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.
The complex physiological signal transduction networks that respond to the dual challenges of inflammatory and oxidative stress are major factors that promote the development of cardiovascular ...pathologies. These signaling networks contribute to the development of age-related diseases, suggesting crosstalk between the development of aging and cardiovascular disease. Inhibition and/or attenuation of these signaling networks also delays the onset of disease. Therefore, a concept of targeting the signaling networks that are involved in inflammation and oxidative stress may represent a novel treatment paradigm for many types of heart disease. In this review, we discuss the molecular mechanisms associated with the physiological responses to inflammation and oxidative stress especially in heart failure with preserved ejection fraction and emphasize the nature of the crosstalk of these signaling processes as well as possible therapeutic implications for cardiovascular medicine.
Atrial fibrillation (AF) and heart failure often coexist, but their interaction is poorly understood. Clinical data indicate that the arrhythmic component of AF may contribute to left ventricular ...(LV) dysfunction.
This study investigates the effects and molecular mechanisms of AF on the human LV.
Ventricular myocardium from patients with aortic stenosis and preserved LV function with sinus rhythm or rate-controlled AF was studied. LV myocardium from patients with sinus rhythm and patients with AF showed no differences in fibrosis. In functional studies, systolic Ca
transient amplitude of LV cardiomyocytes was reduced in patients with AF, while diastolic Ca
levels and Ca
transient kinetics were not statistically different. These results were confirmed in LV cardiomyocytes from nonfailing donors with sinus rhythm or AF. Moreover, normofrequent AF was simulated in vitro using arrhythmic or rhythmic pacing (both at 60 bpm). After 24 hours of AF-simulation, human LV cardiomyocytes from nonfailing donors showed an impaired Ca
transient amplitude. For a standardized investigation of AF-simulation, human iPSC-cardiomyocytes were tested. Seven days of AF-simulation caused reduced systolic Ca
transient amplitude and sarcoplasmic reticulum Ca
load likely because of an increased diastolic sarcoplasmic reticulum Ca
leak. Moreover, cytosolic Na
concentration was elevated and action potential duration was prolonged after AF-simulation. We detected an increased late Na
current as a potential trigger for the detrimentally altered Ca
/Na
-interplay. Mechanistically, reactive oxygen species were higher in the LV of patients with AF. CaMKII (Ca
/calmodulin-dependent protein kinase IIδc) was found to be more oxidized at Met281/282 in the LV of patients with AF leading to an increased CaMKII activity and consequent increased RyR2 phosphorylation. CaMKII inhibition and ROS scavenging ameliorated impaired systolic Ca
handling after AF-simulation.
AF causes distinct functional and molecular remodeling of the human LV. This translational study provides the first mechanistic characterization and the potential negative impact of AF in the absence of tachycardia on the human ventricle.
Cardiac type 2 ryanodine receptors (RyR
s) play a pivotal role in cellular electrophysiology and contractility. Increased RyR
-mediated diastolic sarcoplasmic reticulum (SR) Ca
release is linked to ...heart failure (HF) and arrhythmias. Dantrolene, a drug used for the treatment of malignant hyperthermia, is known to stabilize RyRs in skeletal muscle.
The purpose of this study was to investigate the effects of dantrolene on arrhythmogenic triggers and contractile function in human atrial fibrillation (AF) and HF cardiomyocytes (CM).
Human CM were isolated from either patients with HF (ventricular) or patients with AF (atrial), and Ca
imaging, patch-clamp, or muscle strip experiments were performed.
After exposure to dantrolene, human atrial AF and left ventricular HF CM showed significant reductions in proarrhythmic SR Ca
spark frequency and diastolic SR Ca
leak. Moreover, dantrolene decreased the frequency of Ca
waves and spontaneous Ca
transients in HF CM. Patch-clamp experiments revealed that dantrolene significantly suppressed delayed afterdepolarizations in HF and AF CM. Importantly, dantrolene had no effect on action potential duration in AF or in HF CM. In addition, dantrolene had neutral effects on contractile force of human isometrically twitching ventricular HF trabeculae.
Our study showed that dantrolene beneficially influenced disrupted SR Ca
homeostasis in human HF and AF CM. Cellular arrhythmogenic triggers were potently suppressed by dantrolene, whereas action potential duration and contractility were not affected. As a clinically approved drug for the treatment of malignant hyperthermia, dantrolene may be a potential antiarrhythmic drug for patients with rhythm disorders and merits further clinical investigation.
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 NH4+ 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 μmol/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 μmol/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.
Abstract
An interplay between Ca
2+
/calmodulin-dependent protein kinase IIδc (CaMKIIδc) and late Na
+
current (I
NaL
) is known to induce arrhythmias in the failing heart. Here, we elucidate the ...role of the sodium channel isoform Na
V
1.8 for CaMKIIδc-dependent proarrhythmia. In a CRISPR-Cas9-generated human iPSC-cardiomyocyte homozygous knock-out of Na
V
1.8, we demonstrate that Na
V
1.8 contributes to I
NaL
formation. In addition, we reveal a direct interaction between Na
V
1.8 and CaMKIIδc in cardiomyocytes isolated from patients with heart failure (HF). Using specific blockers of Na
V
1.8 and CaMKIIδc, we show that Na
V
1.8-driven I
NaL
is CaMKIIδc-dependent and that Na
V
1.8-inhibtion reduces diastolic SR-Ca
2+
leak in human failing cardiomyocytes. Moreover, increased mortality of CaMKIIδc-overexpressing HF mice is reduced when a Na
V
1.8 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.
Aims
The EMPA‐REG OUTCOME study showed reduced mortality and hospitalization due to heart failure (HF) in diabetic patients treated with empagliflozin. Overexpression and Ca2+‐dependent activation of ...Ca2+/calmodulin‐dependent kinase II (CaMKII) are hallmarks of HF, leading to contractile dysfunction and arrhythmias. We tested whether empagliflozin reduces CaMKII‐ activity and improves Ca2+‐handling in human and murine ventricular myocytes.
Methods and results
Myocytes from wild‐type mice, mice with transverse aortic constriction (TAC) as a model of HF, and human failing ventricular myocytes were exposed to empagliflozin (1 μmol/L) or vehicle. CaMKII activity was assessed by CaMKII–histone deacetylase pulldown assay. Ca2+ spark frequency (CaSpF) as a measure of sarcoplasmic reticulum (SR) Ca2+ leak was investigated by confocal microscopy. Na+i was measured using Na+/Ca2+‐exchanger (NCX) currents (whole‐cell patch clamp). Compared with vehicle, 24 h empagliflozin exposure of murine myocytes reduced CaMKII activity (1.6 ± 0.7 vs. 4.2 ± 0.9, P < 0.05, n = 10 mice), and also CaMKII‐dependent ryanodine receptor phosphorylation (0.8 ± 0.1 vs. 1.0 ± 0.1, P < 0.05, n = 11 mice), with similar results upon TAC. In murine myocytes, empagliflozin reduced CaSpF (TAC: 1.7 ± 0.3 vs. 2.5 ± 0.4 1/100 μm−1 s−1, P < 0.05, n = 4 mice) but increased SR Ca2+ load and Ca2+ transient amplitude. Importantly, empagliflozin also significantly reduced CaSpF in human failing ventricular myocytes (1 ± 0.2 vs. 3.3 ± 0.9, P < 0.05, n = 4 patients), while Ca2+ transient amplitude was increased (F/F0: 0.53 ± 0.05 vs. 0.36 ± 0.02, P < 0.05, n = 3 patients). In contrast, 30 min exposure with empagliflozin did not affect CaMKII activity nor Ca2+‐handling but significantly reduced Na+i.
Conclusions
We show for the first time that empagliflozin reduces CaMKII activity and CaMKII‐dependent SR Ca2+ leak. Reduced Ca2+ leak and improved Ca2+ transients may contribute to the beneficial effects of empagliflozin in HF.
The SGLT2 inhibitor empagliflozin improved cardiovascular outcomes in patients with diabetes. As the cardiac mechanisms remain elusive, we investigated the long-term effects (up to 2 months) of ...empagliflozin on excitation-contraction (EC)-coupling in human cardiomyocytes derived from induced pluripotent stem cells (iPSC-CM) in a blinded manner. IPSC from 3 donors, differentiated into pure iPSC-CM (4 differentiations), were treated with a clinically relevant concentration of empagliflozin (0.5 μmol/l) or vehicle control. Treatment, data acquisition, and analysis were conducted externally blinded. Epifluorescence microscopy measurements in iPSC-CM showed that empagliflozin has neutral effects on Ca
2+
transient amplitude, diastolic Ca
2+
levels, Ca
2+
transient kinetics, or sarcoplasmic Ca
2+
load after 2 weeks or 8 weeks of treatment. Confocal microscopy determining possible effects on proarrhythmogenic diastolic Ca
2+
release events showed that in iPSC-CM, Ca
2+
spark frequency and leak was not altered after chronic treatment with empagliflozin. Finally, in patch-clamp experiments, empagliflozin did not change action potential duration, amplitude, or resting membrane potential compared with vehicle control after long-term treatment. Next-generation RNA sequencing (NGS) and mapped transcriptome profiles of iPSC-CMs untreated and treated with empagliflozin for 8 weeks showed no differentially expressed EC-coupling genes. In line with NGS data, Western blots indicate that empagliflozin has negligible effects on key EC-coupling proteins. In this blinded study, direct treatment of iPSC-CM with empagliflozin for a clinically relevant duration of 2 months did not influence cardiomyocyte EC-coupling and electrophysiology. Therefore, it is likely that other mechanisms independent of cardiomyocyte EC-coupling are responsible for the beneficial treatment effect of empagliflozin.
Key messages
This blinded study investigated the clinically relevant long-term effects (up to 2 months) of empagliflozin on cardiomyocyte excitation-contraction (EC)-coupling.
Human cardiomyocytes derived from induced pluripotent stem cells (iPSC-CM) were used to study a human model including a high repetition number of experiments.
Empagliflozin has neutral effects on cardiomyocyte Ca
2+
transients, sarcoplasmic Ca
2+
load, and diastolic sarcoplasmic Ca
2+
leak.
In patch-clamp experiments, empagliflozin did not change the action potential.
Next-generation RNA sequencing, mapped transcriptome profiles, and Western blots of iPSC-CM untreated and treated with empagliflozin showed no differentially expressed EC-coupling candidates.
Abstract
Aims
In heart failure (HF), enhanced persistent Na+ current (INaL) exerts detrimental effects on cellular electrophysiology and can induce arrhythmias. However, the underlying regulatory ...mechanisms remain unclear. Our aim was to potentially investigate the regulation and electrophysiological contribution of neuronal sodium channel NaV1.8 in failing human heart and eventually to reveal a novel anti-arrhythmic therapy.
Methods and results
By western blot, we found that NaV1.8 protein expression is significantly up-regulated, while of the predominant cardiac isoform NaV1.5 is inversely reduced in human HF. Furthermore, to investigate the relation of NaV1.8 regulation with the cellular proarrhythmic events, we performed comprehensive electrophysiology recordings and explore the effect of NaV1.8 on INaL, action potential duration (APD), Ca2+ spark frequency, and arrhythmia induction in human failing cardiomyocytes. NaV1.8 inhibition with the specific blockers A-803467 and PF-01247324 decreased INaL, abbreviated APD and reduced cellular-spontaneous Ca2+-release and proarrhythmic events in human failing cardiomyocytes. Consistently, in mouse cardiomyocytes stressed with isoproterenol, pharmacologic inhibition and genetically knockout of NaV1.8 (SCN10A−/−), were associated with reduced INaL and abbreviated APD.
Conclusion
We provide first evidence of differential regulation of NaV1.8 and NaV1.5 in the failing human myocardium and their contribution to arrhythmogenesis due to generation of INaL. We propose inhibition of NaV1.8 thus constitutes a promising novel approach for selective anti-arrhythmic therapy in HF.
Pharmacologic approaches for the treatment of atrial arrhythmias are limited due to side effects and low efficacy. Thus, the identification of new antiarrhythmic targets is of clinical interest. ...Recent genome studies suggested an involvement of SCN10A sodium channels (Na
V
1.8) in atrial electrophysiology. This study investigated the role and involvement of Na
V
1.8 (SCN10A) in arrhythmia generation in the human atria and in mice lacking Na
V
1.8. Na
V
1.8 mRNA and protein were detected in human atrial myocardium at a significant higher level compared to ventricular myocardium. Expression of Na
V
1.8 and Na
V
1.5 did not differ between myocardium from patients with atrial fibrillation and sinus rhythm. To determine the electrophysiological role of Na
V
1.8, we investigated isolated human atrial cardiomyocytes from patients with sinus rhythm stimulated with isoproterenol. Inhibition of Na
V
1.8 by A-803467 or PF-01247324 showed no effects on the human atrial action potential. However, we found that Na
V
1.8 significantly contributes to late Na
+
current and consequently to an increased proarrhythmogenic diastolic sarcoplasmic reticulum Ca
2+
leak in human atrial cardiomyocytes. Selective pharmacological inhibition of Na
V
1.8 potently reduced late Na
+
current, proarrhythmic diastolic Ca
2+
release, delayed afterdepolarizations as well as spontaneous action potentials. These findings could be confirmed in murine atrial cardiomyocytes from wild-type mice and also compared to SCN10A
−/−
mice (genetic ablation of Na
V
1.8). Pharmacological Na
V
1.8 inhibition showed no effects in SCN10A
−/−
mice. Importantly, in vivo experiments in SCN10A
−/−
mice showed that genetic ablation of Na
V
1.8 protects against atrial fibrillation induction. This study demonstrates that Na
V
1.8 is expressed in the murine and human atria and contributes to late Na
+
current generation and cellular arrhythmogenesis. Blocking Na
V
1.8 selectively counteracts this pathomechanism and protects against atrial arrhythmias. Thus, our translational study reveals a new selective therapeutic target for treating atrial arrhythmias.
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