Single dose of intravenous miR199a-5p delivery targeting ischemic heart for long-term repair of myocardial infarction.

Long-term treatment of myocardial infarction is challenging despite medical advances. Tissue engineering shows promise for MI repair, but implantation complexity and uncertain outcomes pose obstacles. microRNAs regulate genes involved in apoptosis, angiogenesis, and myocardial contraction, making them valuable for long-term repair. In this study, we find downregulated miR-199a-5p expression in MI. Intramyocardial injection of miR-199a-5p into the infarcted region of male rats revealed its dual protective effects on the heart. Specifically, miR-199a-5p targets AGTR1, diminishing early oxidative damage post-myocardial infarction, and MARK4, which influences long-term myocardial contractility and enhances cardiac function. To deliver miR-199a-5p efficiently and specifically to ischemic myocardial tissue, we use CSTSMLKAC peptide to construct P-MSN/miR199a-5p nanoparticles. Intravenous administration of these nanoparticles reduces myocardial injury and protects cardiac function. Our findings demonstrate the effectiveness of P-MSN/miR199a-5p nanoparticles in repairing MI through enhanced contraction and anti-apoptosis. miR199a-5p holds significant therapeutic potential for long-term repair of myocardial infarction.

Cardiotoxicity of the diamide insecticide chlorantraniliprole in the intact heart and in isolated cardiomyocytes from the honey bee.

In honey bees, circulation of blood (hemolymph) is driven by the peristaltic contraction of the heart vessel located in the dorsal part of the abdomen. Chlorantraniliprole (CHL) is an insecticide of the anthranilic diamide class which main mode of action is to alter the function of intracellular Ca2+ release channels (known as RyRs, for ryanodine receptors). In the honey bee, it was recently found to be more toxic when applied on the dorsal part of the abdomen, suggesting a direct cardiotoxicity. In the present study, a short-term exposure of semi-isolated bee hearts to CHL (0.1-10 µM) induces alterations of cardiac contraction. These alterations range from a slow-down of systole and diastole kinetics, to bradycardia and cardiac arrest. The bees heart wall is made of a single layer of semi-circular cardiomyocytes arranged concentrically all along the long axis of tube lumen. Since the heart tube is suspended to the cuticle through long tubular muscles fibers (so-called alary muscle cells), the CHL effects in ex-vivo heart preparations could result from the modulation of RyRs present in these skeletal muscle fibers as well as cardiomyocytes RyRs themselves. In order to specifically assess effects of CHL on cardiomyocytes, for the first time, intact heart cells were enzymatically dissociated from bees. Exposure of cardiomyocytes to CHL induces an increase in cytoplasmic calcium, cell contraction at the highest concentrations and depletion of intracellular stores. Electrophysiological properties of isolated cardiomyocytes were described, with a focus on voltage-gated Ca2+ channels responsible for the cardiac action potentials depolarization phase. Two types of Ca2+ currents were measured under voltage-clamp. Exposure to CHL was accompanied by a decrease in voltage-activated Ca2+ currents densities. Altogether, these results show that chlorantraniliprole can cause cardiac defects in honey bees.

Tracking single hiPSC-derived cardiomyocyte contractile function using CONTRAX an efficient pipeline for traction force measurement.

Cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs) are powerful in vitro models to study the mechanisms underlying cardiomyopathies and cardiotoxicity. Quantification of the contractile function in single hiPSC-CMs at high-throughput and over time is essential to disentangle how cellular mechanisms affect heart function. Here, we present CONTRAX, an open-access, versatile, and streamlined pipeline for quantitative tracking of the contractile dynamics of single hiPSC-CMs over time. Three software modules enable: parameter-based identification of single hiPSC-CMs; automated video acquisition of >200 cells/hour; and contractility measurements via traction force microscopy. We analyze >4,500 hiPSC-CMs over time in the same cells under orthogonal conditions of culture media and substrate stiffnesses; +/- drug treatment; +/- cardiac mutations. Using undirected clustering, we reveal converging maturation patterns, quantifiable drug response to Mavacamten and significant deficiencies in hiPSC-CMs with disease mutations. CONTRAX empowers researchers with a potent quantitative approach to develop cardiac therapies.

Omecamtiv mecarbil and Mavacamten target the same myosin pocket despite opposite effects in heart contraction.

Inherited cardiomyopathies are common cardiac diseases worldwide, leading in the late stage to heart failure and death. The most promising treatments against these diseases are small molecules directly modulating the force produced by ß-cardiac myosin, the molecular motor driving heart contraction. Omecamtiv mecarbil and Mavacamten are two such molecules that completed phase 3 clinical trials, and the inhibitor Mavacamten is now approved by the FDA. In contrast to Mavacamten, Omecamtiv mecarbil acts as an activator of cardiac contractility. Here, we reveal by X-ray crystallography that both drugs target the same pocket and stabilize a pre-stroke structural state, with only few local differences. All-atom molecular dynamics simulations reveal how these molecules produce distinct effects in motor allostery thus impacting force production in opposite way. Altogether, our results provide the framework for rational drug development for the purpose of personalized medicine.

Inhibition of ER stress using tauroursodeoxycholic acid rescues obesity-evoked cardiac remodeling and contractile anomalies through regulation of ferroptosis.

Interrupted ER homeostasis contributes to the etiology of obesity cardiomyopathy although it remains elusive how ER stress evokes cardiac anomalies in obesity. Our study evaluated the impact of ER stress inhibition on cardiac anomalies in obesity. Lean and ob/ob obese mice received chemical ER chaperone tauroursodeoxycholic acid (TUDCA, 50 mg/kg/d, p.o.) for 35 days prior to evaluation of glucose sensitivity, echocardiographic, myocardial geometric, cardiomyocyte mechanical and subcellular Ca2+ property, mitochondrial integrity, oxidative stress, apoptosis, and ferroptosis. Intracellular Ca2+ governing domains including sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA) were monitored by45Ca2+uptake and immunoblotting. Our results noted that TUDCA alleviated myocardial remodeling (fibrosis, hypertrophy, enlarged LVESD), echocardiographic anomalies (compromised fractional shortening and ejection fraction), cardiomyocyte contractile dysfunction (amplitude and velocity of cell shortening, relengthening time) and intracellular Ca2+ anomalies (compromised subcellular Ca2+ release, clearance and SERCA function), mitochondrial damage (collapsed membrane potential, downregulated mitochondrial elements and ultrastructural alteration), ER stress (GRP78, eIF2α and ATF4), oxidative stress, apoptosis and ferroptosis [downregulated SLC7A11, GPx4 and upregulated transferrin receptor (TFRC)] without affecting global glucose sensitivity and serum Fe2+ in obese mice. Obesity-evoked change in HSP90, phospholamban and Na+-Ca2+ exchanger was spared by the chemical ER chaperone. Moreover, in vitro results noted that TUDCA, PERK inhibitor GSK2606414, TFRC neutralizing antibody and ferroptosis inhibitor LIP1 mitigated palmitic acid-elicited changes in lipid peroxidation and mechanical function. Our findings favored a role for ferroptosis in obesity cardiomyopathy downstream of ER stress.

Silk fibroin increases the elasticity of alginate-gelatin hydrogels and regulates cardiac cell contractile function in cardiac bioinks.

Silk fibroin (SF) is a natural protein extracted fromBombyx morisilkworm thread. From its common use in the textile industry, it emerged as a biomaterial with promising biochemical and mechanical properties for applications in the field of tissue engineering and regenerative medicine. In this study, we evaluate for the first time the effects of SF on cardiac bioink formulations containing cardiac spheroids (CSs). First, we evaluate if the SF addition plays a role in the structural and elastic properties of hydrogels containing alginate (Alg) and gelatin (Gel). Then, we test the printability and durability of bioprinted SF-containing hydrogels. Finally, we evaluate whether the addition of SF controls cell viability and function of CSs in Alg-Gel hydrogels. Our findings show that the addition of 1% (w/v) SF to Alg-Gel hydrogels makes them more elastic without affecting cell viability. However, fractional shortening (FS%) of CSs in SF-Alg-Gel hydrogels increases without affecting their contraction frequency, suggesting an improvement in contractile function in the 3D cultures. Altogether, our findings support a promising pathway to bioengineer bioinks containing SF for cardiac applications, with the ability to control mechanical and cellular features in cardiac bioinks.

Activation of Cardiac δ2-Opioid Receptors Increases Heart Tolerance to Reperfusion.

Coronary occlusion (45 min) and reperfusion (120 min) in male Wistar rats in vivo, as well as total ischemia (45 min) of an isolated rat heart followed by reperfusion (30 min) were reproduced. The selective δ2-opioid receptor agonist deltorphin II (0.12 mg/kg and 152 nmol/liter) was administered intravenously 5 min before reperfusion in vivo or added to the perfusion solution at the beginning of reperfusion of the isolated heart. The peripheral opioid receptor antagonist naloxone methiodide and δ2-opioid receptor antagonist naltriben were used in doses of 5 and 0.3 mg/kg, respectively. It was found that the infarct-limiting effect of deltorphin II is associated with the activation of δ2-opioid receptors. We have demonstrated that deltorphin II can improve the recovery of the contractility of the isolated heart after total ischemia.

Impact of quetiapine on ion channels and contractile dynamics in rat ventricular myocyte.

Antipsychotic drugs often lead to adverse effects, including those related to the cardiovascular system. Of these, quetiapine is known to cause significant changes in the QT interval although the underlying mechanism remains mysterious, prompting us to examine its effects on cardiac electrophysiological properties. Therefore, we investigated the effect of quetiapine on contraction, action potential (AP), and the associated membrane currents such as L-type Ca2+ and K+ using the whole-cell patch clamp method to examine its impacts on isolated rat ventricular myocytes. Our results showed that (1) quetiapine reduces cell contractility in a concentration-dependent manner and (2) leads to a significant prolongation in the duration of AP in isolated ventricular myocytes. This effect was both concentration and frequency-dependent; (3) quetiapine significantly decreased the Ca2+, transient outward K+, and steady-state K+ currents. However, only high concentration of quetiapine (100 µM) could significantly change the activation and reactivation kinetics of L-type Ca2+ channels. This study demonstrates that QT extension induced by quetiapine is mainly associated with the prolongation of AP. Moreover, quetiapine caused a significant decrease in contractile force and excitability of ventricular myocytes by suppressing Ca2+ and K+ currents.

Aficamten for Symptomatic Obstructive Hypertrophic Cardiomyopathy.

BACKGROUND: One of the major determinants of exercise intolerance and limiting symptoms among patients with obstructive hypertrophic cardiomyopathy (HCM) is an elevated intracardiac pressure resulting from left ventricular outflow tract obstruction. Aficamten is an oral selective cardiac myosin inhibitor that reduces left ventricular outflow tract gradients by mitigating cardiac hypercontractility. METHODS: In this phase 3, double-blind trial, we randomly assigned adults with symptomatic obstructive HCM to receive aficamten (starting dose, 5 mg; maximum dose, 20 mg) or placebo for 24 weeks, with dose adjustment based on echocardiography results. The primary end point was the change from baseline to week 24 in the peak oxygen uptake as assessed by cardiopulmonary exercise testing. The 10 prespecified secondary end points (tested hierarchically) were change in the Kansas City Cardiomyopathy Questionnaire clinical summary score (KCCQ-CSS), improvement in the New York Heart Association (NYHA) functional class, change in the pressure gradient after the Valsalva maneuver, occurrence of a gradient of less than 30 mm Hg after the Valsalva maneuver, and duration of eligibility for septal reduction therapy (all assessed at week 24); change in the KCCQ-CSS, improvement in the NYHA functional class, change in the pressure gradient after the Valsalva maneuver, and occurrence of a gradient of less than 30 mm Hg after the Valsalva maneuver (all assessed at week 12); and change in the total workload as assessed by cardiopulmonary exercise testing at week 24. RESULTS: A total of 282 patients underwent randomization: 142 to the aficamten group and 140 to the placebo group. The mean age was 59.1 years, 59.2% were men, the baseline mean resting left ventricular outflow tract gradient was 55.1 mm Hg, and the baseline mean left ventricular ejection fraction was 74.8%. At 24 weeks, the mean change in the peak oxygen uptake was 1.8 ml per kilogram per minute (95% confidence interval [CI], 1.2 to 2.3) in the aficamten group and 0.0 ml per kilogram per minute (95% CI, -0.5 to 0.5) in the placebo group (least-squares mean between-group difference, 1.7 ml per kilogram per minute; 95% CI, 1.0 to 2.4; P<0.001). The results for all 10 secondary end points were significantly improved with aficamten as compared with placebo. The incidence of adverse events appeared to be similar in the two groups. CONCLUSIONS: Among patients with symptomatic obstructive HCM, treatment with aficamten resulted in a significantly greater improvement in peak oxygen uptake than placebo. (Funded by Cytokinetics; SEQUOIA-HCM ClinicalTrials.gov number, NCT05186818.).

The left ventricle increases contractility in response to baroreceptor unloading, which is sympathetically mediated in the anesthetized rat.

Contemporary discussion of the baroreflex includes the efferent vascular-sympathetic and cardiovagal arms. Since sympathetic postganglionic neurons also innervate the left ventricle (LV), it is often assumed that the LV produces a sympathetically mediated increase in contractility during baroreceptor unloading, but this has not been characterized using a load-independent index of contractility. We aimed to determine 1) whether LV contractility increases in response to baroreceptor unloading and 2) whether such increases are mediated via the sympathetic or parasympathetic arm of the autonomic nervous system. Ten male Wistar rats were anesthetized (urethane) and instrumented with arterial and LV pressure-volume catheters to measure mean arterial pressure (MAP) and load-independent LV contractility [maximal rate of increase in pressure adjusted to end-diastolic volume (PAdP/dtmax)], respectively. Rats were placed in a servo-controlled lower-body negative pressure (LBNP) chamber to reduce MAP by 10% for 60 s to mechanically unload baroreceptors under control conditions. LBNP was repeated in each animal following infusions of cardiac autonomic blockers using esmolol (sympathetic), atropine (parasympathetic), and esmolol + atropine. Under control conditions, PAdP/dtmax increased during baroreceptor unloading (26 ± 6 vs. 31 ± 9 mmHg·s-1·µL-1, P = 0.031). During esmolol, there was no increase in LV contractility during baroreceptor unloading (11 ± 2 vs. 12 ± 2, P = 0.125); however, during atropine, there was an increase in LV contractility during baroreceptor unloading (26 ± 6 vs. 31 ± 9, P = 0.019). During combined esmolol and atropine, there was a small increase in contractility versus control (13 ± 3 vs. 15 ± 4, P = 0.046). Our results demonstrate that, in anesthetized rats, LV contractility increases in response to baroreceptor unloading, which is largely sympathetically mediated.NEW & NOTEWORTHY This study empirically demonstrates a sympathetically mediated increase in LV contractility in response to baroreceptor unloading using a load-independent index of cardiac contractility in the anesthetized rat.

Interferometric Biosensor for High Sensitive Label-Free Recording of HiPS Cardiomyocytes Contraction in Vitro.

Heart disease remains a leading cause of global mortality, underscoring the need for advanced technologies to study cardiovascular diseases and develop effective treatments. We introduce an innovative interferometric biosensor for high-sensitivity and label-free recording of human induced pluripotent stem cell (hiPSC) cardiomyocyte contraction in vitro. Using an optical cavity, our device captures interference patterns caused by the contraction-induced displacement of a thin flexible membrane. First, we demonstrate the capability to quantify spontaneous contractions and discriminate between contraction and relaxation phases. We calculate a contraction-induced vertical membrane displacement close to 40 nm, which implies a traction stress of 34 ± 4 mN/mm2. Finally, we investigate the effects of a drug compound on contractility amplitude, revealing a significant reduction in contractile forces. The label-free and high-throughput nature of our biosensor may enhance drug screening processes and drug development for cardiac treatments. Our interferometric biosensor offers a novel approach for noninvasive and real-time assessment of cardiomyocyte contraction.

Discovery of Nelutroctiv (CK-136), a Selective Cardiac Troponin Activator for the Treatment of Cardiovascular Diseases Associated with Reduced Cardiac Contractility.

Cardiac myosin activation has been shown to be a viable approach for the treatment of heart failure with reduced ejection fraction. Here, we report the discovery of nelutroctiv (CK-136), a selective cardiac troponin activator intended for patients with cardiovascular conditions where cardiac contractility is reduced. Discovery of nelutroctiv began with a high-throughput screen that identified compound 1R, a muscle selective cardiac sarcomere activator devoid of phosphodiesterase-3 activity. Optimization of druglike properties for 1R led to the replacement of the sulfonamide and aniline substituents which resulted in improved pharmacokinetic (PK) profiles and a reduced potential for human drug-drug interactions. In vivo echocardiography assessment of the optimized leads showed concentration dependent increases in fractional shortening and an improved pharmacodynamic window compared to myosin activator CK-138. Overall, nelutroctiv was found to possess the desired selectivity, a favorable pharmacodynamic window relative to myosin activators, and a preclinical PK profile to support clinical development.

Short-Chain Fatty Acid Butyrate Is an Inotropic Agent With Vasorelaxant and Cardioprotective Properties.

BACKGROUND: The heart can metabolize the microbiota-derived short-chain fatty acid butyrate. Butyrate may have beneficial effects in heart failure, but the underlying mechanisms are unknown. We tested the hypothesis that butyrate elevates cardiac output by mechanisms involving direct stimulation of cardiac contractility and vasorelaxation in rats. METHODS AND RESULTS: We examined the effects of butyrate on (1) in vivo hemodynamics using parallel echocardiographic and invasive blood pressure measurements, (2) isolated perfused hearts in Langendorff systems under physiological conditions and after ischemia and reperfusion, and (3) isolated coronary arteries mounted in isometric wire myographs. We tested Na-butyrate added to injection solutions or physiological buffers and compared its effects with equimolar doses of NaCl. Butyrate at plasma concentrations of 0.56 mM increased cardiac output by 48.8±14.9%, stroke volume by 38.5±12.1%, and left ventricular ejection fraction by 39.6±6.2%, and lowered systemic vascular resistance by 33.5±6.4% without affecting blood pressure or heart rate in vivo. In the range between 0.1 and 5 mM, butyrate increased left ventricular systolic pressure by up to 23.7±3.4% in isolated perfused hearts and by 9.4±2.9% following ischemia and reperfusion, while reducing myocardial infarct size by 81.7±16.9%. Butyrate relaxed isolated coronary septal arteries concentration dependently with an EC50=0.57 mM (95% CI, 0.23-1.44). CONCLUSIONS: We conclude that butyrate elevates cardiac output through mechanisms involving increased cardiac contractility and vasorelaxation. This effect of butyrate was not associated with adverse myocardial injury in damaged hearts exposed to ischemia and reperfusion.

Cardiac dysfunction in sucrose-fed rats is associated with alterations of phospholamban phosphorylation and TNF-α levels.

INTRODUCTION: High sucrose intake is linked to cardiovascular disease, a major global cause of mortality worldwide. Calcium mishandling and inflammation play crucial roles in cardiac disease pathophysiology. OBJECTIVE: Evaluate if sucrose-induced obesity is related to deterioration of myocardial function due to alterations in the calcium-handling proteins in association with proinflammatory cytokines. METHODS: Wistar rats were divided into control and sucrose groups. Over eight weeks, Sucrose group received 30% sucrose water. Cardiac function was determined in vivo using echocardiography and in vitro using papillary muscle assay. Western blotting was used to detect calcium handling protein; ELISA assay was used to assess TNF-α and IL-6 levels. RESULTS: Sucrose led to cardiac dysfunction. RYR2, SERCA2, NCX, pPBL Ser16 and L-type calcium channels were unchanged. However, pPBL-Thr17, and TNF-α levels were elevated in the S group. CONCLUSION: Sucrose induced cardiac dysfunction and decreased myocardial contractility in association with altered pPBL-Thr17 and elevated cardiac pro-inflammatory TNF-α.

In silico study of the mechanisms of hypoxia and contractile dysfunction during ischemia and reperfusion of hiPSC cardiomyocytes.

Interconnected mechanisms of ischemia and reperfusion (IR) has increased the interest in IR in vitro experiments using human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). We developed a whole-cell computational model of hiPSC-CMs including the electromechanics, a metabolite-sensitive sarcoplasmic reticulum Ca2+-ATPase (SERCA) and an oxygen dynamics formulation to investigate IR mechanisms. Moreover, we simulated the effect and action mechanism of levosimendan, which recently showed promising anti-arrhythmic effects in hiPSC-CMs in hypoxia. The model was validated using hiPSC-CM and in vitro animal data. The role of SERCA in causing relaxation dysfunction in IR was anticipated to be comparable to its function in sepsis-induced heart failure. Drug simulations showed that levosimendan counteracts the relaxation dysfunction by utilizing a particular Ca2+-sensitizing mechanism involving Ca2+-bound troponin C and Ca2+ flux to the myofilament, rather than inhibiting SERCA phosphorylation. The model demonstrates extensive characterization and promise for drug development, making it suitable for evaluating IR therapy strategies based on the changing levels of cardiac metabolites, oxygen and molecular pathways.