomimetic (gain of function, GoF) or ablated (loss of function, LoF) mutations of the sodium channel Nav1.5 at Ser571 associated with, respectively, increased and stabilized INa,L. At ∼18 mo of age, WT mice developed prolonged duration of the QT interval of the electrocardiogram and impaired diastolic left ventricular (LV) filling, defects that were reversed by INa,L inhibition. Prolonged repolarization and impaired LV filling occurred prematurely in adult (∼5 mo) GoF mutant mice, whereas these alterations were largely attenuated in aging LoF mutant animals. Ca2+ transient decay and kinetics of myocyte shortening/relengthening were delayed in aged (∼24 mo) WT myocytes, with respect to adult cells. In contrast, delayed Ca2+ transients and contractile dynamics occurred at adult stage in GoF myocytes and further deteriorated in old age. Conversely, myocyte mechanics were minimally affected in aging LoF cells. Collectively, these results document that Nav1.5 phosphorylation at Ser571 and the late Na+ current modulate the modality of myocyte relaxation, constituting the mechanism linking delayed ventricular repolarization and diastolic dysfunction.NEW & NOTEWORTHY We have investigated the impact of the late Na current (INa,L) on cardiac and myocyte function with aging by using genetically engineered animals with enhanced or stabilized INa,L, due to phosphomimetic or phosphoablated mutations of Nav1.5. Our findings support the notion that phosphorylation of Nav1.5 at Ser571 prolongs myocardial repolarization and impairs diastolic function, contributing to the manifestations of the aging myopathy.

Apelin-13 reverses bupivacaine-induced cardiotoxicity: an experimental study.

INTRODUCTION: Cardiac arrest or arrhythmia caused by bupivacaine may be refractory to treatment. Apelin has been reported to directly increase the frequency of spontaneous activation and the propagation of action potentials, ultimately promoting cardiac contractility. This study aimed to investigate the effects of apelin-13 in reversing cardiac suppression induced by bupivacaine in rats. METHODS: A rat model of cardiac suppression was established by a 3-min continuous intravenous infusion of bupivacaine at the rate of 5 mg.kg-1.min-1, and serial doses of apelin-13 (50, 150 and 450 µg.kg-1) were administered to rescue cardiac suppression to identify its dose-response relationship. We used F13A, an inhibitor of Angiotensin Receptor-Like 1 (APJ), and Protein Kinase C (PKC) inhibitor chelerythrine to reverse the effects of apelin-13. Moreover, the protein expressions of PKC, Nav1.5, and APJ in ventricular tissues were measured using Western blotting and immunofluorescence assay. RESULTS: Compared to the control rats, the rats subjected to continuous intravenous administration of bupivacaine had impaired hemodynamic stability. Administration of apelin-13, in a dose-dependent manner, significantly improved hemodynamic parameters in rats with bupivacaine-induced cardiac suppression (p < 0.05), and apelin-13 treatment also significantly upregulated the protein expressions of p-PKC and Nav1.5 (p < 0.05), these effects were abrogated by F13A or chelerythrine (p < 0.05). CONCLUSION: Exogenous apelin-13, at least in part, activates the PKC signaling pathway through the apelin/APJ system to improve cardiac function in a rat model of bupivacaine-induced cardiac suppression.

Exenatide reduces atrial fibrillation susceptibility by inhibiting hKv1.5 and hNav1.5 channels.

Exenatide, a promising cardioprotective agent, protects against cardiac structural remodeling and diastolic dysfunction. Combined blockade of sodium and potassium channels is valuable for managing atrial fibrillation (AF). Here, we explored whether exenatide displayed anti-AF effects by inhibiting human Kv1.5 and Nav1.5 channels. We used the whole-cell patch-clamp technique to investigate the effects of exenatide on hKv1.5 and hNav1.5 channels expressed in human embryonic kidney 293 cells and studied the effects of exenatide on action potential (AP) and other cardiac ionic currents in rat atrial myocytes. Additionally, an electrical mapping system was used to explore the effects of exenatide on electrical properties and AF activity in isolated rat hearts. Finally, a rat AF model, established using acetylcholine and calcium chloride, was employed to evaluate the anti-AF potential of exenatide in rats. Exenatide reversibly suppressed IKv1.5 with IC50 of 3.08 µM, preferentially blocked the hKv1.5 channel in its closed state, and positively shifted the voltage-dependent activation curve. Exenatide also reversibly inhibited INav1.5 with IC50 of 3.30 µM, negatively shifted the voltage-dependent inactivation curve, and slowed its recovery from inactivation with significant use-dependency at 5 and 10 Hz. Furthermore, exenatide prolonged AP duration and suppressed the sustained K+ current (Iss) and transient outward K+ current (Ito), but without inhibition of L-type Ca2+ current (ICa,L) in rat atrial myocytes. Exenatide prevented AF incidence and duration in rat hearts and rats. These findings demonstrate that exenatide inhibits IKv1.5 and INav1.5in vitro and reduces AF susceptibility in isolated rat hearts and rats.

Molecular genetic screening after non-ischaemic sudden cardiac arrest and no overt cardiomyopathy in real life: A major tool for the aetiological diagnostic work-up.

BACKGROUND: With the development of advanced sequencing techniques, genetic testing has emerged as a valuable tool for the work-up of non-ischaemic sudden cardiac arrest (SCA). AIMS: To evaluate the effectiveness of genetic testing in patients with unexplained SCA, according to clinical phenotype. METHODS: All patients who underwent molecular genetic testing for non-ischaemic SCA with no left ventricular cardiomyopathy between 2012 and 2021 in two French university hospitals were included. RESULTS: Of 66 patients (mean age 36.7±11.9years, 54.5% men), 21 (31.8%; 95% confidence interval 22.4-45.3%) carried a genetic variant: eight (12.1%) had a pathogenic or likely pathogenic (P/LP) variant and 13 (19.7%) had a variant of uncertain significance (VUS). Among 37 patients (56.1%) with no phenotypic clues, genetic testing identified a P/LP variant in five (13.5%), mainly in RYR2 (n=3) and SCN5A (n=2), and a VUS in nine (24.3%). None of the nine patients with phenotypic evidence of channelopathies had P/LP variants, but two had VUS in RYR2 and NKX2.5. Among the 20 patients with suspected arrhythmogenic cardiomyopathy, three P/LP variants (15.0%) and two VUS (10.0%) were found in DSC2, PKP2, SCN5A and DSG2, TRPM4, respectively. Genetic testing was performed sooner after cardiac arrest (P<0.001) and results were obtained more rapidly (P=0.02) after versus before 2016. CONCLUSION: This study highlights the utility of molecular genetic testing with a genetic variant of interest identified in one-third of patients with unexplained SCA. Genetic testing was beneficial even in patients without phenotypic clues, with one-fourth of patients carrying a P/LP variant that could have direct implications.

The dispensability of 14-3-3 proteins for the regulation of human cardiac sodium channel Nav1.5.

BACKGROUND: 14-3-3 proteins are ubiquitous proteins that play a role in cardiac physiology (e.g., metabolism, development, and cell cycle). Furthermore, 14-3-3 proteins were proposed to regulate the electrical function of the heart by interacting with several cardiac ion channels, including the voltage-gated sodium channel Nav1.5. Given the many cardiac arrhythmias associated with Nav1.5 dysfunction, understanding its regulation by the protein partners is crucial. AIMS: In this study, we aimed to investigate the role of 14-3-3 proteins in the regulation of the human cardiac sodium channel Nav1.5. METHODS AND RESULTS: Amongst the seven 14-3-3 isoforms, only 14-3-3η (encoded by YWHAH gene) weakly co-immunoprecipitated with Nav1.5 when heterologously co-expressed in tsA201 cells. Total and cell surface expression of Nav1.5 was however not modified by 14-3-3η overexpression or inhibition with difopein, and 14-3-3η did not affect physical interaction between Nav1.5 α-α subunits. The current-voltage relationship and the amplitude of Nav1.5-mediated sodium peak current density were also not changed. CONCLUSIONS: Our findings illustrate that the direct implication of 14-3-3 proteins in regulating Nav1.5 is not evident in a transformed human kidney cell line tsA201.

Genetic Testing in Brugada Syndrome: A 30-Year Experience.

BACKGROUND: A pathogenic/likely pathogenic variant can be found in 20% to 25% of patients with Brugada syndrome (BrS) and a pathogenic/likely pathogenic variant in SCN5A is associated with a worse prognosis. The aim of this study is to define the diagnostic yield of a large gene panel with American College of Medical Genetics and Genomics variant classification and to assess prognosis of SCN5A and non-SCN5A variants. METHODS: All patients with BrS, were prospectively enrolled in the Universitair Ziekenhuis Brussel registry between 1992 and 2022. Inclusion criteria for the study were (1) BrS diagnosis; (2) genetic analysis performed with a large gene panel; (3) classification of variants following American College of Medical Genetics and Genomics guidelines. Patients with a pathogenic/likely pathogenic variant in SCN5A were defined as SCN5A+. Patients with a reported variant in a non-SCN5A gene or with no reported variants were defined as patients with SCN5A-. All variants were classified as missense or predicted loss of function. RESULTS: A total of 500 BrS patients were analyzed. A total of 104 patients (20.8%) were SCN5A+ and 396 patients (79.2%) were SCN5A-. A non-SCN5A gene variant was found in 75 patients (15.0%), of whom, 58 patients (77.3%) had a missense variant and 17 patients (22.7%) had a predicted loss of function variant. At a follow-up of 84.0 months, 48 patients (9.6%) experienced a ventricular arrhythmia (VA). Patients without any variant had higher VA-free survival, compared with carriers of a predicted loss of function variant in SCN5A+ or non-SCN5A genes. There was no difference in VA-free survival between patients without any variant and missense variant carriers in SCN5A+ or non-SCN5A genes. At Cox analysis, SCN5A+ or non-SCN5A predicted loss of function variant was an independent predictor of VA. CONCLUSIONS: In a large BrS cohort, the yield for SCN5A+ is 20.8%. A predicted loss of function variant carrier is an independent predictor of VA.

Cardiac GR Mediates the Diurnal Rhythm in Ventricular Arrhythmia Susceptibility.

BACKGROUND: Ventricular arrhythmias (VAs) demonstrate a prominent day-night rhythm, commonly presenting in the morning. Transcriptional rhythms in cardiac ion channels accompany this phenomenon, but their role in the morning vulnerability to VAs and the underlying mechanisms are not understood. We investigated the recruitment of transcription factors that underpins transcriptional rhythms in ion channels and assessed whether this mechanism was pertinent to the heart's intrinsic diurnal susceptibility to VA. METHODS AND RESULTS: Assay for transposase-accessible chromatin with sequencing performed in mouse ventricular myocyte nuclei at the beginning of the animals' inactive (ZT0) and active (ZT12) periods revealed differentially accessible chromatin sites annotating to rhythmically transcribed ion channels and distinct transcription factor binding motifs in these regions. Notably, motif enrichment for the glucocorticoid receptor (GR; transcriptional effector of corticosteroid signaling) in open chromatin profiles at ZT12 was observed, in line with the well-recognized ZT12 peak in circulating corticosteroids. Molecular, electrophysiological, and in silico biophysically-detailed modeling approaches demonstrated GR-mediated transcriptional control of ion channels (including Scn5a underlying the cardiac Na+ current, Kcnh2 underlying the rapid delayed rectifier K+ current, and Gja1 responsible for electrical coupling) and their contribution to the day-night rhythm in the vulnerability to VA. Strikingly, both pharmacological block of GR and cardiomyocyte-specific genetic knockout of GR blunted or abolished ion channel expression rhythms and abolished the ZT12 susceptibility to pacing-induced VA in isolated hearts. CONCLUSIONS: Our study registers a day-night rhythm in chromatin accessibility that accompanies diurnal cycles in ventricular myocytes. Our approaches directly implicate the cardiac GR in the myocyte excitability rhythm and mechanistically link the ZT12 surge in glucocorticoids to intrinsic VA propensity at this time.

Scar architecture affects the electrophysiological characteristics of induced ventricular arrhythmias in hypertrophic cardiomyopathy.

AIMS: Late gadolinium enhancement cardiac magnetic resonance (LGE-CMR) detects myocardial scarring, a risk factor for ventricular arrhythmias (VAs) in hypertrophic cardiomyopathy (HCM). The LGE-CMR distinguishes core, borderzone (BZ) fibrosis, and BZ channels, crucial components of re-entry circuits. We studied how scar architecture affects inducibility and electrophysiological traits of VA in HCM. METHODS AND RESULTS: We correlated scar composition with programmed ventricular stimulation-inducible VA features using LGE intensity maps. Thirty consecutive patients were enrolled. Thirteen (43%) were non-inducible, 6 (20%) had inducible non-sustained, and 11 (37%) had inducible sustained mono (MMVT)- or polymorphic VT/VF (PVT/VF). Of 17 induced VA, 13 (76%) were MMVT that either ended spontaneously, persisted as sustained monomorphic, or degenerated into PVT/VF. Twenty-seven patients (90%) had LGE. Of these, 17 (57%) had non-sustained or sustained inducible VA. Scar mass significantly increased (P = 0.002) from non-inducible to inducible non-sustained and sustained VA patients in both the BZ and core components. Borderzone channels were found in 23%, 67%, and 91% of non-inducible, inducible non-sustained, and inducible sustained VA patients (P = 0.003). All 13 patients induced with MMVT or monomorphic-initiated PVT/VF had LGE. The origin of 10/13 of these VTs matched scar location, with 8/10 of these LGE regions showing BZ channels. During follow-up (20 months, interquartile range: 7-37), one patient with BZ channels and inducible PVT had an ICD shock for VF. CONCLUSION: Scar architecture determines inducibility and electrophysiological traits of VA in HCM. Larger studies should explore the role of complex LGE patterns in refining risk assessment in HCM patients.

Polymorphic Variants of SCN5A Gene (rs41312433 and rs1805124) Associated with Coronary Artery Affliction in Patients with Severe Arrhythmias.

Several mutations in this gene for the α subunit of the cardiac sodium channel have been identified in a heterogeneous subset of cardiac rhythm syndromes, including Brugada syndrome, progressive cardiac conduction defect, sick sinus node syndrome, atrial fibrillation and dilated cardiomyopathy. The aim of our study was to associate some SCN5A polymorphic variants directly with confirmed coronary stenoses in patients with non-LQTS ventricular fibrillation/flutter treated by an implantable cardioverter defibrillator. MATERIALS AND METHODS: A group of 32 unrelated individuals, aged 63 ± 12 years, was included in the study. All the patients were examined, diagnosed and treated with an implantable cardioverter defibrillator at the Department of Internal Cardiology Medicine, Faculty Hospital Brno. The control group included 87 persons of similar age without afflicted coronary circulation, which was confirmed coronagraphically. Genomic DNA was extracted from samples of peripheral blood according to the standard protocol. Two SCN5A polymorphisms-IVS9-3C/A (rs41312433) and A1673G (rs1805124, H558R)-were examined in association with coronary artery stenosis in the patients. RESULTS: In the case-control study, no significant differences in genotype distribution/allelic frequencies were observed for IVS9-3c>a and A1673G gene polymorphisms between patients with severe arrhythmias and healthy persons. The distribution of SCN5A double genotypes was not significantly different among different types of arrhythmias according to their ejection fraction in arrhythmic patients (p = 0.396). The ventricular arrhythmias with an ejection fraction below 40% were found to be 10.67 times more frequent in patients with multiple coronary stenosis with clinically valid sensitivity, specificity and power tests. In the genotype-phenotype study, we observed a significant association of both SCN5A polymorphisms with the stenosis of coronary vessels in the patients with severe arrhythmia. The double genotype of polymorphisms IVS9-3C/A together with A1673G (CCAA) as well as their simple genotypes were associated with significant multiple stenosis of coronary arteries (MVS) with high sensitivity and specificity (p = 0.05; OR = 5 (95% CI 0.99-23.34); sensitivity 0.70; specificity 0.682; power test 0.359) Moreover, when a concrete stenotic coronary artery was associated with SCN5A genotypes, the CCAA double genotype was observed to be five times more frequent in patients with significant stenosis in the right coronary artery (RCA) compared to those without affliction of this coronary artery (p = 0.05; OR = 5 (95% CI 0.99-23.34); sensitivity 0.682; specificity 0.700; power test 0.359). The CCAA genotype was also more frequent in patients without RCA affliction with MVS (p = 0.008); in patients with ACD affliction but without MVS (p = 0.008); and in patients with both ACD affliction and MVS compared to those without ACD affliction and MVS (p = 0.005). CONCLUSIONS: Our study presents a highly sensitive and specific association of two polymorphisms in SCN5A with significant coronary artery stenoses in patients with potentially fatal ventricular arrhythmias. At the same time, these polymorphisms were not associated with arrhythmias themselves. Thus, SCN5A gene polymorphic variants may form a part of germ cell gene predisposition to ischemia.