Plakophilin-2 Haploinsufficiency Causes Calcium Handling Deficits and Modulates the Cardiac Response Towards Stress.

Human variants in plakophilin-2 (PKP2) associate with most cases of familial arrhythmogenic cardiomyopathy (ACM). Recent studies show that PKP2 not only maintains intercellular coupling, but also regulates transcription of genes involved in Ca cycling and cardiac rhythm. ACM penetrance is low and it remains uncertain, which genetic and environmental modifiers are crucial for developing the cardiomyopathy.

A common co-morbidity modulates disease expression and treatment efficacy in inherited cardiac sodium channelopathy.

Aims: Management of patients with inherited cardiac ion channelopathy is hindered by variability in disease severity and sudden cardiac death (SCD) risk. Here, we investigated the modulatory role of hypertrophy on arrhythmia and SCD risk in sodium channelopathy.

Methods And Results: Follow-up data was collected from 164 individuals positive for the SCN5A-1795insD founder mutation and 247 mutation-negative relatives.

Calmodulin/CaMKII inhibition improves intercellular communication and impulse propagation in the heart and is antiarrhythmic under conditions when fibrosis is absent.

Aim: In healthy hearts, ventricular gap junctions are mainly composed by connexin43 (Cx43) and localize in the intercalated disc, enabling appropriate electrical coupling. In diseased hearts, Cx43 is heterogeneously down-regulated, whereas activity of calmodulin/calcium-calmodulin protein kinase II (CaM/CaMKII) signalling increases. It is unclear if CaM/CaMKII affects Cx43 expression/localization or impulse propagation.

Beat-to-Beat Variability in Preload Unmasks Latent Risk of Torsade de Pointes in Anesthetized Chronic Atrioventricular Block Dogs.

Background: Beat-to-beat variability in ventricular repolarization (BVR) associates with increased arrhythmic risk. Proarrhythmic remodeling in the dog with chronic AV-block (CAVB) compromises repolarization reserve and associates with increased BVR, which further increases upon dofetilide infusion and correlates with Torsade de Pointes (TdP) arrhythmias. It was hypothesized that these pro-arrhythmia-associated increases in BVR are induced by beat-to-beat variability in preload.

Changes in Cx43 and NaV1.5 expression precede the occurrence of substantial fibrosis in calcineurin-induced murine cardiac hypertrophy.

In mice, the calcium-dependent phosphatase calcineurin A (CnA) induces a transcriptional pathway leading to pathological cardiac hypertrophy. Interestingly, induction of CnA has been frequently noticed in human hypertrophic and failing hearts. Independently, the arrhythmia vulnerability of such hearts has been regularly associated with remodeling of parameters determining electrical conduction (expression level of connexin43 (Cx43) and NaV1.

Nfat and miR-25 cooperate to reactivate the transcription factor Hand2 in heart failure.

Although aberrant reactivation of embryonic gene programs is intricately linked to pathological heart disease, the transcription factors driving these gene programs remain ill-defined. Here we report that increased calcineurin/Nfat signalling and decreased miR-25 expression integrate to re-express the basic helix-loop-helix (bHLH) transcription factor dHAND (also known as Hand2) in the diseased human and mouse myocardium. In line, mutant mice overexpressing Hand2 in otherwise healthy heart muscle cells developed a phenotype of pathological hypertrophy.

Combined Na(+)/Ca(2+) exchanger and L-type calcium channel block as a potential strategy to suppress arrhythmias and maintain ventricular function.

Background: L-type calcium channel (LTCC) and Na(+)/Ca(2+) exchanger (NCX) have been implicated in repolarization-dependent arrhythmias, but also modulate calcium and contractility. Although LTCC inhibition is negative inotropic, NCX inhibition has the opposite effect. Combined block may, therefore, offer an advantage for hemodynamics and antiarrhythmic efficiency, particularly in diseased hearts.

Relevance of calmodulin/CaMKII activation for arrhythmogenesis in the AV block dog.

Background: The calcium-dependent signaling molecules calcineurin and calcium/calmodulin-dependent protein kinase II (CaMKII) both have been linked to decompensated hypertrophy and arrhythmias. CaMKII is also believed to be involved in acute modulation of ion channels.

Objective: The purpose of this study was to determine the role of calcineurin and CaMKII in a dog model of compensated hypertrophy and a long QT phenotype.

Sodium current deficit and arrhythmogenesis in a murine model of plakophilin-2 haploinsufficiency.

Aims: The shRNA-mediated loss of expression of the desmosomal protein plakophilin-2 leads to sodium current (I(Na)) dysfunction. Whether pkp2 gene haploinsufficiency leads to I(Na) deficit in vivo remains undefined. Mutations in pkp2 are detected in arrhythmogenic right ventricular cardiomyopathy (ARVC).

Reduced Cx43 expression triggers increased fibrosis due to enhanced fibroblast activity.

Background: Arrhythmogenic ventricular remodeling is hallmarked by both reduced gap junction expression and increased collagen deposition. We hypothesized that reduced connexin43 (Cx43) expression is responsible for enhanced fibrosis in the remodeled heart, resulting in an arrhythmogenic substrate. Therefore, we investigated the effect of normal or reduced Cx43 expression on the formation of fibrosis in a physiological (aging) and pathophysiological (transverse aortic constriction [TAC]) mouse model.

Reduced heterogeneous expression of Cx43 results in decreased Nav1.5 expression and reduced sodium current that accounts for arrhythmia vulnerability in conditional Cx43 knockout mice.

Background: Reduced expression of connexin43 (Cx43) and sodium channel (Nav1.5) and increased expression of collagen (fibrosis) are important determinants of impulse conduction in the heart.

Objective: To study the importance and interaction of these factors at very low Cx43 expression, inducible Cx43 knockout mice with and without inducible ventricular tachycardia (VT) were compared through electrophysiology and immunohistochemistry.

Effects of K201 on repolarization and arrhythmogenesis in anesthetized chronic atrioventricular block dogs susceptible to dofetilide-induced torsade de pointes.

The novel antiarrhythmic drug K201 (4-[3-{1-(4-benzyl)piperidinyl}propionyl]-7-methoxy-2,3,4,5-tetrahydro-1,4-benzothiazepine monohydrochloride) is currently in development for treatment of atrial fibrillation. K201 not only controls intracellular calcium release by the ryanodine receptors, but also possesses a ventricular action that might predispose to torsade de pointes arrhythmias. The anti- and proarrhythmic effects of K201 were investigated in the anesthetized canine chronic atrioventricular block model.

Ventricular remodelling is a prerequisite for the induction of dofetilide-induced torsade de pointes arrhythmias in the anaesthetized, complete atrio-ventricular-block dog.

Introduction: A number of predisposing factors have been suggested to be contributing to drug-induced torsade de pointes (TdP) arrhythmias: short-long-short (SLS) sequence, bradycardia, timing of drug administration, anaesthesia, ventricular remodelling, and altered ventricular activation due to ventricular ectopic beats (SLS) or idioventricular rhythm (IVR). Chronic atrio-ventricular (AV)-block (CAVB) dogs are susceptible to dofetilide-induced TdP.

Methods And Results: In 32 anaesthetized animals, the relevance of ventricular remodelling for TdP susceptibility was studied by dofetilide [0.

Direct detection of myocardial fibrosis by MRI.

Excessive collagen deposition is a major hallmark of cardiac disease. Fibrosis reduces cardiac function and plays a major role in cardiac arrhythmogeneity. Despite the clinical importance, there is no non-invasive technique for direct detection of myocardial fibrosis yet.

Peroxisome proliferator-activated receptor (PPAR) gene profiling uncovers insulin-like growth factor-1 as a PPARalpha target gene in cardioprotection.

Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear receptor family of ligand-activated transcription factors and consist of the three isoforms, PPARα, PPARβ/δ, and PPARγ. Considerable evidence indicates the importance of PPARs in cardiovascular lipid homeostasis and diabetes, yet the isoform-dependent cardiac target genes remain unknown. Here, we constructed murine ventricular clones allowing stable expression of siRNAs to achieve specifically knockdown for each of the PPAR isoforms.

MicroRNA-199b targets the nuclear kinase Dyrk1a in an auto-amplification loop promoting calcineurin/NFAT signalling.

MicroRNAs (miRs) are a class of single-stranded, non-coding RNAs of about 22 nucleotides in length. Increasing evidence implicates miRs in myocardial disease processes. Here we show that miR-199b is a direct calcineurin/NFAT target gene that increases in expression in mouse and human heart failure, and targets the nuclear NFAT kinase dual-specificity tyrosine-(Y)-phosphorylation regulated kinase 1a (Dyrk1a), constituting a pathogenic feed forward mechanism that affects calcineurin-responsive gene expression.

Anesthesia and arrhythmogenesis in the chronic atrioventricular block dog model.

Background: Drug-induced torsade de pointes (TdP) arrhythmias can readily be induced in anesthetized dogs with remodeled hearts [chronic complete atrioventricular block (CAVB) dogs]. Similar studies in conscious CAVB dogs reveal lower TdP incidences. Regulations forced us to reconsider our anesthetic regimen, which consist of pentobarbital followed by halothane (P + H).

MEF2 transcriptional activity maintains mitochondrial adaptation in cardiac pressure overload.

Aims: The transcription factor MEF2 is a downstream target for several hypertrophic signalling pathways in the heart, suggesting that MEF2 may act as a valuable therapeutic target in the treatment of heart failure.

Methods And Results: In this study, we investigated the potential benefits of overall MEF2 inhibition in a mouse model of chronic pressure overloading, by subjecting transgenic mice expressing a dominant negative form of MEF2 (DN-MEF2 Tg) in the heart, to transverse aortic constriction (TAC). Histological analysis revealed no major differences in cardiac remodelling between DN-MEF2 Tg and control mice after TAC.

NF-kappaB activation is required for adaptive cardiac hypertrophy.

Aims: We have previously shown that cardiac-specific inhibition of NF-kappaB attenuates angiotensin II (AngII)-induced left ventricular (LV) hypertrophy in vivo. We now tested whether NF-kappaB inhibition is able to block LV remodelling upon chronic pressure overload and chronic AngII stimulation.

Methods And Results: Cardiac-restricted NF-kappaB inhibition was achieved by expression of a stabilized IkappaBalpha mutant (IkappaBalphaDeltaN) in cells with an active alpha-myosin heavy chain (alphaMHC) promoter employing the Cre/lox technique.

Combined reduction of intercellular coupling and membrane excitability differentially affects transverse and longitudinal cardiac conduction.

Aims: Reduced excitability and gap junction expression are commonly found in electrically remodelled diseased hearts, but their contribution to slow conduction and arrhythmias is unclear. In this study, we have investigated the effect of isolated and combined reductions in membrane excitability and intercellular coupling on impulse propagation and arrhythmogeneity in genetically modified mice.

Methods And Results: Cx43 and Scn5a(1798insD/+) heterozygous (HZ) mice were crossbred to create a mixed offspring: wild-type (WT, n = 15), Cx43 HZ (n = 14), Scn5a(1798insD/+) (Scn5a) HZ (n = 17), and Cx43/Scn5a(1798insD/+) (Cx43/Scn5a) HZ (n = 15) mice.

Beta-Catenin downregulation attenuates ischemic cardiac remodeling through enhanced resident precursor cell differentiation.

We analyzed the effect of conditional, alphaMHC-dependent genetic beta-catenin depletion and stabilization on cardiac remodeling following experimental infarct. beta-Catenin depletion significantly improved 4-week survival and left ventricular (LV) function (fractional shortening: CT(Deltaex3-6): 24 +/- 1.9%; beta-cat(Deltaex3-6): 30.

In calcineurin-induced cardiac hypertrophy expression of Nav1.5, Cx40 and Cx43 is reduced by different mechanisms.

Alterations in expression levels of Na(v)1.5, Cx43 and Cx40 have been frequently reported in cardiac disease and are associated with the development of arrhythmias, but little is known about the underlying molecular mechanisms. In this study we investigated electrical conduction and expression of Na(v)1.

NFATc2 is a necessary mediator of calcineurin-dependent cardiac hypertrophy and heart failure.

One major intracellular signaling pathway involved in heart failure employs the phosphatase calcineurin and its downstream transcriptional effector nuclear factor of activated T-cells (NFAT). In vivo evidence for the involvement of NFAT factors in heart failure development is still ill defined. Here we reveal that nfatc2 transcripts outnumber those from other nfat genes in the unstimulated heart by severalfold.

Loss of muscle-specific RING-finger 3 predisposes the heart to cardiac rupture after myocardial infarction.

RING-finger proteins commonly function as ubiquitin ligases that mediate protein degradation by the ubiquitin-proteasome pathway. Muscle-specific RING-finger (MuRF) proteins are striated muscle-restricted components of the sarcomere that are thought to possess ubiquitin ligase activity. We show that mice lacking MuRF3 display normal cardiac function but are prone to cardiac rupture after acute myocardial infarction.

EUK-8, a superoxide dismutase and catalase mimetic, reduces cardiac oxidative stress and ameliorates pressure overload-induced heart failure in the harlequin mouse mutant.

Objectives: The purpose of this study was to identify apoptosis-inducing factor (AIF) as a cardiac mitochondrial antioxidant and assess the efficacy of EUK-8, a salen-manganese catalytic free radical scavenger, to protect the AIF-deficient myocardium against pressure overload.

Background: Oxidative stress has been postulated to provoke cell death and pathologic remodeling in heart failure. We recently characterized the apoptosis-inducing factor-deficient harlequin (Hq) mouse mutant to display excessive pressure overload-induced oxidative stress, cell death, accelerated progression to heart failure, and a reduced capacity of subsarcolemmal mitochondria to scavenge free radicals, suggesting a role for AIF as a novel mitochondrial antioxidant.