Antiarrhythmische Effekte der Ryanodinrezeptorstabilisierung durch Dantrolen im humanen Myokard

Kardiale Arrhythmien bestimmen maßgeblich die hohe Morbidität und Mortalität kardiovaskulärer Erkrankungen. Der kardiale Ryanodinrezeptors 2 (RyR2) ist pathophysiologisch für die zelluläre Arrhythmogenese in der Herzinsuffizienz und beim Vorhofflimmern bedeutsam. Eine Destabilisierung des RyR2 kann zu zellulären Ca2+-Freisetzungen führen, die Arrhythmien triggern können. Die vorliegende Arbeit hat mögliche antiarrhythmische Effekte der RyR2-Stabilisierung durch Dantrolen in isolierten humanen ventrikulären Herzzellen von Patienten mit Herzinsuffizienz, sowie in atrialen Zellen von Patienten mit Vorhofflimmern oder Sinusrhythmus untersucht. Mittels Konfokalmikroskopie unter Verwendung des Ca2+-Farbstoffes Fluo-3 konnte nachgewiesen werden, dass Dantrolen (10 µmol/l) arrhythmogene diastolische Ca2+-Freisetzungen in ventrikulären Kardiomyozyten von Patienten mit Herzinsuffizienz und in atrialen Kardiomyozyten von Patienten mit Vorhofflimmern vermindert. Unter Verwendung der Patch-Clamp-Technik wurde konsekutiv die Inzidenz von zellulären Nachdepolarisationen, die durch die elektrogene Ca2+-Depletion entstehen können, untersucht. Diese zellulären arrhythmogenen Trigger, welche typischerweise vermehrt bei Herzinsuffizienz und bei Vorhofflimmern auftreten, waren in Dantrolen-behandelten Kardiomyozyten von Patienten mit Herzinsuffizienz oder Vorhofflimmern signifikant supprimiert. Mittels Konfokalmikroskopie wurden in atrialen Kardiomyozyten von Patienten mit Sinusrhythmus, bei denen keine RyR2-Destabilisierung vorbeschrieben ist, basal weniger diastolische Ca2+-Freisetzungen als in Kardiomyozyten von Patienten mit Vorhofflimmern beobachtet. Im Gegensatz zum Vorhofflimmern zeigten sich jedoch keine Effekte von Dantrolen auf diastolische Ca2+-Freisetzungen in atrialen Kardiomyozyten von Patienten mit Sinusrhythmus. Hierdurch wird ein selektiver antiarrhythmischer Wirkmechanismus von Dantrolen, der sich auf destabilisierte, „kranke“ RyR2 beschränkt, suggeriert. Abschließend zeigte Dantrolen keine Effekte auf die Aktionspotentialdauer oder auf die Kontraktilität von ventrikulären Myokardtrabekeln aus humanen Herzen. Dies stellt einen Vorteil der Substanz gegenüber zahlreich negativ inotrop wirkenden Antiarrhythmika dar. Diese Arbeit konnte somit erstmals antiarrhythmische Eigenschaften der RyR2-Stabilisierung mittels Dantrolen im menschlichen Myokard demonstrieren und diese in den Kontext kardialer Pathologien einordnen. Zugleich konnten Einflüsse von Dantrolen auf das kardiale Aktionspotential und die Kontraktilität des Myokards ausgeschlossen werden. Mit dem Nachweis der antiarrhythmischen Effekte im humanen Myokard liefert diese Arbeit eine translationale Grundlage für die weitere (klinische) Erforschung von Dantrolen zur Behandlung von kardialen Arrhythmien

Potential Mechanisms of SGLT2 Inhibitors for the Treatment of Heart Failure With Preserved Ejection Fraction

Heart failure with preserved ejection fraction (HFpEF) is an unsolved and growing concern in cardiovascular medicine. While no treatment options that improve prognosis in HFpEF patients has been established so far, SGLT2 inhibitors (SGLT2i) are currently being investigated for the treatment of HFpEF patients. SGLT2i have already been shown to mitigate comorbidities associated with HFpEF such as type 2 diabetes and chronic renal disease, however, more recently there has been evidence that they may also directly improve diastolic function. In this article, we discuss some potential beneficial mechanisms of SGLT2i in the pathophysiology of HFpEF with focus on contractile function

SGLT2 Inhibitors and Their Mode of Action in Heart Failure—Has the Mystery Been Unravelled?

Purpose of review SGLT2 inhibitors (SGLT2i) are new drugs for patients with heart failure (HF) irrespective of diabetes. However, the mechanisms of SGLT2i in HF remain elusive. This article discusses the current clinical evidence for using SGLT2i in different types of heart failure and provides an overview about the possible underlying mechanisms. Recent findings Clinical and basic data strongly support and extend the use of SGLT2i in HF. Improvement of conventional secondary risk factors is unlikely to explain the prognostic benefits of these drugs in HF. However, different multidirectional mechanisms of SGLT2i could improve HF status including volume regulation, cardiorenal mechanisms, metabolic effects, improved cardiac remodelling, direct effects on cardiac contractility and ion-homeostasis, reduction of inflammation and oxidative stress as well as an impact on autophagy and adipokines. Summary Further translational studies are needed to determine the mechanisms of SGLT2i in HF. However, basic and clinical evidence encourage the use of SGLT2i in HFrEF and possibly HFpEF

Empagliflozin inhibits Na + /H + exchanger activity in human atrial cardiomyocytes

Aims Recent clinical trials have proven gliflozins to be cardioprotective in diabetic and non-diabetic patients. However, the underlying mechanisms are incompletely understood. A potential inhibition of cardiac Na+/H(+)exchanger 1 (NHE1) has been suggested in animal models. We investigated the effect of empagliflozin on NHE1 activity in human atrial cardiomyocytes. Methods and results Expression of NHE1 was assessed in human atrial and ventricular tissue via western blotting. NHE activity was measured as the maximal slope of pH recovery after NH(4)(+)pulse in isolated carboxy-seminaphtarhodafluor 1 (SNARF1)-acetoxymethylester-loaded murine ventricular and human atrial cardiomyocytes. NHE1 is abundantly expressed in human atrial and ventricular tissue. Interestingly, compared with patients without heart failure (HF), atrial NHE1 expression was significantly increased in patients with HF with preserved ejection fraction and atrial fibrillation. The largest increase in atrial and ventricular NHE1 expression, however, was observed in patients with end-stage HF undergoing heart transplantation. Importantly, acute exposure to empagliflozin (1 mu mol/L, 10 min) significantly inhibited NHE activity to a similar extent in human atrial myocytes and mouse ventricular myocytes. This inhibition was also achieved by incubation with the well-described selective NHE inhibitor cariporide (10 mu mol/L, 10 min). Conclusions This is the first study systematically analysing NHE1 expression in human atrial and ventricular myocardium of HF patients. We show that empagliflozin inhibits NHE in human cardiomyocytes. The extent of NHE inhibition was comparable with cariporide and may potentially contribute to the improved outcome of patients in clinical trials

The functional consequences of sodium channel NaV 1.8 in human left ventricular hypertrophy

Aims In hypertrophy and heart failure, the proarrhythmic persistent Na+ current (I-NaL) is enhanced. We aimed to investigate the electrophysiological role of neuronal sodium channel Na(V)1.8 in human hypertrophied myocardium. Methods and results Myocardial tissue of 24 patients suffering from symptomatic severe aortic stenosis and concomitant significant afterload-induced hypertrophy with preserved ejection fraction was used and compared with 12 healthy controls. We performed quantitative real-time PCR and western blot and detected a significant up-regulation of Na(V)1.8 mRNA (2.34-fold) and protein expression (1.96-fold) in human hypertrophied myocardium compared with healthy hearts. Interestingly, Na(V)1.5 protein expression was significantly reduced in parallel (0.60-fold). Using whole-cell patch-clamp technique, we found that the prominent I-NaL was significantly reduced after addition of novel Na(V)1.8-specific blockers either A-803467 (30 nM) or PF-01247324 (1 mu M) in human hypertrophic cardiomyocytes. This clearly demonstrates the relevant contribution of Na(V)1.8 to this proarrhythmic current. We observed a significant action potential duration shortening and performed confocal microscopy, demonstrating a 50% decrease in proarrhythmic diastolic sarcoplasmic reticulum (SR)-Ca2+ leak and SR-Ca2+ spark frequency after exposure to both Na(V)1.8 inhibitors. Conclusions We show for the first time that the neuronal sodium channel Na(V)1.8 is up-regulated on mRNA and protein level in the human hypertrophied myocardium. Furthermore, inhibition of Na(V)1.8 reduced augmented I-NaL, abbreviated the action potential duration, and decreased the SR-Ca2+ leak. The findings of our study suggest that Na(V)1.8 could be a promising antiarrhythmic therapeutic target and merits further investigation

Structural Analysis of Mitochondrial Dynamics—From Cardiomyocytes to Osteoblasts: A Critical Review

Mitochondria play a crucial role in cell physiology and pathophysiology. In this context, mitochondrial dynamics and, subsequently, mitochondrial ultrastructure have increasingly become hot topics in modern research, with a focus on mitochondrial fission and fusion. Thus, the dynamics of mitochondria in several diseases have been intensively investigated, especially with a view to developing new promising treatment options. However, the majority of recent studies are performed in highly energy-dependent tissues, such as cardiac, hepatic, and neuronal tissues. In contrast, publications on mitochondrial dynamics from the orthopedic or trauma fields are quite rare, even if there are common cellular mechanisms in cardiovascular and bone tissue, especially regarding bone infection. The present report summarizes the spectrum of mitochondrial alterations in the cardiovascular system and compares it to the state of knowledge in the musculoskeletal system. The present paper summarizes recent knowledge regarding mitochondrial dynamics and gives a short, but not exhaustive, overview of its regulation via fission and fusion. Furthermore, the article highlights hypoxia and its accompanying increased mitochondrial fission as a possible link between cardiac ischemia and inflammatory diseases of the bone, such as osteomyelitis. This opens new innovative perspectives not only for the understanding of cellular pathomechanisms in osteomyelitis but also for potential new treatment options

Enhanced Heart Failure in Redox‐Dead Cys17Ser PKARIα Knock‐In Mice

Background PKARIα (protein kinase A type I‐α regulatory subunit) is redox‐active independent of its physiologic agonist cAMP. However, it is unknown whether this alternative mechanism of PKARIα activation may be of relevance to cardiac excitation–contraction coupling. Methods and Results We used a redox‐dead transgenic mouse model with homozygous knock‐in replacement of redox‐sensitive cysteine 17 with serine within the regulatory subunits of PKARIα (KI). Reactive oxygen species were acutely evoked by exposure of isolated cardiac myocytes to AngII (angiotensin II, 1 µmol/L). The long‐term relevance of oxidized PKARIα was investigated in KI mice and their wild‐type (WT) littermates following transverse aortic constriction (TAC). AngII increased reactive oxygen species in both groups but with RIα dimer formation in WT only. AngII induced translocation of PKARI to the cell membrane and resulted in protein kinase A–dependent stimulation of ICa (L‐type Ca current) in WT with no effect in KI myocytes. Consequently, Ca transients were reduced in KI myocytes as compared with WT cells following acute AngII exposure. Transverse aortic constriction–related reactive oxygen species formation resulted in RIα oxidation in WT but not in KI mice. Within 6 weeks after TAC, KI mice showed an enhanced deterioration of contractile function and impaired survival compared with WT. In accordance, compared with WT, ventricular myocytes from failing KI mice displayed significantly reduced Ca transient amplitudes and lack of ICa stimulation. Conversely, direct pharmacological stimulation of ICa using Bay K8644 rescued Ca transients in AngII‐treated KI myocytes and contractile function in failing KI mice in vivo. Conclusions Oxidative activation of PKARIα with subsequent stimulation of ICa preserves cardiac function in the setting of acute and chronic oxidative stress

Detrimental proarrhythmogenic interaction of Ca2+/calmodulin-dependent protein kinase II and NaV1.8 in heart failure

An interplay between Ca2+/calmodulin-dependent protein kinase IIδc (CaMKIIδc) and late Na+ current (INaL) is known to induce arrhythmias in the failing heart. Here, we elucidate the role of the sodium channel isoform NaV1.8 for CaMKIIδc-dependent proarrhythmia. In a CRISPR-Cas9-generated human iPSC-cardiomyocyte homozygous knock-out of NaV1.8, we demonstrate that NaV1.8 contributes to INaL formation. In addition, we reveal a direct interaction between NaV1.8 and CaMKIIδc in cardiomyocytes isolated from patients with heart failure (HF). Using specific blockers of NaV1.8 and CaMKIIδc, we show that NaV1.8-driven INaL is CaMKIIδc-dependent and that NaV1.8-inhibtion reduces diastolic SR-Ca2+ leak in human failing cardiomyocytes. Moreover, increased mortality of CaMKIIδc-overexpressing HF mice is reduced when a NaV1.8 knock-out is introduced. Cellular and in vivo experiments reveal reduced ventricular arrhythmias without changes in HF progression. Our work therefore identifies a proarrhythmic CaMKIIδc downstream target which may constitute a prognostic and antiarrhythmic strategy

Tachycardiomyopathy entails a dysfunctional pattern of interrelated mitochondrial functions

Tachycardiomyopathy is characterised by reversible left ventricular dysfunction, provoked by rapid ventricular rate. While the knowledge of mitochondria advanced in most cardiomyopathies, mitochondrial functions await elucidation in tachycardiomyopathy. Pacemakers were implanted in 61 rabbits. Tachypacing was performed with 330 bpm for 10 days (n = 11, early left ventricular dysfunction) or with up to 380 bpm over 30 days (n = 24, tachycardiomyopathy, TCM). In n = 26, pacemakers remained inactive (SHAM). Left ventricular tissue was subjected to respirometry, metabolomics and acetylomics. Results were assessed for translational relevance using a human-based model: induced pluripotent stem cell derived cardiomyocytes underwent field stimulation for 7 days (TACH–iPSC–CM). TCM animals showed systolic dysfunction compared to SHAM (fractional shortening 37.8 ± 1.0% vs. 21.9 ± 1.2%, SHAM vs. TCM, p < 0.0001). Histology revealed cardiomyocyte hypertrophy (cross-sectional area 393.2 ± 14.5 µm2 vs. 538.9 ± 23.8 µm2, p < 0.001) without fibrosis. Mitochondria were shifted to the intercalated discs and enlarged. Mitochondrial membrane potential remained stable in TCM. The metabolite profiles of ELVD and TCM were characterised by profound depletion of tricarboxylic acid cycle intermediates. Redox balance was shifted towards a more oxidised state (ratio of reduced to oxidised nicotinamide adenine dinucleotide 10.5 ± 2.1 vs. 4.0 ± 0.8, p < 0.01). The mitochondrial acetylome remained largely unchanged. Neither TCM nor TACH–iPSC–CM showed relevantly increased levels of reactive oxygen species. Oxidative phosphorylation capacity of TCM decreased modestly in skinned fibres (168.9 ± 11.2 vs. 124.6 ± 11.45 pmol·O2·s−1·mg−1 tissue, p < 0.05), but it did not in isolated mitochondria. The pattern of mitochondrial dysfunctions detected in two models of tachycardiomyopathy diverges from previously published characteristic signs of other heart failure aetiologies