A gain-of-function HCN4 mutant in the HCN domain is responsible for inappropriate sinus tachycardia in a Spanish family.

In a family with inappropriate sinus tachycardia (IST), we identified a mutation (p.V240M) of the hyperpolarization-activated cyclic nucleotide-gated type 4 (HCN4) channel, which contributes to the pacemaker current (If) in human sinoatrial node cells. Here, we clinically study fifteen family members and functionally analyze the p.V240M variant. Macroscopic (IHCN4) and single-channel currents were recorded using patch-clamp in cells expressing human native (WT) and/or p.V240M HCN4 channels. All p.V240M mutation carriers exhibited IST that was accompanied by cardiomyopathy in adults. IHCN4 generated by p.V240M channels either alone or in combination with WT was significantly greater than that generated by WT channels alone. The variant, which lies in the N-terminal HCN domain, increased the single-channel conductance and opening frequency and probability of HCN4 channels. Conversely, it did not modify the channel sensitivity for cAMP and ivabradine or the level of expression at the membrane. Treatment with ivabradine based on functional data reversed the IST and the cardiomyopathy of the carriers. In computer simulations, the p.V240M gain-of-function variant increases If and beating rate and thus explains the IST of the carriers. The results demonstrate the importance of the unique HCN domain in HCN4, which stabilizes the channels in the closed state.

Drug-induced atrial fibrillation. A narrative review of a forgotten adverse effect.

Atrial fibrillation (AF) is the most common cardiac arrhythmia and is associated with an increased morbidity and mortality. There is clinical evidence that an increasing number of cardiovascular and non-cardiovascular drugs, mainly anticancer drugs, can induce AF either in patients with or without pre-existing cardiac disorders, but drug-induced AF (DIAF) has not received the attention that it might deserve. In many cases DIAF is asymptomatic and paroxysmal and patients recover sinus rhythm spontaneously, but sometimes, DIAF persists, and it is necessary to perform a cardioversion. Furthermore, DIAF is not mentioned in clinical guidelines on the treatment of AF. The risk of DIAF increases in elderly and in patients treated with polypharmacy and with risk factors and comorbidities that commonly coexist with AF. This is the case of cancer patients. Under these circumstances ascribing causality of DIAF to a given drug often represents a clinical challenge. We review the incidence, the pathophysiological mechanisms, risk factors, clinical relevance, and treatment of DIAF. Because of the limited information presently available, further research is needed to obtain a deeper insight into DIAF. Meanwhile, it is important that clinicians are aware of the problem that DIAF represents, recognize which drugs may cause DIAF, and consider the possibility that a drug may be responsible for a new-onset AF episode.

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.

Zfhx3 Transcription Factor Represses the Expression of SCN5A Gene and Decreases Sodium Current Density (INa).

The ZFHX3 and SCN5A genes encode the zinc finger homeobox 3 (Zfhx3) transcription factor (TF) and the human cardiac Na+ channel (Nav1.5), respectively. The effects of Zfhx3 on the expression of the Nav1.5 channel, and in cardiac excitability, are currently unknown. Additionally, we identified three Zfhx3 variants in probands diagnosed with familial atrial fibrillation (p.M1260T) and Brugada Syndrome (p.V949I and p.Q2564R). Here, we analyzed the effects of native (WT) and mutated Zfhx3 on Na+ current (INa) recorded in HL-1 cardiomyocytes. ZFHX3 mRNA can be detected in human atrial and ventricular samples. In HL-1 cardiomyocytes, transfection of Zfhx3 strongly reduced peak INa density, while the silencing of endogenous expression augmented it (from -65.9 ± 8.9 to -104.6 ± 10.8 pA/pF; n ≥ 8, p < 0.05). Zfhx3 significantly reduced the transcriptional activity of human SCN5A, PITX2, TBX5, and NKX25 minimal promoters. Consequently, the mRNA and/or protein expression levels of Nav1.5 and Tbx5 were diminished (n ≥ 6, p < 0.05). Zfhx3 also increased the expression of Nedd4-2 ubiquitin-protein ligase, enhancing Nav1.5 proteasomal degradation. p.V949I, p.M1260T, and p.Q2564R Zfhx3 produced similar effects on INa density and time- and voltage-dependent properties in WT. WT Zfhx3 inhibits INa as a result of a direct repressor effect on the SCN5A promoter, the modulation of Tbx5 increasing on the INa, and the increased expression of Nedd4-2. We propose that this TF participates in the control of cardiac excitability in human adult cardiac tissue.

Cardiac Kir2.1 and NaV1.5 Channels Traffic Together to the Sarcolemma to Control Excitability.

RATIONALE: In cardiomyocytes, NaV1.5 and Kir2.1 channels interact dynamically as part of membrane bound macromolecular complexes. OBJECTIVE: The objective of this study was to test whether NaV1.5 and Kir2.1 preassemble during early forward trafficking and travel together to common membrane microdomains. METHODS AND RESULTS: In patch-clamp experiments, coexpression of trafficking-deficient mutants Kir2.1Δ314-315 or Kir2.1R44A/R46A with wild-type (WT) NaV1.5WT in heterologous cells reduced inward sodium current compared with NaV1.5WT alone or coexpressed with Kir2.1WT. In cell surface biotinylation experiments, expression of Kir2.1Δ314-315 reduced NaV1.5 channel surface expression. Glycosylation analysis suggested that NaV1.5WT and Kir2.1WT channels associate early in their biosynthetic pathway, and fluorescence recovery after photobleaching experiments demonstrated that coexpression with Kir2.1 increased cytoplasmic mobility of NaV1.5WT, and vice versa, whereas coexpression with Kir2.1Δ314-315 reduced mobility of both channels. Viral gene transfer of Kir2.1Δ314-315 in adult rat ventricular myocytes and human induced pluripotent stem cell-derived cardiomyocytes reduced inward rectifier potassium current and inward sodium current, maximum diastolic potential and action potential depolarization rate, and increased action potential duration. On immunostaining, the AP1 (adaptor protein complex 1) colocalized with NaV1.5WT and Kir2.1WT within areas corresponding to t-tubules and intercalated discs. Like Kir2.1WT, NaV1.5WT coimmunoprecipitated with AP1. Site-directed mutagenesis revealed that NaV1.5WT channels interact with AP1 through the NaV1.5Y1810 residue, suggesting that, like for Kir2.1WT, AP1 can mark NaV1.5 channels for incorporation into clathrin-coated vesicles at the trans-Golgi. Silencing the AP1 ϒ-adaptin subunit in human induced pluripotent stem cell-derived cardiomyocytes reduced inward rectifier potassium current, inward sodium current, and maximum diastolic potential and impaired rate-dependent action potential duration adaptation. CONCLUSIONS: The NaV1.5-Kir2.1 macromolecular complex pre-assembles early in the forward trafficking pathway. Therefore, disruption of Kir2.1 trafficking in cardiomyocytes affects trafficking of NaV1.5, which may have important implications in the mechanisms of arrhythmias in inheritable cardiac diseases.

Tbx20 controls the expression of the KCNH2 gene and of hERG channels.

Long QT syndrome (LQTS) exhibits great phenotype variability among family members carrying the same mutation, which can be partially attributed to genetic factors. We functionally analyzed the KCNH2 (encoding for Kv11.1 or hERG channels) and TBX20 (encoding for the transcription factor Tbx20) variants found by next-generation sequencing in two siblings with LQTS in a Spanish family of African ancestry. Affected relatives harbor a heterozygous mutation in KCNH2 that encodes for p.T152HfsX180 Kv11.1 (hERG). This peptide, by itself, failed to generate any current when transfected into Chinese hamster ovary (CHO) cells but, surprisingly, exerted "chaperone-like" effects over native hERG channels in both CHO cells and mouse atrial-derived HL-1 cells. Therefore, heterozygous transfection of native (WT) and p.T152HfsX180 hERG channels generated a current that was indistinguishable from that generated by WT channels alone. Some affected relatives also harbor the p.R311C mutation in Tbx20. In human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), Tbx20 enhanced human KCNH2 gene expression and hERG currents (IhERG) and shortened action-potential duration (APD). However, Tbx20 did not modify the expression or activity of any other channel involved in ventricular repolarization. Conversely, p.R311C Tbx20 did not increase KCNH2 expression in hiPSC-CMs, which led to decreased IhERG and increased APD. Our results suggest that Tbx20 controls the expression of hERG channels responsible for the rapid component of the delayed rectifier current. On the contrary, p.R311C Tbx20 specifically disables the Tbx20 protranscriptional activity over KCNH2 Therefore, TBX20 can be considered a KCNH2-modifying gene.

Cardiac electrical defects in progeroid mice and Hutchinson-Gilford progeria syndrome patients with nuclear lamina alterations.

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disease caused by defective prelamin A processing, leading to nuclear lamina alterations, severe cardiovascular pathology, and premature death. Prelamin A alterations also occur in physiological aging. It remains unknown how defective prelamin A processing affects the cardiac rhythm. We show age-dependent cardiac repolarization abnormalities in HGPS patients that are also present in the Zmpste24-/- mouse model of HGPS. Challenge of Zmpste24-/- mice with the ß-adrenergic agonist isoproterenol did not trigger ventricular arrhythmia but caused bradycardia-related premature ventricular complexes and slow-rate polymorphic ventricular rhythms during recovery. Patch-clamping in Zmpste24-/- cardiomyocytes revealed prolonged calcium-transient duration and reduced sarcoplasmic reticulum calcium loading and release, consistent with the absence of isoproterenol-induced ventricular arrhythmia. Zmpste24-/- progeroid mice also developed severe fibrosis-unrelated bradycardia and PQ interval and QRS complex prolongation. These conduction defects were accompanied by overt mislocalization of the gap junction protein connexin43 (Cx43). Remarkably, Cx43 mislocalization was also evident in autopsied left ventricle tissue from HGPS patients, suggesting intercellular connectivity alterations at late stages of the disease. The similarities between HGPS patients and progeroid mice reported here strongly suggest that defective cardiac repolarization and cardiomyocyte connectivity are important abnormalities in the HGPS pathogenesis that increase the risk of arrhythmia and premature death.

New Therapeutic Approaches for the Treatment of Hyperkalemia in Patients Treated with Renin-Angiotensin-Aldosterone System Inhibitors.

Hyperkalemia (serum potassium > 5.5 mEq/L) is a common clinical problem in patients with chronic kidney disease, hypertension, diabetes, and heart failure. It can result from increased K+ intake, impaired distribution between intracellular and extracellular spaces, and most frequently, decreased renal excretion. Patients at the highest risk of hyperkalemia are treated with renin-angiotensin-aldosterone system inhibitors (RAASIs) as they improve cardiovascular and renal outcomes and are strongly recommended in clinical guidelines. However, RAASIs cause or increase the risk of hyperkalemia, a key limitation to fully titrate RAASIs in patients who are most likely to benefit from treatment. Until recently, drugs for the treatment of hyperkalemia presented limited efficacy and/or safety concerns and there was an unmet need of new drugs to control hyperkalemia while maintaining RAASI therapy. We provide an overview of the mechanisms involved in K+ homeostasis and the epidemiology and management of hyperkalemia as a complication in cardiovascular patients and, finally, analyze the efficacy and safety of two new polymer-based, non-systemic agents, patiromer calcium and sodium zirconium cyclosilicate (ZS-9), designed to increase fecal K+ loss and to normalize elevated serum K+ levels and chronically maintain K+ homeostasis in hyperkalemic patients treated with RAASIs.

miR-208b upregulation interferes with calcium handling in HL-1 atrial myocytes: Implications in human chronic atrial fibrillation.

MicroRNAs (miR) have considerable potential as therapeutic tools in cardiac diseases. Alterations in atrial miR are involved in the development of atrial fibrillation (AF), but the molecular mechanism underlying their contribution to atrial remodeling in chronic atrial fibrillation (CAF) is only partially understood. Here we used miR array to analyze the miR profile of atrial biopsies from sinus rhythm (SR) and CAF patients. qRT-PCR identified a distinctive CAF-miR signature and described conserved miR-208b upregulation in human and ovine AF atrial tissue. We used bioinformatics analysis to predict genes and signaling pathways as putative miR-208b targets, which highlighted genes from the cardiac muscle gene program and from canonical WNT, gap-junction and Ca2+ signaling networks. Results from analysis of miR-208b-overexpressing HL-1 atrial myocytes and from myocytes isolated from CAF patients showed that aberrant miR-208b levels reduced the expression and function of L-type Ca2+ channel subunits (CACNA1C and CACNB2) as well as the sarcoplasmic reticulum-Ca2+ pump SERCA2. These findings clearly pointed to CAF-specific upregulated miR-208b as an important mediator in Ca2+ handling impairment during atrial remodeling.

Coexistent HCN4 and GATA5 Rare Variants and Atrial Fibrillation in a Large Spanish Family.

BACKGROUND: Familial association of atrial fibrillation (AF) can involve single gene variants related to known arrhythmogenic mechanisms; however, genome-wide association studies often disclose complex genetic variants in familial and nonfamilial AF, making it difficult to relate to known pathogenetic mechanisms. METHODS: The finding of 4 siblings with AF led to studying 47 members of a family. Long-term Holter monitoring (average 298 hours) ruled out silent AF. Whole-exome sequencing was performed, and variants shared by the index cases were filtered and prioritised according to current recommendations. HCN4 currents (IHCN4) were recorded in Chinese hamster ovary cells expressing human p.P1163H or native HCN4 channels with the use of the patch-clamp technique, and topologically associating domain analyses of GATA5 variant were performed. RESULTS: The clinical study diagnosed 2 more AF cases. Five family members carried the heterozygous p.P1163H HCN4 variant, 14 carried the intronic 20,61040536,G,A GATA5 rare variant, and 9 carried both variants (HCN4+GATA5). Five of the 6 AF cases (onset age ranging from 33 to 70 years) carried both variants and 1 carried the GATA5 variant alone. Multivariate analysis showed that the presence of HCN4+GATA5 variants significantly increased AF risk (odds ratio 32.7, 95% confidence interval 1.8-591.4) independently from age, hypertension, and overweight. Functional testing showed that IHCN4 generated by heterozygous p.P1163H were normal. Topologically associating domain analysis suggested that GATA5 could affect the expression of many genes, including those encoding microRNA-1. CONCLUSION: The coincidence of 2 rare gene variants was independently associated with AF, but functional studies do not allow the postulation of the arrhythmogenic mechanisms involved.

Flecainide increases Kir2.1 currents by interacting with cysteine 311, decreasing the polyamine-induced rectification.

Both increase and decrease of cardiac inward rectifier current (I(K1)) are associated with severe cardiac arrhythmias. Flecainide, a widely used antiarrhythmic drug, exhibits ventricular proarrhythmic effects while effectively controlling ventricular arrhythmias associated with mutations in the gene encoding Kir2.1 channels that decrease I(K1) (Andersen syndrome). Here we characterize the electrophysiological and molecular basis of the flecainide-induced increase of the current generated by Kir2.1 channels (I(Kir2.1)) and I(K1) recorded in ventricular myocytes. Flecainide increases outward I(Kir2.1) generated by homotetrameric Kir2.1 channels by decreasing their affinity for intracellular polyamines, which reduces the inward rectification of the current. Flecainide interacts with the HI loop of the cytoplasmic domain of the channel, Cys311 being critical for the effect. This explains why flecainide does not increase I(Kir2.2) and I(Kir2.3), because Kir2.2 and Kir2.3 channels do not exhibit a Cys residue at the equivalent position. We further show that incubation with flecainide increases expression of functional Kir2.1 channels in the membrane, an effect also determined by Cys311. Indeed, flecainide pharmacologically rescues R67W, but not R218W, channel mutations found in Andersen syndrome patients. Moreover, our findings provide noteworthy clues about the structural determinants of the C terminus cytoplasmic domain of Kir2.1 channels involved in the control of gating and rectification.

A Cantú syndrome mutation produces dual effects on KATP channels by disrupting ankyrin B regulation.

ATP-sensitive potassium (KATP) channels composed of Kir6.x and sulfonylurea receptor (SURs) subunits couple cellular metabolism to electrical activity. Cantú syndrome (CS) is a rare disease caused by mutations in the genes encoding Kir6.1 (KCNJ8) and SUR2A (ABCC9) that produce KATP channel hyperactivity due to a reduced channel block by physiological ATP concentrations. We functionally characterized the p.S1054Y SUR2A mutation identified in two CS carriers, who exhibited a mild phenotype although the mutation was predicted as highly pathogenic. We recorded macroscopic and single-channel currents in CHO and HEK-293 cells and measured the membrane expression of the channel subunits by biotinylation assays in HEK-293 cells. The mutation increased basal whole-cell current density and at the single-channel level, it augmented opening frequency, slope conductance, and open probability (Po), and promoted the appearance of multiple conductance levels. p.S1054Y also reduced Kir6.2 and SUR2A expression specifically at the membrane. Overexpression of ankyrin B (AnkB) prevented these gain- and loss-of-function effects, as well as the p.S1054Y-induced reduction of ATP inhibition of currents measured in inside-out macropatches. Yeast two-hybrid assays suggested that SUR2A WT and AnkB interact, while p.S1054Y interaction with AnkB is decreased. The p.E322K Kir6.2 mutation, which prevents AnkB binding to Kir6.2, produced similar biophysical alterations than p.S1054Y. Our results are the first demonstration of a CS mutation whose functional consequences involve the disruption of AnkB effects on KATP channels providing a novel mechanism by which CS mutations can reduce ATP block. Furthermore, they may help explain the mild phenotype associated with this mutation.

Cancer Chemotherapy-Induced Sinus Bradycardia: A Narrative Review of a Forgotten Adverse Effect of Cardiotoxicity.

Cardiotoxicity is a common adverse effect of anticancer drugs (ACDs), including the so-called targeted drugs, and increases morbidity and mortality in patients with cancer. Attention has focused mainly on ACD-induced heart failure, myocardial ischemia, hypertension, thromboembolism, QT prolongation, and tachyarrhythmias. Yet, although an increasing number of ACDs can produce sinus bradycardia (SB), this proarrhythmic effect remains an underappreciated complication, probably because of its low incidence and severity since most patients are asymptomatic. However, SB merits our interest because its incidence increases with the aging of the population and cancer is an age-related disease and because SB represents a risk factor for QT prolongation. Indeed, several ACDs that produce SB also prolong the QT interval. We reviewed published reports on ACD-induced SB from January 1971 to November 2020 using the PubMed and EMBASE databases. Published reports from clinical trials, case reports, and recent reviews were considered. This review describes the associations between ACDs and SB, their clinical relevance, risk factors, and possible mechanisms of onset and treatment.

Sex-related differences in the pharmacological treatment of heart failure.

Heart failure (HF) represents a leading cause of morbidity and mortality. However, HF trials highlighted many differences between men and women with HF. Thus, women represent approximately a quarter of people with HF with reduced ejection fraction (HFrEF), while they account for over half of those with HF with preserved EF (HFpEF). There are also sex-related differences (SRDs) in the pharmacokinetics, pharmacodynamics and safety profile of some guideline-recommended drugs for the treatment of HF. As compared with men, women with HFrEF are less often treated with guideline-recommended HF drugs, experience more frequent and severe adverse reactions when these drugs are prescribed at the same doses in both sexes, and recent evidence suggests that women might need lower doses than men, bringing into question which are the optimal doses of HF drugs in women and men separately. However, information on SRDs in drug efficacy and safety in patients with HFrEF is very limited due to the underrepresentation of women and the lack of sex-specific evaluations of drug efficacy and safety in HF clinical trials. As a consequence, current clinical guidelines do not provide sex-specific recommendations, even when significant differences exist, at least, in drug safety. The aim of this article is to review the SRDs in the pharmacokinetics, efficacy and safety of guideline-recommended HF drugs and to identify emerging areas of research to improve our understanding of the SRDs, because a better understanding of these differences is the first step to achieve a personalized treatment of HF in women and men.

Empagliflozin and Dapagliflozin Increase Na+ and Inward Rectifier K+ Current Densities in Human Cardiomyocytes Derived from Induced Pluripotent Stem Cells (hiPSC-CMs).

Dapagliflozin (dapa) and empagliflozin (empa) are sodium-glucose cotransporter-2 inhibitors (SGLT2is) that reduce morbidity and mortality in heart failure (HF) patients. Sodium and inward rectifier K+ currents (INa and IK1), carried by Nav1.5 and Kir2.1 channels, respectively, are responsible for cardiac excitability, conduction velocity, and refractoriness. In HF patients, Nav1.5 and Kir2.1 expression are reduced, enhancing risk of arrhythmia. Incubation with dapa or empa (24-h,1 µM) significantly increased INa and IK1 densities recorded in human-induced pluripotent stem cell-cardiomyocytes (hiPSC-CMs) using patch-clamp techniques. Dapa and empa, respectively, shifted to more hyperpolarized potentials the INa activation and inactivation curves. Identical effects were observed in Chinese hamster ovary (CHO) cells that were incubated with dapa or empa and transiently expressed human Nav1.5 channels. Conversely, empa but not dapa significantly increased human Kir2.1 currents in CHO cells. Dapa and empa effects on INa and IK1 were also apparent in Ca-calmodulin kinase II-silenced CHO cells. Cariporide, a Na+/H+ exchanger type 1 (NHE1) inhibitor, did not increase INa or IK1 in hiPSC-CMs. Dapa and empa at therapeutic concentrations increased INa and IK1 in healthy human cardiomyocytes. These SGLT2is could represent a new class of drugs with a novel and long-pursued antiarrhythmic mechanism of action.

Tbx5 variants disrupt Nav1.5 function differently in patients diagnosed with Brugada or Long QT Syndrome.

AIMS: The transcription factor Tbx5 controls cardiogenesis and drives Scn5a expression in mice. We have identified two variants in TBX5 encoding p. D111Y and p. F206L Tbx5, respectively, in two unrelated patients with structurally normal hearts diagnosed with long QT (LQTS) and Brugada (BrS) syndrome. Here, we characterized the consequences of each variant to unravel the underlying disease mechanisms. METHODS AND RESULTS: We combined clinical analysis with in vivo and in vitro electrophysiological and molecular techniques in human-induced pluripotent stem-cell-derived cardiomyocytes (hiPSC-CMs), HL-1 cells, and cardiomyocytes from mice trans-expressing human wild-type (WT) or mutant proteins. Tbx5 increased transcription of SCN5A encoding cardiac Nav1.5 channels, while repressing CAMK2D and SPTBN4 genes encoding Ca/calmodulin kinase IIδ (CaMKIIδ) and ßIV-spectrin, respectively. These effects significantly increased Na current (INa) in hiPSC-CMs and in cardiomyocytes from mice trans-expressing Tbx5. Consequently, action potential (AP) amplitudes increased and QRS interval narrowed in the mouse electrocardiogram. p. F206L Tbx5 bound to the SCN5A promoter failed to transactivate it, thus precluding the pro-transcriptional effect of WT Tbx5. Therefore, p. F206L markedly decreased INa in hiPSC-CM, HL-1 cells and mouse cardiomyocytes. The INa decrease in p. F206L trans-expressing mice translated into QRS widening and increased flecainide sensitivity. p. D111Y Tbx5 increased SCN5A expression but failed to repress CAMK2D and SPTBN4. The increased CaMKIIδ and ßIV-spectrin significantly augmented the late component of INa (INaL) which, in turn, significantly prolonged AP duration in both hiPSC-CMs and mouse cardiomyocytes. Ranolazine, a selective INaL inhibitor, eliminated the QT and QTc intervals prolongation seen in p. D111Y trans-expressing mice. CONCLUSIONS: In addition to peak INa, Tbx5 critically regulates INaL and the duration of repolarization in human cardiomyocytes. Our original results suggest that TBX5 variants associate with and modulate the intensity of the electrical phenotype in LQTS and BrS patients.

Facing the challenge of polypharmacy when prescribing for older people with cardiovascular disease. A review by the European Society of Cardiology Working Group on Cardiovascular Pharmacotherapy.

Population ageing has resulted in an increasing number of older people living with chronic diseases (multimorbidity) requiring five or more medications daily (polypharmacy). Ageing produces important changes in the cardiovascular system and represents the most potent single cardiovascular risk factor. Cardiovascular diseases (CVDs) constitute the greatest burden for older people, their caregivers, and healthcare systems. Cardiovascular pharmacotherapy in older people is complex because age-related changes in body composition, organ function, homeostatic mechanisms, and comorbidities modify the pharmacokinetic and pharmacodynamic properties of many commonly used cardiovascular and non-cardiovascular drugs. Additionally, polypharmacy increases the risk of adverse drug reactions and drug interactions, which in turn can lead to increased morbi-mortality and healthcare costs. Unfortunately, evidence of drug efficacy and safety in older people with multimorbidity and polypharmacy is limited because these individuals are frequently underrepresented/excluded from clinical trials. Moreover, clinical guidelines are largely written with a single-disease focus and only occasionally address the issue of coordination of care, when and how to discontinue treatments, if required, or how to prioritize recommendations for patients with multimorbidity and polypharmacy. This review analyses the main challenges confronting healthcare professionals when prescribing in older people with CVD, multimorbidity, and polypharmacy. Our goal is to provide information that can contribute to improving drug prescribing, efficacy, and safety, as well as drug adherence and clinical outcomes.

Nitric oxide increases cardiac IK1 by nitrosylation of cysteine 76 of Kir2.1 channels.

RATIONALE: The cardiac inwardly rectifying K(+) current (I(K1)) plays a critical role in modulating excitability by setting the resting membrane potential and shaping phase 3 of the cardiac action potential. OBJECTIVE: This study aims to analyze the effects of nitric oxide (NO) on human atrial I(K1) and on Kir2.1 channels, the major isoform of inwardly rectifying channels present in the human heart. METHODS AND RESULTS: Currents were recorded in enzymatically isolated myocytes and in transiently transfected CHO cells, respectively. NO at myocardial physiological concentrations (25 to 500 nmol/L) increased inward and outward I(K1) and I(Kir2.1). These effects were accompanied by hyperpolarization of the resting membrane potential and a shortening of the duration of phase 3 of the human atrial action potential. The I(Kir2.1) increase was attributable to an increase in the open probability of the channel. Site-directed mutagenesis analysis demonstrated that NO effects were mediated by the selective S-nitrosylation of Kir2.1 Cys76 residue. Single ion monitoring experiments performed by liquid chromatography/tandem mass spectrometry suggested that the primary sequence that surrounds Cys76 determines its selective S-nitrosylation. Chronic atrial fibrillation, which produces a decrease in NO bioavailability, decreased the S-nitrosylation of Kir2.1 channels in human atrial samples as demonstrated by a biotin-switch assay, followed by Western blot. CONCLUSIONS: The results demonstrated that, under physiological conditions, NO regulates human cardiac I(K1) through a redox-related process.

Chronic (-)-epicatechin improves vascular oxidative and inflammatory status but not hypertension in chronic nitric oxide-deficient rats.

The present study analysed the effects of the flavanol (-)-epicatechin in rats after chronic inhibition of NO synthesis with NG-nitro-L-arginine methyl ester (L-NAME), at doses equivalent to those achieved in the studies involving human subjects. Wistar rats were randomly divided into four groups: (1) control-vehicle, (2) L-NAME, (3) L-NAME-epicatechin 2 (L-NAME-Epi 2) and (4) L-NAME-epicatechin 10 (L-NAME-Epi 10). Rats were daily given by oral administration for 4 weeks: vehicle, (-)-epicatechin 2 or 10 mg/kg. Animals in the L-NAME groups daily received L-NAME 75 mg/100 ml in drinking-water. The evolution in systolic blood pressure and heart rate, and morphological and plasma variables, proteinuria, vascular superoxide, reactivity and protein expression at the end of the experiment were analysed. Chronic (-)-epicatechin treatment did not modify the development of hypertension and only weakly affected the endothelial dysfunction induced by L-NAME but prevented the cardiac hypertrophy, the renal parenchyma and vascular lesions and proteinuria, and blunted the prostanoid-mediated enhanced endothelium-dependent vasoconstrictor responses and the cyclo-oxygenase-2 and endothelial NO synthase (eNOS) up-regulation. Furthermore, (-)-epicatechin also increased Akt and eNOS phosphorylation and prevented the L-NAME-induced increase in systemic (plasma malonyldialdehyde and urinary 8-iso-PGF2α) and vascular (dihydroethidium staining, NADPH oxidase activity and p22phox up-regulation) oxidative stress, proinflammatory status (intercellular adhesion molecule-1, IL-1ß and TNFα up-regulation) and extracellular-signal-regulated kinase 1/2 phosphorylation. The present study shows for the first time that chronic oral administration of (-)-epicatechin does not improve hypertension but reduced pro-atherogenic pathways such as oxidative stress and proinflammatory status of the vascular wall induced by blockade of NO production.