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Key points Chronic fetal hypoxia is one of the most common complications of pregnancy and can programme cardiac abnormalities in adult offspring including ventricular remodelling, diastolic dysfunction and sympathetic dominance. However, the underlying mechanisms at the level of the cardiomyocyte are unknown, preventing the identification of targets for therapeutic intervention. Therefore, we aimed to link echocardiographic data with cardiomyocyte function to reveal cellular mechanism for cardiac dysfunction in rat offspring from hypoxic pregnancy. Further, we investigated the potential of maternal treatment with melatonin as antenatal antioxidant therapy. Wistar rats were randomly allocated into normoxic (21% O2) or hypoxic (13% O2) pregnancy with or without melatonin treatment (5 µg/ml; normoxic melatonin in the maternal drinking water from gestational day 6 to 20 (term = 22 days). After delivery, male and female offspring were maintained to adulthood (16 weeks). Cardiomyocytes were isolated from the left and right ventricles, and calcium (Ca2+) handling was investigated in field‐stimulated myocytes. Systolic and diastolic function was negatively impacted in male and female offspring of hypoxic pregnancy demonstrating biventricular systolic and diastolic dysfunction and compensatory increases in cardiac output. Ca2+ transients from isolated cardiomyocytes in offspring of both sexes in hypoxic pregnancy displayed diastolic dysfunction with a reduced rate of [Ca2+]i recovery. Cardiac and cardiomyocyte dysfunction in male and female adult offspring was ameliorated by maternal antenatal treatment with melatonin in hypoxic pregnancy. Therefore, cardiomyocyte Ca2+ mishandling provides a cellular mechanism explaining functional deficits in hearts of male and female offspring in pregnancies complicated by chronic fetal hypoxia. This study identified significant changes in Ca2+ handling within cardiomyocytes isolated from offspring of hypoxic pregnancy including reduced systolic Ca2+ transients, impaired diastolic recovery of [Ca2+]i and a greater increase in systolic [Ca2+]i amplitude to β‐adrenergic stimulation. These changes in cardiomyocyte Ca2+ handling help to explain dysregulation of biventricular systolic and diastolic dysfunction determined by echocardiography. The data show protection against maladaptive cardiomyocyte calcium handling and thereby improvement in cardiac function in adult offspring of hypoxic pregnancy treated with melatonin with doses lower than those recommended for overcoming jet lag in humans. Melatonin treatment alone in healthy pregnancy did cause some alterations in cardiac structure. Therefore, maternal treatment with melatonin should only be given to pregnancies affected by chronic fetal hypoxia. [ABSTRACT FROM AUTHOR]

Cardiovascular disease such as myocardial infarction is one of the chief reasons for death universally. An extensive variety of natural and synthetic biodegradable polymers have been studied lately for cardiac tissue engineering applications. Cardiac tissue regeneration is a substitute approach for in situ cellular cardiomyoplasties, which is considered to renovate infarcted myocardium utilizing cells, scaffold, and growth factors. Polymer-based scaffolds including graphene and carbon nanotubes can be employed for heart tissue engineering. In this review, we consider the literature in terms of cardiac tissue engineering and discuss the most recent developments in terms of scaffold-based polymers for cardiac tissue regeneration. [ABSTRACT FROM AUTHOR]

Background: Cardiomyocyte senescence plays a crucial role as a pathological mechanism in heart failure (HF). However, the exact triggering factors and underlying causes of HF onset and progression are still not fully understood. Objectives: By integrating multi-omics data, this study aimed to determine the genetic associations between cardiomyocyte and HF using cell senescence-related genes (SRGs). Methods: The study utilized the CellAge database and the SenMayo dataset, combined with high-resolution single-cell RNA sequencing (scRNA-seq) data, to identify SRG and examine differences in cardiac cell expression. To explore the causal relationship with HF using Mendelian Randomization (MR). Genetic variations influencing gene expression, DNA methylation, and protein expression (cis-eQTL, cis-mQTL, and cis-pQTL) were analyzed using the two-sample MR (TSMR) and summary-data-based MR (SMR). Additionally, Bayesian colocalization analysis, germline genetic variation, and bulk RNA data were employed to strengthen the reliability of the results. The application potential of therapeutic targets is ultimately assessed by evaluating their druggability. Results: The expression of 39 SRGs in cardiomyocytes was identified. In the discovery set revealed that CDKN1A (OR = 1.09, 95% confidence interval (CI) 1.02–1.15, FDR = 0.048) could be causally related to HF, and the results are also replicated in the validation set (OR = 1.20, 95% confidence interval (CI) 1.10–1.30, FDR <0.0001). Based on the SMR method, CDKN1A was confirmed as a candidate pathogenic gene for HF, and its methylation (cg03714916, cg08179530) was associated with HF risk loci. The result is validated by Bayesian colocalization analysis, genetic variations, and bulk RNA data. The druggability analysis identified two potential therapeutic drugs. Conclusion: Based on multi-omics data, this study uncovered the reciprocal regulation of cardiomyocyte senescence through CDKN1 A, providing potential targets for HF drug development. [ABSTRACT FROM AUTHOR]

Cardiac regeneration in newborn rodents depends on the ability of pre-existing cardiomyocytes to proliferate and divide. This capacity is lost within the first week of postnatal development when these cells rapidly switch from hyperplasia to hypertrophy, withdraw from the cell cycle, become binucleated, and increase in size. How these dynamic changes in cell size and nucleation impact cardiomyocyte proliferative potential is not well understood. In this study, we innovate the application of a commercially available digital holographic imaging microscope, the Holomonitor M4, to evaluate the proliferative responses of mononucleated and binucleated cardiomyocytes after CHIR99021 treatment, a model proliferative stimulus. This system enables long-term label-free quantitative tracking of primary cardiomyocyte dynamics in real-time with single-cell resolution. Our results confirm that chemical inhibition of glycogen synthase kinase 3 with CHIR99021 promotes complete cell division of both mononucleated and binucleated cardiomyocytes with high frequency. Quantitative tracking of cardiomyocyte volume dynamics during these proliferative events revealed that both mononucleated and binucleated cardiomyocytes reach a similar size-increase threshold prior to attempted cell division. Binucleated cardiomyocytes attempt to divide with lower frequency than mononucleated cardiomyocytes, which may be associated with inadequate increases in cell size. By defining the interrelationship between cardiomyocyte size, nucleation, and cell cycle control, we may better understand the cellular mechanisms that drive the loss of mammalian cardiac regenerative capacity after birth. Digital holographic imaging enables quantitative label-free three-dimensional tracking of cardiomyocyte and non-cardiomyocyte dynamics in vitro. Cardiomyocytes reach a size-increase threshold prior to cell division, which may be impaired by binucleation. Created with BioRender.com. [Display omitted] • Digital holographic imaging enables quantitative label-free three-dimensional monitoring of primary cardiac cultures in vitro. • Primary cardiomyocytes can be readily distinguished from non-cardiomyocytes based on their larger size and slower motility. • Mononucleated and binucleated cardiomyocytes reach a common size-increase threshold prior to cell division. [ABSTRACT FROM AUTHOR]

L'augmentation de troponine hors contexte de syndrome coronarien aigu représente une réalité au quotidien et concerne 40 % des demandes de dosage des services d'urgences. Dans l'infarctus du myocarde (IDM), l'augmentation de la troponine cardiaque est liée à la nécrose des cardiomyocytes. Mais toute lésion des cardiomyocytes, réversible ou irréversible, peut s'accompagner d'une augmentation de troponine cardiaque. Il convient de savoir distinguer les causes sans réel impact délétère, comme l'exercice physique intense, des nombreuses causes pathologiques où la valeur élevée de troponine cardiaque constitue un facteur pronostique : insuffisance rénale, embolie pulmonaire, sepsis, chirurgie non cardiaque. La particularité de l'augmentation de la seule troponine T cardiaque dans différentes atteintes musculaires semble de plus en plus fréquente et amène à s'interroger sur la nécessité de disposer du dosage à la fois de troponine T cardiaque et de troponine I cardiaque. Apart from acute coronary syndrome, increased troponin represents 40 % of requests. In acute myocardial infarction (AMI), the increase in cardiac troponin is linked to cardiomyocyte necrosis. But any reversible or irreversible cardiomyocyte injury can be accompanied with an increase in cardiac troponin. It is important to know how to distinguish causes without any real deleterious -impact, such as intense physical exercise, from the numerous pathological causes where the high value of cardiac troponin constitutes a prognostic factor : renal failure, pulmonary embolism, sepsis, non-cardiac surgery. The increase in cardiac troponin T alone in various muscular disorders seems more and more frequent and leads us to question the necessity of having the dosage of both cardiac troponin T and cardiac troponin l. [ABSTRACT FROM AUTHOR]

Chemically defined, suspension culture conditions are a key requirement in realizing clinical translation of engineered cardiac tissues (ECTs). Building on our previous work producing functional ECT microspheres through differentiation of biomaterial encapsulated human induced pluripotent stem cells (hiPSCs), here we establish the ability to use chemically defined culture conditions, including stem cell media (E8) and cardiac differentiation media (chemically defined differentiation media with three components, CDM3). A custom microfluidic cell encapsulation system was used to encapsulate hiPSCs at a range of initial cell concentrations and diameters in the hybrid biomaterial, poly(ethylene glycol)‐fibrinogen (PF), for the formation of highly spherical and uniform ECT microspheres for subsequent cardiac differentiation. Initial microsphere diameter could be tightly controlled, and microspheres could be produced with an initial diameter between 400 and 800 µm. Three days after encapsulation, cardiac differentiation was initiated through small molecule modulation of Wnt signaling in CDM3. Cardiac differentiation occurred resulting in in situ ECT formation; results showed that this differentiation protocol could be used to achieve cardiomyocyte (CM) contents greater than 90%, although there was relatively high variability in CM content and yield between differentiation batches. Spontaneous contraction of ECT microspheres initiated between Days 7 and 10 of differentiation and ECT microspheres responded to electrical pacing up to 1.5 Hz. Resulting CMs had well‐defined sarcomeres and the gap junction protein, connexin 43, and had appropriate temporal changes in gene expression. In summary, this study demonstrated the proof‐of‐concept to produce functional ECT microspheres with chemically defined media in suspension culture in combination with biomaterial support of microsphere encapsulated hiPSCs. [ABSTRACT FROM AUTHOR]

Cardiotoxicity is a serious challenge cancer patients face today. Various factors are involved in cardiotoxicity. Circular RNAs (circRNAs) are one of the effective factors in the occurrence and prevention of cardiotoxicity. circRNAs can lead to increased proliferation, apoptosis, and regeneration of cardiomyocytes by regulating the molecular pathways, as well as increasing or decreasing gene expression; some circRNAs have a dual role in cardiomyocyte regeneration or death. Identifying each of the pathways related to these processes can be effective on managing patients and preventing cardiotoxicity. In this study, an overview of the molecular pathways involved in cardiotoxicity by circRNAs and their effects on the downstream factors have been discussed. [ABSTRACT FROM AUTHOR]

Dapagliflozin (DAPA), a sodium-glucose cotransporter 2 (SGLT2) inhibitor, is well-recognized for its therapeutic benefits in type 2 diabetes (T2D) and cardiovascular diseases. In this comprehensive in vitro study, we investigated DAPA's effects on cardiomyocytes, aortic endothelial cells (AECs), and stem cell-derived beta cells (SC-β), focusing on its impact on hypertrophy, inflammation, and cellular stress. Our results demonstrate that DAPA effectively attenuates isoproterenol (ISO)-induced hypertrophy in cardiomyocytes, reducing cell size and improving cellular structure. Mechanistically, DAPA mitigates reactive oxygen species (ROS) production and inflammation by activating the AKT pathway, which influences downstream markers of fibrosis, hypertrophy, and inflammation. Additionally, DAPA's modulation of SGLT2, the Na+/H + exchanger 1 (NHE1), and glucose transporter (GLUT 1) type 1 highlights its critical role in maintaining cellular ion balance and glucose metabolism, providing insights into its cardioprotective mechanisms. In aortic endothelial cells (AECs), DAPA exhibited notable anti-inflammatory properties by restoring AKT and phosphoinositide 3-kinase (PI3K) expression, enhancing mitogen-activated protein kinase (MAPK) activation, and downregulating inflammatory cytokines at both the gene and protein levels. Furthermore, DAPA alleviated tumor necrosis factor (TNFα)-induced inflammation and stress responses while enhancing endothelial nitric oxide synthase (eNOS) expression, suggesting its potential to preserve vascular function and improve endothelial health. Investigating SC-β cells, we found that DAPA enhances insulin functionality without altering cell identity, indicating potential benefits for diabetes management. DAPA also upregulated MAFA, PI3K, and NRF2 expression, positively influencing β-cell function and stress response. Additionally, it attenuated NLRP3 activation in inflammation and reduced NHE1 and glucose-regulated protein GRP78 expression, offering novel insights into its anti-inflammatory and stress-modulating effects. Overall, our findings elucidate the multifaceted therapeutic potential of DAPA across various cellular models, emphasizing its role in mitigating hypertrophy, inflammation, and cellular stress through the activation of the AKT pathway and other signaling cascades. These mechanisms may not only contribute to enhanced cardiac and endothelial function but also underscore DAPA's potential to address metabolic dysregulation in T2D. Key message: DAPA effectively attenuates ISO-induced cardiomyocyte hypertrophy by reducing cell size and improving cellular structure. DAPA exhibits anti-inflammatory properties in AECs by restoring AKT and PI3K expression, upregulating MAPK activation, and downregulating inflammatory gene expression. DAPA enhances insulin functionality in SC-β cells without altering cell identity, suggesting potential benefits in diabetes management. DAPA's modulation of SGLT2, NHE1, and GLUT1 expression in cardiomyocytes underscores its role in cellular ion balance and glucose metabolism, contributing to its cardioprotective mechanisms. DAPA alleviates TNFα-induced inflammation and stress responses in AECs, while enhancing eNOS expression, indicating its potential to preserve vascular function. DAPA attenuates NLRP3 activation and reduces NHE1 and GRP78 expression in SC-β cells, offering novel insights into its anti-inflammatory and stress-modulating effects. [ABSTRACT FROM AUTHOR]

Background: We aimed to investigate the effect and potential mechanism of enhancing Neuregulin1 (NRG1)/v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 4 (ErbB4) expression on the differentiation of induced pluripotent stem cells (iPSCs) into cardiomyocytes. Methods: We utilized CRISPR-CAS9 technology to knock in ErbB4 and obtained a single-cell clone IPSN-AAVS1-CMV-ErbB4 (iPSCs-ErbB4). Subsequently, we induced the differentiation of iPSCs into cardiomyocytes and quantified the number of beating embryoid bodies. Furthermore, quantitative real-time PCR assessed the expression of cardiomyocyte markers, including ANP (atrial natriuretic peptide), Nkx2.5 (NK2 transcription factor related locus 5), and GATA4 (GATA binding protein 4). On the 14th day of differentiation, we observed the α-MHC (α-myosin heavy chain)-positive area using immunofluorescent staining and conducted western blotting to detect the expression of cTnT (cardiac troponin) protein and PI3K/Akt signaling pathway-related proteins. Additionally, we intervened the iPSCs-ErbB4 + NRG1 group with the PI3K/Akt inhibitor LY294002 and observed alterations in the expression of cardiomyocyte differentiation-related genes. Results: The number of beating embryoid bodies increased after promoting the expression of NRG1/ErbB4 compared to the iPSCs control group. Cardiomyocyte markers ANP, Nkx2.5, and GATA4 significantly increased on day 14 of differentiation, and the positive area of α-MHC was three times that of the iPSCs control group. Moreover, there was a marked increase in cTnT protein expression. However, there was no significant difference in cardiomyocyte differentiation between the iPSCs-ErbB4 group and the iPSCs control group. Akt phosphorylation was significantly increased in the iPSCs-ErbB4 + NRG1 group. LY294002 significantly reversed the enhancing effect of NRG1/ErbB4 overexpression on Akt phosphorylation as well as the increase in α-MHC and cTnT expression. Conclusions: In conclusion, promoting the expression of NRG1/ErbB4 induced the differentiation of iPSC into cardiomyocytes, possibly through modulation of the PI3K/Akt signaling pathway. [ABSTRACT FROM AUTHOR]

Objective: To explore the role and regulation mechanism of leptin in senescence of mouse cardiomyocytes (MCM). Methods: The mRNA expression levels of senescence related indicators p16, p21, and senescence ⁃ associated secretory phenotype (SASP)in leptin stimulated MCM were examined by qPCR; the protein expressions of p16, p21, γ⁃H2AX, PI3K, AKT, p⁃PI3K, and p⁃AKT were detected by Western blot; the senescence of MCM was detected by β⁃galactosidase staining. PI3K inhibitor (LY294002) was pretreated for 2 h and then stimulated with leptin, the mRNA and protein levels of p16 and p21 were detected by qPCR and Western blot; the mRNA levels of SASP were examined by qPCR; MCM senescence was detected by β⁃galactosidase staining. Results: In MCM stimulated by leptin, the mRNA and protein levels of p16 and p21, as well as the protein level of γ⁃H2AX increased, the mRNA levels of SASP［ (interleukin, IL)⁃1β, tumor necrosis factor (TNF)⁃α、IL⁃6, monocyte chemoattractant protein (MCP)⁃1］were up ⁃regulated, the phosphorylation levels of proteins in PI3K/AKT signaling pathway increased; and β⁃galactosidase staining showed the senescence of MCM. When pretreated with PI3K inhibitor for 2 h, the mRNA and protein levels of p16 and p21, as well as the protein level of γ ⁃ H2AX were down ⁃ regulated, and the expressions of SASP mRNA were down ⁃ regulated, the senescence of MCM was alleviated. Conclusion: Leptin regulates the progression of MCM senescence by activating PI3K/AKT signaling pathway and promoting SASP (IL⁃1β, TNF⁃α, IL⁃6 and MCP⁃1)secretion. [ABSTRACT FROM AUTHOR]

The atrioventricular node (AVN) is a crucial component of the cardiac conduction system. Despite its pivotal role in regulating the transmission of electrical signals between atria and ventricles, a comprehensive understanding of the cellular electrophysiological mechanisms governing AVN function has remained elusive. This paper presents a detailed computational model of mouse AVN cell action potential (AP). Our model builds upon previous work and introduces several key refinements, including accurate representation of membrane currents and exchangers, calcium handling, cellular compartmentalization, dynamic update of intracellular ion concentrations, and calcium buffering. We recalibrated and validated the model against existing and unpublished experimental data. In control conditions, our model reproduces the AVN AP experimental features, (e.g. rate = 175 bpm, experimental range [121, 191] bpm). Notably, our study sheds light on the contribution of L‐type calcium currents, through both Cav1.2 and Cav1.3 channels, in AVN cells. The model replicates several experimental observations, including the cessation of firing upon block of Cav1.3 or INa,r current. If block induces a reduction in beating rate of 11%. In summary, this work presents a comprehensive computational model of mouse AVN cell AP, offering a valuable tool for investigating pacemaking mechanisms and simulating the impact of ionic current blockades. By integrating calcium handling and refining formulation of ionic currents, our model advances understanding of this critical component of the cardiac conduction system, providing a platform for future developments in cardiac electrophysiology. Key points: This paper introduces a comprehensive computational model of mouse atrioventricular node (AVN) cell action potentials (APs).Our model is based on the electrophysiological data from isolated mouse AVN cells and exhibits an action potential and calcium transient that closely match the experimental records.By simulating the effects of blocking specific ionic currents, the model effectively predicts the roles of L‐type Cav1.2 and Cav1.3 channels, T‐type calcium channels, sodium currents (TTX‐sensitive and TTX‐resistant), and the funny current (If) in AVN pacemaking.The study also emphasizes the significance of other ionic currents, including IKr, Ito, IKur, in regulating AP characteristics and cycle length in AVN cells.The model faithfully reproduces the rate dependence of action potentials under pacing, opening the possibility of use in impulse propagation models.The population‐of‐models approach showed the robustness of this new AP model in simulating a wide spectrum of cellular pacemaking in AVN. [ABSTRACT FROM AUTHOR]

S‐Limonene (s‐Lim) is a monocyclic monoterpene found in a variety of plants and has been shown to present antioxidant and cardioprotective activity in experimental models of myocardial infarction. The aim of this study was to evaluate the potential mechanism by which s‐Lim exerts its antiarrhythmic effect, focusing on the blockade of β‐adrenoceptor (β‐AR) and its effects on various in vivo and in vitro parameters, including electrocardiogram (ECG) measurements, left ventricular developed pressure (LVDP), the β‐adrenergic pathway, sarcomeric shortening and L‐type calcium current (ICa,L). In isolated hearts, 10 μM of s‐Lim did not alter the ECG profile or LVPD. s‐Lim increased the heart rate corrected QT interval (QTc) (10.8%) at 50 μM and reduced heart rate at the concentrations of 30 (12.4%) and 50 μM (16.6%). s‐Lim (10 μM) also inhibited the adrenergic response evoked by isoproterenol (ISO) (1 μM) reducing the increased of heart rate, LVDP and ECG changes. In ventricular cardiomyocyte, s‐Lim antagonized the effect of dobutamine by preventing the increase of sarcomeric shortening, demonstrating a similar effect to atenolol (blocker β1‐AR). In vivo, s‐Lim antagonized the effect of ISO (agonists β1‐AR), presenting a similar effect to propranolol (a non‐selective blocker β‐AR). In ventricular cardiomyocyte, s‐Lim did not alter the voltage dependence for ICa,L activation or the ICa,L density. In addition, s‐Lim did not affect changes in the ECG effect mediated by 5 μM forskolin (an activator of adenylate cyclase). In an in vivo caffeine/ISO‐induced arrhythmia model, s‐Lim (1 mg/kg) presented antiarrhythmic action verified by a reduced arrhythmia score, heart rate, and occurrence of ventricular premature beats and inappropriate sinus tachycardia. These findings indicate that the antiarrhythmic activity of s‐Lim is related to blockade of β‐AR in the heart. [ABSTRACT FROM AUTHOR]

Mechanical load is a potent regulator of cardiac structure and function. Although high workload during heart failure is associated with disruption of cardiomyocyte t‐tubules and Ca2+ homeostasis, it remains unclear whether changes in preload and afterload may promote adaptive t‐tubule remodelling. We examined this issue by first investigating isolated effects of stepwise increases in load in cultured rat papillary muscles. Both preload and afterload increases produced a biphasic response, with the highest t‐tubule densities observed at moderate loads, whereas excessively low and high loads resulted in low t‐tubule levels. To determine the baseline position of the heart on this bell‐shaped curve, mice were subjected to mildly elevated preload or afterload (1 week of aortic shunt or banding). Both interventions resulted in compensated cardiac function linked to increased t‐tubule density, consistent with ascension up the rising limb of the curve. Similar t‐tubule proliferation was observed in human patients with moderately increased preload or afterload (mitral valve regurgitation, aortic stenosis). T‐tubule growth was associated with larger Ca2+ transients, linked to upregulation of L‐type Ca2+ channels, Na+–Ca2+ exchanger, mechanosensors and regulators of t‐tubule structure. By contrast, marked elevation of cardiac load in rodents and patients advanced the heart down the declining limb of the t‐tubule–load relationship. This bell‐shaped relationship was lost in the absence of electrical stimulation, indicating a key role of systolic stress in controlling t‐tubule plasticity. In conclusion, modest augmentation of workload promotes compensatory increases in t‐tubule density and Ca2+ cycling, whereas this adaptation is reversed in overloaded hearts during heart failure progression. Key points: Excised papillary muscle experiments demonstrated a bell‐shaped relationship between cardiomyocyte t‐tubule density and workload (preload or afterload), which was only present when muscles were electrically stimulated.The in vivo heart at baseline is positioned on the rising phase of this curve because moderate increases in preload (mice with brief aortic shunt surgery, patients with mitral valve regurgitation) resulted in t‐tubule growth. Moderate increases in afterload (mice and patients with mild aortic banding/stenosis) similarly increased t‐tubule density.T‐tubule proliferation was associated with larger Ca2+ transients, with upregulation of the L‐type Ca2+ channel, Na+–Ca2+ exchanger, mechanosensors and regulators of t‐tubule structure.By contrast, marked elevation of cardiac load in rodents and patients placed the heart on the declining phase of the t‐tubule–load relationship, promoting heart failure progression.The dependence of t‐tubule structure on preload and afterload thus enables both compensatory and maladaptive remodelling, in rodents and humans. [ABSTRACT FROM AUTHOR]

The transient receptor potential melastatin 4 (TRPM4) channel contributes extensively to cardiac electrical activity, especially cardiomyocyte action potential formation. Mechanical stretch can induce changes in heart rate and rhythm, and the mechanosensitive channel Piezo1 is expressed in many cell types within the myocardium. Our previous study showed that TRPM4 and Piezo1 are closely co‐localized in the t‐tubules of ventricular cardiomyocytes and contribute to the Ca2+‐dependent signalling cascade that underlies hypertrophy in response to mechanical pressure overload. However, there was no direct evidence showing that Piezo1 activation was related to TRPM4 activation in situ. In the present study, we employed the HL‐1 mouse atrial myocyte‐like cell line as an in vitro model to investigate whether Piezo1–TRPM4 coupling can affect action potential properties. We used the small molecule Piezo1 agonist, Yoda1, as a surrogate for mechanical stretch to activate Piezo1 and detected the action potential changes in HL‐1 cells using FluoVolt, a fluorescent voltage sensitive dye. Our results demonstrate that Yoda1‐induced activation of Piezo1 changes the action potential frequency in HL‐1 cells. This change in action potential frequency is reduced by Piezo1 knockdown using small intefering RNA. Importantly knockdown or pharmacological inhibition of TRPM4 significantly affected the degree to which Yoda1‐evoked Piezo1 activation influenced action potential frequency. Thus, the present study provides in vitro evidence of a functional coupling between Piezo1 and TRPM4 in a cardiomyocyte‐like cell line. The coupling of a mechanosensitive Ca2+ permeable channel and a Ca2+‐activated TRP channel probably represents a ubiquitous model for the role of TRP channels in mechanosensory transduction. Key points: The transient receptor potential melastatin 4 (TRPM4) and Piezo1 channels have been confirmed to contribute to the Ca2+‐dependent signalling cascade that underlies cardiac hypertrophy in response to mechanical pressure overload. However, there was no direct evidence showing that Piezo1 activation was related to TRPM4 activation in situ.We employed the HL‐1 mouse atrial myocyte‐like cell line as an in vitro model to investigate the effect of Piezo1–TRPM4 coupling on cardiac electrical properties.The results show that both pharmacological and genetic inhibition of TRPM4 significantly affected the degree to which Piezo1 activation influenced action potential frequency in HL‐1 cells.Our findings provide in vitro evidence of a functional coupling between Piezo1 and TRPM4 in a cardiomyocyte‐like cell line.The coupling of a mechanosensitive Ca2+ permeable channel and a Ca2+‐activated TRP channel probably represents a ubiquitous model for the role of TRP channels in mechanosensory transduction in various (patho)physiological processes. [ABSTRACT FROM AUTHOR]

Myocardial infarction (MI) triggers a poor ventricular remodeling response, but the underlying mechanisms remain unclear. Here, the authors show that sentrin‐specific protease 1 (SENP1) is downregulated in post‐MI mice and in patients with severe heart failure. By generating cardiomyocyte‐specific SENP1 knockout and overexpression mice to assess cardiac function and ventricular remodeling responses under physiological and pathological conditions. Increased cardiac fibrosis in the cardiomyocyte‐specific SENP1 deletion mice, associated with increased fibronectin (Fn) expression and secretion in cardiomyocytes, promotes fibroblast activation in response to myocardial injury. Mechanistically, SENP1 deletion in mouse cardiomyocytes increases heat shock protein 90 alpha family class B member 1 (HSP90ab1) SUMOylation with (STAT3) activation and Fn secretion after ventricular remodeling initiated. Overexpression of SENP1 or mutation of the HSP90ab1 Lys72 ameliorates adverse ventricular remodeling and dysfunction after MI. Taken together, this study identifies SENP1 as a positive regulator of cardiac repair and a potential drug target for the treatment of MI. Inhibition of HSP90ab1 SUMOylation stabilizes STAT3 to inhibit the adverse ventricular remodeling response. [ABSTRACT FROM AUTHOR]

Epidemiology has shown a strong relationship between fine particulate matter (PM) exposure and cardiovascular disease. However, it remains unknown whether PM aggravates myocardial ischemia-reperfusion (I/R) injury, and the related mechanisms are unclear. Our previous study has shown that adipose stem cell-derived exosomes (ADSC-Exos) contain high levels of Mir221 and Mir222. The present study investigated the effects of PM exposure on I/R-induced cardiac injury through mitophagy and apoptosis, as well as the potential role of Mir221 and Mir222 in ADSC-Exos. Wild-type, mir221- and mir222-knockout (KO), and Mir221- and Mir222-overexpressing transgenic (TG) mice were intratracheally injected with PM (10 mg/kg). After 24 h, mice underwent left coronary artery ligation for 30 min, followed by 3 h of reperfusion (I/R). H9c2 cardiomyocytes were cultured under 1% O2 for 6 h, then reoxygenated for 12 h (hypoxia-reoxygenation [H/R]). PM aggravated I/R (or H/R) cardiac injury by increasing ROS levels and causing mitochondrial dysfunction, which increased the expression of mitochondrial fission-related proteins (DNM1L/Drp1 and MFF) and mitophagy-related proteins (BNIP3 and MAP1LC3B/LC3B) in vivo and in vitro. Treatment with ADSC-Exos or Mir221- and Mir222-mimics significantly reduced PM+I/R-induced cardiac injury. Importantly, ADSC-Exos contain Mir221 and Mir222, which directly targets BNIP3, MAP1LC3B/LC3B, and BBC3/PUMA, decreasing their expression and ultimately reducing cardiomyocyte mitophagy and apoptosis. The present data showed that ADSC-Exos treatment regulated mitophagy and apoptosis through the Mir221 and Mir222-BNIP3-MAP1LC3B-BBC3/PUMA pathway and significantly reduced the cardiac damage caused by PM+I/R. The present study revealed the novel therapeutic potential of ADSC-Exos in alleviating PM-induced exacerbation of myocardial I/R injury.Abbreviation: ADSC-Exos: adipose-derived stem cell exosomes; AL: autolysosome; ATP: adenosine triphosphate; BBC3/PUMA: BCL2 binding component 3; BNIP3: BCL2/adenovirus E1B interacting protein 3; CASP3: caspase 3; CASP9: caspase 9; CDKN1B/p27: cyclin dependent kinase inhibitor 1B; CVD: cardiovascular disease; DCFH-DA: 2‘,7’-dichlorodihydrofluorescein diacetate; DHE: dihydroethidium; DNM1L/Drp1: dynamin 1-like; EF: ejection fraction; FS: fractional shortening; H/R: hypoxia-reoxygenation; I/R: ischemia-reperfusion; LDH: lactate dehydrogenase; MAP1LC3B/LC3B: microtubule-associated protein 1 light chain 3 beta; MFF: mitochondrial fission factor; miRNA: microRNA; NAC: N-acetylcysteine; OCR: oxygen consumption rate; PIK3C3/Vps34: phosphatidylinositol 3-kinase catalytic subunit type 3; PM: particulate matter; PRKAA1/AMPK: protein kinase AMP-activated catalytic subunit alpha 1; ROS: reactive oxygen species; SQSTM1/p62: sequestosome 1; TEM: transmission electron microscopy; TRP53/p53: transformation related protein 53; TUNEL: terminal deoxynucleotidyl transferase dUTP nick end labeling. [ABSTRACT FROM AUTHOR]

BACKGROUND: Inherited heart disease has a high prevalence and mortality rate, but its pathogenesis has not yet been clarified. Although relevant animal models have been established to provide a foundation for the pathogenesis research of inherited heart disease, the value of these research results has been significantly reduced due to differences among species. Therefore, a new model is needed to explore its occurrence and development. OBJECTIVE: To review the current role of induced pluripotent stem cells in disease modeling and potential application prospects in various inherited heart diseases. METHODS: The first author searched the relevant articles published nearly 13 years in PubMed from January to March 2023. The search terms were “induced pluripotent stem cell, inherited heart disease, congenital heart disease”. Finally, 76 articles were included for analysis. RESULTS AND CONCLUSION: Since 2007, when induced pluripotent stem cells were induced from human somatic cells, many studies have been reported on disease-specific induced pluripotent stem cells. Due to the ability of disease-specific induced pluripotent stem cells to reproduce disease phenotypes, they are expected to become a new research tool for in vitro disease modeling, used to analyze pathogenesis and develop auxiliary drugs. In the research of cardiovascular genetic diseases, cardiomyocytes derived from patient-specific induced pluripotent stem cells contain gene mutations that are involved in cardiac dysplasia. Therefore, it can be used as a new tool to study the potential mechanisms of inherited heart disease. Up to now, induced pluripotent stem cells-derived cardiomyocytes have been widely used to study the molecular mechanisms of various genetic heart diseases, such as cardiac electrophysiological diseases, cardiomyopathy, and some syndromic inherited heart diseases. [ABSTRACT FROM AUTHOR]

BACKGROUND: HIF (hypoxia inducible factor) regulates many aspects of cardiac function. We and others previously showed that chronic HIF activation in the heart in mouse models phenocopies multiple features of ischemic cardiomyopathy in humans, including mitochondrial loss, lipid accumulation, and systolic cardiac dysfunction. In some settings, HIF also causes the loss of peroxisomes. How, mechanistically, HIF promotes cardiac dysfunction is an open question. METHODS: We used mice lacking cardiac pVHL (von Hippel-Lindau protein) to investigate how chronic HIF activation causes multiple features of ischemic cardiomyopathy, such as autophagy induction and lipid accumulation. We performed immunoblot assays, RNA sequencing, mitochondrial and peroxisomal autophagy flux measurements, and live cell imaging on isolated cardiomyocytes. We used CRISPR-Cas9 gene editing in mice to validate a novel mediator of cardiac dysfunction in the setting of chronic HIF activation. RESULTS: We identify a previously unknown pathway by which cardiac HIF activation promotes the loss of mitochondria and peroxisomes. We found that DEPP1 (decidual protein induced by progesterone 1) is induced under hypoxia in a HIF-dependent manner and localizes inside mitochondria. DEPP1 is both necessary and sufficient for hypoxia-induced autophagy and triglyceride accumulation in cardiomyocytes ex vivo. DEPP1 loss increases cardiomyocyte survival in the setting of chronic HIF activation ex vivo, and whole-body Depp1 loss decreases cardiac dysfunction in hearts with chronic HIF activation caused by VHL loss in vivo. CONCLUSIONS: Our findings identify DEPP1 as a key component in the cardiac remodeling that occurs with chronic ischemia. [ABSTRACT FROM AUTHOR]

Post-translational SUMOylation of nuclear and cytosolic proteins maintains homeostasis in eukaryotic cells and orchestrates programmed responses to changes in metabolic demand or extracellular stimuli. In excitable cells, SUMOylation tunes the biophysical properties and trafficking of ion channels. Ion channel SUMOylation status is determined by the opposing enzyme activities of SUMO ligases and deconjugases. Phosphorylation also plays a permissive role in SUMOylation. SUMO deconjugases have been identified for several ion channels, but their corresponding E3 ligases remain unknown. This study shows PIAS3, a.k.a. KChAP, is a bona fide SUMO E3 ligase for Kv4.2 and HCN2 channels in HEK cells, and endogenous Kv4.2 and Kv4.3 channels in cardiomyocytes. PIAS3-mediated SUMOylation at Kv4.2-K579 increases channel surface expression through a rab11a-dependent recycling mechanism. PKA phosphorylation at Kv4.2-S552 reduces the current mediated by Kv4 channels in HEK293 cells, cardiomyocytes, and neurons. This study shows PKA mediated phosphorylation blocks Kv4.2-K579 SUMOylation in HEK cells and cardiomyocytes. Together, these data identify PIAS3 as a key downstream mediator in signaling cascades that control ion channel surface expression. [ABSTRACT FROM AUTHOR]

Objective To explore the possible mechanism of apoptosis induced by SARS-CoV-2 targeting cyclophilin A(CypA)/CD147 receptor pathway. Methods The pEncmv-SARS-CoV-2 S-3xflag recombinant plasmid was constructed and transfected into H9C2 cells. Apoptosis was analyzed by TUNEL assay, flow cytometry and DNA ladder. The interaction between CD147 and SARS-CoV-2 S protein was verified by CoIP. The expressions of apoptosis-related proteins were detected by Western blotting. Results Compared with the control group, TUNEL assay and flow cytometry showed that apoptosis was significantly increased in transfection group (P<0.05), and DNA ladder analysis showed that DNA degradation was obvious. The expression levels of Caspase12 and DR3 were significantly increased (P<0.001), the expression level of FAS was slightly increased(P<0.01), while the expression levels of TRF1 and DR4 were decreased (P<0.01). Conclusion SARS-CoV-2 S protein targets CypA/CD147 receptor to invade host cells, activate endoplasmic network pathway (Caspase12) and death receptor pathway(DR3, FAS), and induce apoptosis of cardiomyocytes through endogenous and exogenous apoptotic pathways. [ABSTRACT FROM AUTHOR]

The aging heart is characterized by a number of structural changes leading to ventricular stiffness, impaired resistance to stress and increased risk of developing heart failure (HF). Genetic or pharmacological removal of senescent cells has recently demonstrated the possibility to relieve some cardiac aging features such as hypertrophy and fibrosis. However, the contribution of the different cell types in cardiac aging remains fragmentary due to a lack of cell‐specific markers. Cardiomyocytes undergo post‐mitotic senescence in response to telomere damage, characterized by persistent DNA damage response and expression of the classical senescence markers p21 and p16, which are shared by many other cell types. In the present study, we used transcriptomic approaches to discover new markers specific for cardiomyocyte senescence. We identified Prominin2 (Prom2), encoding a transmembrane glycoprotein, as the most upregulated gene in cardiomyocytes of aged mice compared to young mice. We showed that Prom2 was upregulated by a p53‐dependent pathway in stress‐induced premature senescence. Prom2 expression correlated with cardiomyocyte hypertrophy in the hearts of aged mice and was increased in atrial samples of patients with HF with preserved ejection fraction. Consistently, Prom2 overexpression was sufficient to drive senescence, hypertrophy and resistance to cytotoxic stress while Prom2 shRNA silencing inhibited these features in doxorubicin‐treated cardiac cells. In conclusion, we identified Prom2 as a new player of cardiac aging, linking cardiomyocyte hypertrophy to senescence. These results could provide a better understanding and targeting of cell‐type specific senescence in age‐associated cardiac diseases. [ABSTRACT FROM AUTHOR]

Objective: To investigate the effect of wogonin (WOG) on hypoxia/reoxygenation (H/R)-induced cardiomyocyte apoptosis by regulating adenosine monophosphate-activated protein kinase (AMPK)/NOD-like receptor protein 3 (NLRP3) signaling pathway. Methods: H9C2 cells were divided into Control group (normal culture), H/R group (H/R induction), L (low dose)-WOG group, M (medium dose)-WOG group, H (high dose)-WOG group (40, 80, 120 µmol/L WOG were added on the basis of H/R induction), WOG+Compound C group (10 µmol/L AMPK inhibitor Compound C was added on the basis of H-WOG group). The effect of WOG on the proliferation of H9C2 cells was detected by methyl thiazolyl tetrazolium (MTT) assay and 5-ethynyl-2'-deoxyuridine (EdU) staining. The effect of WOG on apoptosis of H9C2 cells was detected by flow cytometry. The levels of serum oxidative stress indexes [malondialdehyde (MDA), reactive oxygen species (ROS), superoxide dismutase (SOD)] and inflammatory factors [interleukin (IL)-1B, IL-18] in H9C2 cells were detected by enzyme-linked immunosorbent assay (ELISA). The expression of AMPK, NLRP3, B cell lymphoma-2 (Bcl-2) and Bcl-2 associated X protein (Bax) in H9C2 cells was detected by Western blot (WB). Results: The optical density (OD), EdU positive cell rate, SOD, AMPK and Bcl-2 of H9C2 cells in H/R group were lower than those in Control group, and the apoptosis rate, IL-1B, IL-18, ROS, MDA, NLRP3 and Bax were higher than those in Control group (P<0.05). Compared with H/R group, OD490, EdU positive cell rate, SOD, AMPK and Bcl-2 expression in L-WOG group, M-WOG group and H-WOG group increased, while apoptosis rate, IL-1B, IL-18, ROS, MDA, NLRP3 and Bax decreased (P<0.05). The OD, EdU positive cell rate, SOD, AMPK and Bcl-2 in WOG+Compound C group were lower than those in H-WOG group, and the apoptosis rate, IL-1B, IL-18, ROS, MDA, NLRP3 and Bax expression were higher than those in H-WOG group (P<0.05). Conclusion: WOG can inhibit H/R-induced cardiomyocyte apoptosis, which may be relate to the AMPK/NLRP3 signaling pathway. [ABSTRACT FROM AUTHOR]

Cardiovascular manifestations of coronavirus disease 2019 (COVID-19) include myocardial injury, heart failure, and myocarditis and are associated with long-term disability and mortality. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA and antigens are found in the myocardium of COVID-19 patients, and human cardiomyocytes are susceptible to infection in cell or organoid cultures. While these observations raise the possibility that cardiomyocyte infection may contribute to the cardiac sequelae of COVID-19, a causal relationship between cardiomyocyte infection and myocardial dysfunction and pathology has not been established. Here, we generated a mouse model of cardiomyocyte-restricted infection by selectively expressing human angiotensin-converting enzyme 2 (hACE2), the SARS-CoV-2 receptor, in cardiomyocytes. Inoculation of Myh6-Cre Rosa26loxP-STOP-loxP-hACE2 mice with an ancestral, non-mouse-adapted strain of SARS-CoV-2 resulted in viral replication within the heart, accumulation of macrophages, and moderate left ventricular (LV) systolic dysfunction. Cardiac pathology in this model was transient and resolved with viral clearance. Blockade of monocyte trafficking reduced macrophage accumulation, suppressed the development of LV systolic dysfunction, and promoted viral clearance in the heart. These findings establish a mouse model of SARS-CoV-2 cardiomyocyte infection that recapitulates features of cardiac dysfunctions of COVID-19 and suggests that both viral replication and resultant innate immune responses contribute to cardiac pathology. IMPORTANCE Heart involvement after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection occurs in multiple ways and is associated with worse outcomes in coronavirus disease 2019 (COVID-19) patients. It remains unclear if cardiac disease is driven by primary infection of the heart or immune response to the virus. SARS-CoV-2 is capable of entering contractile cells of the heart in a culture dish. However, it remains unclear how such infection affects the function of the heart in the body. Here, we designed a mouse in which only heart muscle cells can be infected with a SARS-CoV-2 strain to study cardiac infection in isolation from other organ systems. In our model, infected mice show viral infection, worse function, and accumulation of immune cells in the heart. A subset of immune cells facilitates such worsening heart function. As this model shows features similar to those observed in patients, it may be useful for understanding the heart disease that occurs as a part of COVID-19. [ABSTRACT FROM AUTHOR]

One of the many unresolved obstacles in the field of cardiovascular research is an uncompromising in vitro cardiac model. While primary cell sources from animal models offer both advantages and disadvantages, efforts over the past half-century have aimed to reduce their use. Additionally, obtaining a sufficient quantity of human primary cardiomyocytes faces ethical and legal challenges. As the practically unlimited source of human cardiomyocytes from induced pluripotent stem cells (hiPSC-CM) is now mostly resolved, there are great efforts to improve their quality and applicability by overcoming their intrinsic limitations. The greatest bottleneck in the field is the in vitro ageing of hiPSC-CMs to reach a maturity status that closely resembles that of the adult heart, thereby allowing for more appropriate drug developmental procedures as there is a clear correlation between ageing and developing cardiovascular diseases. Here, we review the current state-of-the-art techniques in the most realistic heart models used in disease modelling and toxicity evaluations from hiPSC-CM maturation through heart-on-a-chip platforms and in silico models to the in vitro models of certain cardiovascular diseases. [ABSTRACT FROM AUTHOR]

The discovery of human pluripotent stem cells (hiPSCs) and advances in DNA editing techniques have opened opportunities for personalized cell-based therapies for a wide spectrum of diseases. It has gained importance as a valuable tool to investigate genetic and functional variations in congenital heart defects (CHDs), enabling the customization of treatment strategies. The ability to understand the disease process specific to the individual patient of interest provides this technology with a significant advantage over generic animal models. However, its utility as a disease-in-a-dish model requires identifying effective and efficient differentiation protocols that accurately reproduce disease traits. Currently, iPSC-related research relies heavily on the quality of cells and the properties of the differentiation technique In this review, we discuss the utility of iPSCs in bench CHD research, the molecular pathways involved in the differentiation of cardiomyocytes, and their applications in CHD disease modeling, therapeutics, and drug application. [ABSTRACT FROM AUTHOR]

Pulmonary artery hypertension (PAH) is characterised by increased pulmonary vascular resistance (PVR) resulting in elevated pressure in the pulmonary artery supplying the pulmonary circulation. Disease of the right ventricle (RV) often manifests as a result of PAH placing excessive pressure on the right side of the heart. Although a relatively rare disease in humans, the impact of sustained PAH is severe, with poor outcomes even in treated individuals. As PAH develops, the blood flow is restricted through the pulmonary arteries and the right ventricle hypertrophies due to the increased strain of pumping blood through the pulmonary circulation. With time, RV hypertrophy progresses to right heart failure, impacting the supply of blood to the left ventricle and systemic circulation. Although right heart failure can currently be treated, it cannot be cured. There is therefore a need for more research into the physiological changes that cause the heart to fail under pressure overload. This review aims to evaluate the monocrotaline (MCT) rat model of PAH as a means of studying the cellular mechanisms associated with the development of RV hypertrophy and right heart failure. [ABSTRACT FROM AUTHOR]

Context: The prevalence of obesity was 8% in 2002 and increased to 11.5% in 2011 in Indonesia. The results of the 2018 Basic Health Research stated that 13.6% of people over the age of 18 were overweight, and 21.8% were obese. Aims: To analyze the effect of intermittent restriction diet and continuous exercise with moderate intensity on individuals exposed to a high-calorie diet. Methods: A randomized posttest control group design was used as the research design, with 6 mice in each group as the study subjects. The groups in this study were the control group with a high-calorie diet (CON+), the intermittent restriction diet group (IRD), the moderate-intensity continuous exercise group (MEX), the combined intermittent restriction diet group and the moderate-intensity continuous exercise (HYB) group. The measured variables were cardiomyocyte diameter and brain natriuretic peptide (BNP) expression. Results: The results of this study were cardiomyocyte diameter CON+ (27.94 ± 5.65 µm), IRD (20.99 ± 11.80 µm), MEX (25.08 ± 9.14 µm), HYB (24.52 ± 5.90 µm) with p=0.578. Expression scores of BNP CON+ (6.00 ± 1.50), IRD (4.67 ± 1.00), MEX (5.42±2.09), HYB (6.27 ± 1.54) with p=0.335. These results showed no significant difference, but the intermittent restriction diet showed the most optimal mean with the lowest mean cardiomyocyte diameter and BNP expression. Conclusions: There was no effect between the combination of an intermittent restriction diet and moderate-intensity continuous exercise on cardiomyocyte diameter and BNP expression. However, there is a potential that an intermittent restriction diet has the optimal effect in preventing changes in the cardiac structure.

Abstract Abstract Dapagliflozin (DAPA), a sodium-glucose cotransporter 2 (SGLT2) inhibitor, is well-recognized for its therapeutic benefits in type 2 diabetes (T2D) and cardiovascular diseases. In this comprehensive in vitro study, we investigated DAPA’s effects on cardiomyocytes, aortic endothelial cells (AECs), and stem cell-derived beta cells (SC-β), focusing on its impact on hypertrophy, inflammation, and cellular stress. Our results demonstrate that DAPA effectively attenuates isoproterenol (ISO)-induced hypertrophy in cardiomyocytes, reducing cell size and improving cellular structure. Mechanistically, DAPA mitigates reactive oxygen species (ROS) production and inflammation by activating the AKT pathway, which influences downstream markers of fibrosis, hypertrophy, and inflammation. Additionally, DAPA’s modulation of SGLT2, the Na+/H + exchanger 1 (NHE1), and glucose transporter (GLUT 1) type 1 highlights its critical role in maintaining cellular ion balance and glucose metabolism, providing insights into its cardioprotective mechanisms. In aortic endothelial cells (AECs), DAPA exhibited notable anti-inflammatory properties by restoring AKT and phosphoinositide 3-kinase (PI3K) expression, enhancing mitogen-activated protein kinase (MAPK) activation, and downregulating inflammatory cytokines at both the gene and protein levels. Furthermore, DAPA alleviated tumor necrosis factor (TNFα)-induced inflammation and stress responses while enhancing endothelial nitric oxide synthase (eNOS) expression, suggesting its potential to preserve vascular function and improve endothelial health. Investigating SC-β cells, we found that DAPA enhances insulin functionality without altering cell identity, indicating potential benefits for diabetes management. DAPA also upregulated MAFA, PI3K, and NRF2 expression, positively influencing β-cell function and stress response. Additionally, it attenuated NLRP3 activation in inflammation and reduced NHE1 and glucose-regulated protein GRP78 expression, offering novel insights into its anti-inflammatory and stress-modulating effects. Overall, our findings elucidate the multifaceted therapeutic potential of DAPA across various cellular models, emphasizing its role in mitigating hypertrophy, inflammation, and cellular stress through the activation of the AKT pathway and other signaling cascades. These mechanisms may not only contribute to enhanced cardiac and endothelial function but also underscore DAPA’s potential to address metabolic dysregulation in T2D. Graphical abstract

Abstract Background We aimed to investigate the effect and potential mechanism of enhancing Neuregulin1 (NRG1)/v-erb-b2 avian erythroblastic leukemia viral oncogene homolog 4 (ErbB4) expression on the differentiation of induced pluripotent stem cells (iPSCs) into cardiomyocytes. Methods We utilized CRISPR-CAS9 technology to knock in ErbB4 and obtained a single-cell clone IPSN-AAVS1-CMV-ErbB4 (iPSCs-ErbB4). Subsequently, we induced the differentiation of iPSCs into cardiomyocytes and quantified the number of beating embryoid bodies. Furthermore, quantitative real-time PCR assessed the expression of cardiomyocyte markers, including ANP (atrial natriuretic peptide), Nkx2.5 (NK2 transcription factor related locus 5), and GATA4 (GATA binding protein 4). On the 14th day of differentiation, we observed the α-MHC (α-myosin heavy chain)-positive area using immunofluorescent staining and conducted western blotting to detect the expression of cTnT (cardiac troponin) protein and PI3K/Akt signaling pathway-related proteins. Additionally, we intervened the iPSCs-ErbB4 + NRG1 group with the PI3K/Akt inhibitor LY294002 and observed alterations in the expression of cardiomyocyte differentiation-related genes. Results The number of beating embryoid bodies increased after promoting the expression of NRG1/ErbB4 compared to the iPSCs control group. Cardiomyocyte markers ANP, Nkx2.5, and GATA4 significantly increased on day 14 of differentiation, and the positive area of α-MHC was three times that of the iPSCs control group. Moreover, there was a marked increase in cTnT protein expression. However, there was no significant difference in cardiomyocyte differentiation between the iPSCs-ErbB4 group and the iPSCs control group. Akt phosphorylation was significantly increased in the iPSCs-ErbB4 + NRG1 group. LY294002 significantly reversed the enhancing effect of NRG1/ErbB4 overexpression on Akt phosphorylation as well as the increase in α-MHC and cTnT expression. Conclusions In conclusion, promoting the expression of NRG1/ErbB4 induced the differentiation of iPSC into cardiomyocytes, possibly through modulation of the PI3K/Akt signaling pathway.

Abstract Post-translational SUMOylation of nuclear and cytosolic proteins maintains homeostasis in eukaryotic cells and orchestrates programmed responses to changes in metabolic demand or extracellular stimuli. In excitable cells, SUMOylation tunes the biophysical properties and trafficking of ion channels. Ion channel SUMOylation status is determined by the opposing enzyme activities of SUMO ligases and deconjugases. Phosphorylation also plays a permissive role in SUMOylation. SUMO deconjugases have been identified for several ion channels, but their corresponding E3 ligases remain unknown. This study shows PIAS3, a.k.a. KChAP, is a bona fide SUMO E3 ligase for Kv4.2 and HCN2 channels in HEK cells, and endogenous Kv4.2 and Kv4.3 channels in cardiomyocytes. PIAS3-mediated SUMOylation at Kv4.2-K579 increases channel surface expression through a rab11a-dependent recycling mechanism. PKA phosphorylation at Kv4.2-S552 reduces the current mediated by Kv4 channels in HEK293 cells, cardiomyocytes, and neurons. This study shows PKA mediated phosphorylation blocks Kv4.2-K579 SUMOylation in HEK cells and cardiomyocytes. Together, these data identify PIAS3 as a key downstream mediator in signaling cascades that control ion channel surface expression.

BACKGROUND: Studies have shown that cardiomyocyte apoptosis is closely related to cardiac decompensation and the cardiac aging process. Appropriate exercise can alter heart pump function in patients with heart failure as well as attenuate aging-induced cardiomyocyte apoptosis, hypertrophy, and fibrotic damage. OBJECTIVE: To investigate the effects of long-term aerobic exercise on cardiomyocyte apoptosis and the thioredoxin system in aging rats. METHODS: Thirty-six male Sprague-Dawley rats were selected and divided into three age groups: 3-month-old young group, 9-month-old middle-aged group, and 18-month-old elderly group, with 12 rats in each group. Within each age group, rats were randomly assigned to sedentary and exercise subgroups (n=6 per group). The sedentary groups did not undergo any exercise intervention. The exercise groups were acclimated to a treadmill environment and subsequently subjected to treadmill exercise for 45 minutes per day, at a speed of 15 m/min, 5 days per week for 10 weeks in total. At 24 hours after the final intervention, ELISA was employed to measure serum levels of cardiac troponin I and creatine kinase-MB in rats. TUNEL assay was utilized to detect cardiomyocyte apoptosis, while western blot assay was employed to assess the protein expression of Bax, Bcl-2, Caspase 3, thioredoxin-1, thioredoxin-2, thioredoxin reductase-1, thioredoxin reductase-2, thioredoxin-interacting protein, apoptosis signal-regulating kinase 1, and P38 mitogen-activated protein kinase in rat myocardial tissue. RESULTS AND CONCLUSION: Serum levels of cardiac troponin I and creatine kinase-MB in the elderly sedentary group were significantly higher than those in the young and middle-aged sedentary groups and elderly exercise group (P < 0.01). Serum levels of cardiac troponin I and creatine kinase-MB in the elderly sedentary group were significantly higher than those in the young and middle-aged exercise groups and elderly exercise group (P < 0.01). Positive apoptotic cells in rat myocardial tissue, along with increased protein expression of Bax and Caspase 3, exhibited an age-related upward trend, while Bcl-2 protein expression showed a declining trend. In comparison with the sedentary groups within each age category, the number of apoptotic cardiomyocytes and the expression of Bax and Caspase 3 proteins were reduced to different degrees, and the expression of Bcl-2 protein was increased to different degrees in the corresponding exercise groups. Compared with the young sedentary group, middle-aged sedentary group and elderly exercise group, elderly sedentary rats showed a significant decrease in the expression of myocardial thioredoxin 1, thioredoxin 2, thioredoxin reductase 1, and thioredoxin reductase 2 proteins (P < 0.05, P < 0.01). The expression of myocardial thioredoxin 1, thioredoxin 2, and thioredoxin reductase 2 proteins was lower in the elderly exercise group than in the young exercise group (P < 0.05, P < 0.01), while the expression of thioredoxin reductase 1 and thioredoxin reductase 2 proteins was lower in the elderly exercise group than in the middle-aged exercise group (P < 0.01). The protein expression of thioredoxin-interacting protein, apoptosis signal-regulating kinase 1, and P38 mitogen-activated protein kinase in rat myocardium was significantly higher in the elderly sedentary group than the young sedentary group, middleaged sedentary group and elderly exercise group (P < 0.01). The protein expression of thioredoxin-interacting protein, apoptosis signal-regulating kinase 1, and P38 mitogen-activated protein kinase in rat myocardium was significantly higher in the elderly exercise group than the young exercise group and middleaged exercise group (P < 0.01). To conclude, aerobic exercise may enhance the anti-apoptotic effects of thioredoxin by down-regulating the expression of thioredoxin-interacting protein in aging rat hearts, leading to the downregulation of apoptosis signal-regulated kinase 1 and P38 mitogen-activated kinase protein, thereby alleviating myocardial cell apoptosis in aging rat hearts. [ABSTRACT FROM AUTHOR]

New approach methodologies (NAMs) for predicting embryotoxicity and developmental toxicity are urgently needed for generating human relevant data, while reducing turnover time and costs, and alleviating ethical concerns related to the use of animal models. We have previously developed the PluriLum assay, a NKX2.5-reporter gene 3D model using human-induced pluripotent stem cells (hiPSCs) that are genetically modified to enable the assessment of adverse effects of chemicals on the early-stage embryo. Aiming at improving the predictive value of the PluriLum assay for future screening purposes, we sought to introduce standardization steps to the protocol, improving the overall robustness of the PluriLum assay, as well as a shortening of the assay protocol. First, we showed that the initial size of embryoid bodies (EBs) is crucial for a proper differentiation into cardiomyocytes and overall reproducibility of the assay. When the starting diameter of the EBs exceeds 500 µm, robust differentiation can be anticipated. In terms of reproducibility, exposure to the fungicide epoxiconazole at smaller initial diameters resulted in a larger variation of the derived data, compared to more reliable concentration–response curves obtained using spheroids with larger initial diameters. We further investigated the ideal length of the differentiation protocol, resulting in a shortening of the PluriLum assay by 24 h to 7 days. Following exposure to the teratogens all-trans and 13-cis retinoic acid, both cardiomyocyte contraction and measurement of NKX2.5-derived luminescence were recorded with a similar or increased sensitivity after 6 days of differentiation when compared to the original 7 days. Finally, we have introduced an efficient step for enzymatic dissociation of the EBs at assay termination. This allows for an even splitting of the individual EBs and testing of additional endpoints other than the NKX2.5-luciferase reporter, which was demonstrated in this work by the simultaneous assessment of ATP levels. In conclusion, we have introduced standardizations and streamlined the PluriLum assay protocol to improve its suitability as a NAM for screening of a large number of chemicals for developmental toxicity testing. [ABSTRACT FROM AUTHOR]

Background. The purpose of the present study was to investigate the effect of aerobic-resistance training on the population of cardiac multipotent cells, thickness, and diameter of the ventricular through the C-Kit pathway of male rats during the growth stages. Methods. Overall, 30 male Wistar rats in three age groups of 2 (n = 10), 8 (n = 10), and 96 (n = 10) weeks were accidentally divided into exercise and control groups. Resistance training programs (resistance ladder, 3 days a week) and aerobics (treadmill running, 3 days a week) were performed for 6 weeks. Hematoxylin-Eosin staining was done the heart tissue and histological images were prepared. Afterward, C-Kit gene expression was examined by the real-time polymerase chain reaction method. The statistical method of one-way analysis of variance (P ≥ 0.05) and Tukey’s test were used at a significant level (P ≥ 0.01). Results. The trained neonatal group had higher values in heart weight to body weight index, and the trained neonatal and trained young adult groups had higher values in left ventricular thickness (P ≥ 0.01). The decrease in left ventricular internal diameter in the trained young adult group was significant compared to the control group (P ≥ 0.01). There was a significant increase in C-Kit gene expression in trained young adult and trained old groups (P ≥ 0.01). Conclusion. Aerobic-resistance training can be an effective stimulus for increasing the number of cardiac stem cells and creating structural adaptations of the heart, including increasing thickness and ventricular diameter in neonatal and young adult groups. Practical Implications. Participation in aerobic-resistance training programs leads to the improvement of the cardiac structure and an increase in cardiac stem cells.

Abstract The adult mammalian heart has extremely limited cardiac regenerative capacity. Most cardiomyocytes live in a state of permanent cell-cycle arrest and are unable to re-enter the cycle. Cardiomyocytes switch from cell proliferation to a maturation state during neonatal development. Although several signaling pathways are involved in this transition, the molecular mechanisms by which these inputs coordinately regulate cardiomyocyte maturation are not fully understood. Retinoic acid (RA) plays a pivotal role in development, morphogenesis, and regeneration. Despite the importance of RA signaling in embryo heart development, little is known about its function in the early postnatal period. We found that mRNA expression of aldehyde dehydrogenase 1 family member A2 (Aldh1a2), which encodes the key enzyme for synthesizing all-trans retinoic acid (ATRA) and is an important regulator for RA signaling, was transiently upregulated in neonatal mouse ventricles. Single-cell transcriptome analysis and immunohistochemistry revealed that Aldh1a2 expression was enriched in cardiac fibroblasts during the early postnatal period. Administration of ATRA inhibited cardiomyocyte proliferation in cultured neonatal rat cardiomyocytes and human cardiomyocytes. RNA-seq analysis indicated that cell proliferation-related genes were downregulated in prenatal rat ventricular cardiomyocytes treated with ATRA, while cardiomyocyte maturation-related genes were upregulated. These findings suggest that RA signaling derived from cardiac fibroblasts is one of the key regulators of cardiomyocyte proliferation and maturation during neonatal heart development.

Abstract Background Stem cell-derived extracellular vesicles (EVs) are an emerging class of therapeutics with excellent biocompatibility, bioactivity and pro-regenerative capacity. One of the potential targets for EV-based medicines are cardiovascular diseases (CVD). In this work we used EVs derived from human induced pluripotent stem cells (hiPSCs; hiPS-EVs) cultured under different oxygen concentrations (21, 5 and 3% O2) to dissect the molecular mechanisms responsible for cardioprotection. Methods EVs were isolated by ultrafiltration combined with size exclusion chromatography (UF + SEC), followed by characterization by nanoparticle tracking analysis, atomic force microscopy (AFM) and Western blot methods. Liquid chromatography and tandem mass spectrometry coupled with bioinformatic analyses were used to identify differentially enriched proteins in various oxygen conditions. We directly compared the cardioprotective effects of these EVs in an oxygen-glucose deprivation/reoxygenation (OGD/R) model of cardiomyocyte (CM) injury. Using advanced molecular biology, fluorescence microscopy, atomic force spectroscopy and bioinformatics techniques, we investigated intracellular signaling pathways involved in the regulation of cell survival, apoptosis and antioxidant response. The direct effect of EVs on NRF2-regulated signaling was evaluated in CMs following NRF2 inhibition with ML385. Results We demonstrate that hiPS-EVs derived from physiological hypoxia at 5% O2 (EV-H5) exert enhanced cytoprotective function towards damaged CMs compared to EVs derived from other tested oxygen conditions (normoxia; EV-N and hypoxia 3% O2; EV-H3). This resulted from higher phosphorylation rates of Akt kinase in the recipient cells after transfer, modulation of AMPK activity and reduced apoptosis. Furthermore, we provide direct evidence for improved calcium signaling and sustained contractility in CMs treated with EV-H5 using AFM measurements. Mechanistically, our mass spectrometry and bioinformatics analyses revealed differentially enriched proteins in EV-H5 associated with the antioxidant pathway regulated by NRF2. In this regard, EV-H5 increased the nuclear translocation of NRF2 protein and enhanced its transcription in CMs upon OGD/R. In contrast, inhibition of NRF2 with ML385 abolished the protective effect of EVs on CMs. Conclusions In this work, we demonstrate a superior cardioprotective function of EV-H5 compared to EV-N and EV-H3. Such EVs were most effective in restoring redox balance in stressed CMs, preserving their contractile function and preventing cell death. Our data support the potential use of hiPS-EVs derived from physiological hypoxia, as cell-free therapeutics with regenerative properties for the treatment of cardiac diseases.

Abstract Background Cardiomyocytes (CMs) derived from human induced pluripotent stem cells (hiPSCs) by traditional methods are a mix of atrial and ventricular CMs and many other non-cardiomyocyte cells. Retinoic acid (RA) plays an important role in regulation of the spatiotemporal development of the embryonic heart. Methods CMs were derived from hiPSC (hi-PCS-CM) using different concentrations of RA (Control without RA, LRA with 0.05μM and HRA with 0.1 μM) between day 3-6 of the differentiation process. Engineered heart tissues (EHTs) were generated by assembling hiPSC-CM at high cell density in a low collagen hydrogel. Results In the HRA group, hiPSC-CMs exhibited highest expression of contractile proteins MYH6, MYH7 and cTnT. The expression of TBX5, NKX2.5 and CORIN, which are marker genes for left ventricular CMs, was also the highest in the HRA group. In terms of EHT, the HRA group displayed the highest contraction force, the lowest beating frequency, and the highest sensitivity to hypoxia and isoprenaline, which means it was functionally more similar to the left ventricle. RNAsequencing revealed that the heightened contractility of EHT within the HRA group can be attributed to the promotion of augmented extracellular matrix strength by RA. Conclusion By interfering with the differentiation process of hiPSC with a specific concentration of RA at a specific time, we were able to successfully induce CMs and EHTs with a phenotype similar to that of the left ventricle or right ventricle.

Myocardial infarction results in extensive cardiomyocyte apoptosis, leading to the formation of noncontractile scar tissue. Given the limited regenerative capacity of adult mammalian cardiomyocytes, direct reprogramming of cardiac fibroblasts (CFs) into cardiomyocytes represents a promising therapeutic strategy for myocardial repair, and small molecule drugs might offer a more attractive alternative to gene editing approaches in terms of safety and clinical feasibility. This study aimed to reprogram rat CFs into cardiomyocytes using a small molecular chemical mixture comprising CHIR99021, Valproic acid, Dorsomorphin, SB431542, and Forskolin. Immunofluorescence analysis revealed a significant increase in the expression of cardiomyocyte‐specific markers, including cardiac troponin T (cTnT), Connexin 43 (Cx43), α‐actinin, and Tbx5. Changes in intracellular calcium ion levels and Ca2+ signal transfer between adjacent cells were monitored using a calcium ion fluorescence probe. mRNA sequencing analysis demonstrated the upregulation of genes associated with cardiac morphogenesis, myocardial differentiation, and muscle fiber contraction during CF differentiation induced by the small‐molecule compounds. Conversely, the expression of fibroblast‐related genes was downregulated. These findings suggest that chemical‐induced cell fate conversion of rat CFs into cardiomyocyte‐like cells is feasible, offering a potential therapeutic solution for myocardial injury.

Activating Ca2+-sensitive enzymes of oxidative metabolism while preventing calcium overload that leads to mitochondrial and cellular injury requires dynamic control of mitochondrial Ca2+ uptake. This is ensured by the mitochondrial calcium uptake (MICU)1/2 proteins that gate the pore of the mitochondrial calcium uniporter (mtCU). MICU1 is relatively sparse in the heart, and recent studies claimed the mammalian heart lacks MICU1 gating of mtCU. However, genetic models have not been tested. We find that MICU1 is present in a complex with MCU in nonfailing human hearts. Furthermore, using murine genetic models and pharmacology, we show that MICU1 and MICU2 control cardiac mitochondrial Ca2+ influx, and that MICU1 deletion alters cardiomyocyte mitochondrial calcium signaling and energy metabolism. MICU1 loss causes substantial compensatory changes in the mtCU composition and abundance, increased turnover of essential MCU regulator (EMRE) early on and, later, of MCU, that limit mitochondrial Ca2+ uptake and allow cell survival. Thus, both the primary consequences of MICU1 loss and the ensuing robust compensation highlight MICU1's relevance in the beating heart. [ABSTRACT FROM AUTHOR]

Hypertrophic cardiomyopathy (HCM) is a cardiovascular pathology that is caused by variants in genes encoding sarcomere-associated proteins. However, the clinical significance of numerous variants in HCM-associated genes is still unknown. CRISPR/Cas9 is a tool of nucleotide sequence editing that allows for the unraveling of different biological tasks. In this study, introducing a mutation with CRISPR/Cas9 into induced pluripotent stem cells (iPSCs) of a healthy donor and the directed differentiation of the isogenic iPSC lines into cardiomyocytes were used to assess the pathogenicity of a variant of unknown significance, p.M659I (c.1977G > A) in MYH7, which was found previously in an HCM patient. Using two single-stranded donor oligonucleotides with and without the p.M659I (c.1977G > A) mutation, together with CRISPR/Cas9, an iPSC line heterozygous at the p.M659I (c.1977G > A) variant in MYH7 was generated. No CRISPR/Cas9 off-target activity was observed. The iPSC line with the introduced p.M659I (c.1977G > A) mutation in MYH7 retained its pluripotent state and normal karyotype. Compared to the isogenic control, cardiomyocytes derived from the iPSCs with the introduced p.M659I (c.1977G > A) mutation in MYH7 recapitulated known HCM features: enlarged size, elevated diastolic calcium level, changes in the expression of HCM-related genes, and disrupted energy metabolism. These findings indicate the pathogenicity of the variant. [ABSTRACT FROM AUTHOR]

Synaptopodin 2-like protein (SYNPO2L) is localized in the sarcomere of cardiomyocytes and is involved in heart morphogenesis. However, the molecular function of SYNPO2L in the heart is not fully understood. We investigated the interaction of SYNPO2L with sarcomeric α-actinin and actin filaments in cultured mouse cardiomyocytes. Immunofluorescence studies showed that SYNPO2L colocalized with α-actinin and actin filaments at the Z-discs of the sarcomere. Recombinant SYNPO2La or SYNPO2Lb caused a bundling of the actin filaments in the absence of α-actinin and enhanced the α-actinin-dependent formation of actin bundles. In addition, high-speed atomic force microscopy revealed that SYNPO2La directly bound to α-actinin via its globular ends. The interaction between α-actinin and SYNPO2La fixed the movements of the two proteins on the actin filaments. These results strongly suggest that SYNPO2L cooperates with α-actinin during actin bundle formation to facilitate sarcomere formation and maintenance. [ABSTRACT FROM AUTHOR]

From birth to adulthood, the mammalian heart grows primarily through increasing cardiomyocyte (CM) size, which is known as maturational hypertrophic growth. The Hippo-YAP signaling pathway is well known for regulating heart development and regeneration, but its roles in CM maturational hypertrophy have not been clearly addressed. Vestigial-like 4 (VGLL4) is a crucial component of the Hippo-YAP pathway, and it functions as a suppressor of YAP/TAZ, the terminal transcriptional effectors of this signaling pathway. To develop an in vitro model for studying CM maturational hypertrophy, we compared the biological effects of T3 (triiodothyronine), Dex (dexamethasone), and T3/Dex in cultured neonatal rat ventricular myocytes (NRVMs). The T3/Dex combination treatment stimulated greater maturational hypertrophy than either the T3 or Dex single treatment. Using T3/Dex treatment of NRVMs as an in vitro model, we found that activation of VGLL4 suppressed CM maturational hypertrophy. In the postnatal heart, activation of VGLL4 suppressed heart growth, impaired heart function, and decreased CM size. On the molecular level, activation of VGLL4 inhibited the PI3K-AKT pathway, and disrupting VGLL4 and TEAD interaction abolished this inhibition. In conclusion, our data suggest that VGLL4 suppresses CM maturational hypertrophy by inhibiting the YAP/TAZ-TEAD complex and its downstream activation of the PI3K-AKT pathway. [ABSTRACT FROM AUTHOR]

Atrial fibrillation (AF) is the most common cardiac rhythm disorder, often occurring in the setting of atrial distension and elevated myocardialstretch. While various mechano-electrochemical signal transduction pathways have been linked to AF development and progression, the underlying molecular mechanisms remain poorly understood, hampering AF therapies. In this review, we describe different aspects of stretch-induced electro-anatomical remodeling as seen in animal models and in patients with AF. Specifically, we focus on cellular and molecular mechanisms that are responsible for mechano-electrochemical signal transduction and the development of ectopic beats triggering AF from pulmonary veins, the most common source of paroxysmal AF. Furthermore, we describe structural changes caused by stretch occurring before and shortly after the onset of AF as well as during AF progression, contributing to longstanding forms of AF. We also propose mechanical stretch as a new dimension to the concept "AF begets AF", in addition to underlying diseases. Finally, we discuss the mechanisms of these electro-anatomical alterations in a search for potential therapeutic strategies and the development of novel antiarrhythmic drugs targeted at the components of mechano-electrochemical signal transduction not only in cardiac myocytes, but also in cardiac non-myocyte cells. • Atrial fibrillation (AF) often occurs in the setting of atrial distension and elevated myocardial stretch. • Stretch triggers AF by promoting ectopic automaticity, electro-anatomical remodeling, and impaired contractile function. • Chronically, these suppress atrial contractility leading to dilation and further stretch,promoting longstanding AF. • Targeting downstream stretch pathways may prevent AF progression and decrease the risk of associated complications. • Non-invasive assessment of atrial structure and function can predict and stratify arrhythmia risk in vulnerable patients. [ABSTRACT FROM AUTHOR]

Carbon black nanoparticles (CBNPs) are widely distributed in the environment and are increasingly recognized as a contributor in the development of cardiovascular disease. A variety of cardiac injuries and diseases result from structural and functional damage to cardiomyocytes. This study explored the mechanisms of CBNPs-mediated myocardial toxicity. CBNPs were given to mice through intra-tracheal instillation and it was demonstrated that the particles can be taken up into the cardiac tissue. Exposure to CBNPs induced cardiomyocyte inflammation and apoptosis. In combination with in vitro experiments, we showed that CBNPs increased the ROS and induced mitochondria fragmentation. Functionally, CBNPs-exposed cardiomyocyte exhibited depolarization of the mitochondrial membrane potential, release of cytochrome c , and activation of pro-apoptotic BAX, thereby initiating programmed cell death. On the other hand, CBNPs impaired autophagy, leading to the inadequate removal of dysfunctional mitochondria. The excess accumulation of damaged mitochondria further stimulated NF-κB activation and triggered the NLRP3 inflammasome pathway. Both the antioxidant N -acetylcysteine and the autophagy activator rapamycin were effective to attenuate the damage of CBNPs on cardiomyocytes. Taken together, this study elucidated the potential mechanism underlying CBNPs-induced myocardial injury and provided a scientific reference for the evaluation and prevention of the CBNPs-related heart risk. [Display omitted] • CBNPs can be taken up into cardiomyocytes through intra-tracheal instillation and induce myocardial toxicity. • ROS mediates myocardial cell damage through disruption of mitochondrial quality control. • Inhibition of mitophagy triggers the inflammation and mitochondrial-dependent apoptosis in myocardial cell. [ABSTRACT FROM AUTHOR]

A thorough characterization of induced pluripotent stem cells (iPSCs) used with in vitro models or therapeutics is essential. Even iPSCs derived from a single donor can exhibit variability within and between cell lines, which can lead to heterogeneity in results and hinder the promising future of cell replacement therapies. In this study, the cell seeding density of human and rhesus monkey iPSCs was tested to maximize the cell line-specific yield of the generated cardiomyocytes. We found that, despite using the same iPSC generation and differentiation protocols, the cell seeding density for the cell line-specific best differentiation efficiency could differ by a factor of four for the four cell lines used here. In addition, the cell lines showed differences in the range of cell seeding densities that they could tolerate without the severe loss of differentiation efficiency. Overall, our data show that the cell seeding density is a critical parameter for the differentiation inefficiency of primate iPSCs to cardiomyocytes and that iPSCs generated with the same episomal approach still exhibit considerable heterogeneity. Therefore, individual characterization of iPSC lines is required, and functional comparability with in vivo processes must be ensured to warrant the translatability of in vitro research with iPSCs. [ABSTRACT FROM AUTHOR]

Altered ankyrin-R (AnkR; encoded by ANK1) expression is associated with diastolic function, left ventricular remodeling, and heart failure with preserved ejection fraction (HFpEF). First identified in erythrocytes, the role of AnkR in other tissues, particularly the heart, is less studied. Here, we identified the expression of both canonical and small isoforms of AnkR in the mouse myocardium. We demonstrate that cardiac myocytes primarily express small AnkR (sAnkR), whereas cardiac fibroblasts predominantly express canonical AnkR. As canonical AnkR expression in cardiac fibroblasts is unstudied, we focused on expression and localization in these cells. AnkR is expressed in both the perinuclear and cytoplasmic regions of fibroblasts with considerable overlap with the trans-Golgi network protein 38, TGN38, suggesting a potential role in trafficking. To study the role of AnkR in fibroblasts, we generated mice lacking AnkR in activated fibroblasts (Ank1-ifKO mice). Notably, Ank1-ifKO mice fibroblasts displayed reduced collagen compaction, supportive of a novel role of AnkR in normal fibroblast function. At the whole animal level, in response to a heart failure model, Ank1-ifKO mice displayed an increase in fibrosis and T-wave inversion compared with littermate controls, while preserving cardiac ejection fraction. Collagen type I fibers were decreased in the Ank1-ifKO mice, suggesting a novel function of AnkR in the maturation of collagen fibers. In summary, our findings illustrate the novel expression of AnkR in cardiac fibroblasts and a potential role in cardiac function in response to stress. [ABSTRACT FROM AUTHOR]

Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) are a cell model now widely used to investigate pathophysiological features of cardiac tissue. Given the invaluable contribution hiPSC-CM could make for studies on cardio-metabolic disorders by defining a postnatal metabolic phenotype, our work herein focused on monitoring the insulin response in CM derived from the hiPSC line UKBi015-B. Western blot analysis on total cell lysates obtained from hiPSC-CM showed increased phosphorylation of both AKT and AS160 following insulin treatment, but failed to highlight any changes in the expression dynamics of the glucose transporter GLUT4. By contrast, the Western blot analysis of membrane fractions, rather than total lysates, revealed insulin-induced plasma membrane translocation of GLUT4, which is known to also occur in postnatal CM. Thus, these findings suggest that hiPSC-derived CMs exhibit an insulin response reminiscent to that of adult CMs regarding intracellular signaling and GLUT4 translocation to the plasma membrane, representing a suitable cellular model in the cardio-metabolic research field. Moreover, our studies also demonstrate the relevance of analyzing membrane fractions rather than total lysates in order to monitor GLUT4 dynamics in response to metabolic regulators in hiPSC-CMs. [ABSTRACT FROM AUTHOR]

Factors responsible for cardiomyocyte proliferation could serve as potential therapeutics to stimulate endogenous myocardial regeneration following insult, such as ischemic injury. A previously published forward genetics approach on cardiomyocyte cell cycle and ploidy led us to the transcription factor, Runx1. Here, we examine the effect of Runx1 on cardiomyocyte cell cycle during postnatal development and cardiac regeneration using cardiomyocyte-specific gain- and loss-of-function mouse models. RUNX1 is expressed in cardiomyocytes during early postnatal life, decreases to negligible levels by 3 wk of age, and increases upon myocardial injury, all consistent with observed rates of cardiomyocyte cell-cycle activity. Loss of Runx1 transiently stymied cardiomyocyte cell-cycle activity during normal postnatal development, a result that corrected itself and did not extend to the context of neonatal heart regeneration. On the other hand, cardiomyocyte-specific Runx1 overexpression resulted in an expansion of diploid cardiomyocytes in uninjured hearts and expansion of 4 N cardiomyocytes in the context of neonatal cardiac injury, suggesting Runx1 overexpression is sufficient to induce cardiomyocyte cell-cycle responses. Persistent overexpression of Runx1 for >1 mo continued to promote cardiomyocyte cell-cycle activity resulting in substantial hyperpolyploidization (≥8 N DNA content). This persistent cell-cycle activation was accompanied by ventricular dilation and adverse remodeling, raising the concern that continued cardiomyocyte cell cycling can have detrimental effects. NEW & NOTEWORTHY:Runx1 is sufficient but not required for cardiomyocyte cell cycle. [ABSTRACT FROM AUTHOR]

Hypercholesterolemia is one of the most important risk factors for cardiovascular diseases. However, it is mostly associated with vascular dysfunction and atherosclerotic lesions, while evidence of direct effects of hypercholesterolemia on cardiomyocytes and heart function is still incomplete and controversial. In this study, we assessed the direct effects of hypercholesterolemia on heart function and the electro-contractile properties of isolated cardiomyocytes. After 5 weeks, male Swiss mice fed with AIN-93 diet added with 1.25% cholesterol (CHO), developed an increase in total serum cholesterol levels and cardiomyocytes cholesterol content. These changes led to altered electrocardiographic records, with a shortening of the QT interval. Isolated cardiomyocytes displayed a shortening of the action potential duration with increased rate of depolarization, which was explained by increased IK, reduced ICa.L and altered INa voltage-dependent inactivation. Also, reduced diastolic [Ca2+]i was found with preserved adrenergic response and cellular contraction function. However, contraction of isolated hearts is impaired in isolated CHO hearts, before and after ischemia/reperfusion, although CHO heart was less susceptible to arrhythmic contractions. Overall, our results demonstrate that early hypercholesterolemia-driven increase in cellular cholesterol content is associated with direct modulation of the heart and cardiomyocytes' excitability, Ca2+ handling, and contraction. [ABSTRACT FROM AUTHOR]

Background. The purpose of the present study was to investigate the effect of aerobic)resistance training on the population of cardiac multipotent cells, thickness, and diameter of the ventricular through the C-Kit pathway of male rats during the growth stages. Methods. Overall, 30 male Wistar rats in three age groups of 2 (n = 10), 8 (n = 10), and 96 (n = 10) weeks were accidentally divided into exercise and control groups. Resistance training programs (resistance ladder, 3 days a week) and aerobics (treadmill running, 3 days a week) were performed for 6 weeks. Hematoxylin-Eosin staining was done the heart tissue and histological images were prepared. Afterward, C-Kit gene expression was examined by the real-time polymerase chain reaction method. The statistical method of one-way analysis of variance (P ≥ 0.05) and Tukey’s test were used at a significant level (P ≥ 0.01). Results. The trained neonatal group had higher values in heart weight to body weight index, and the trained neonatal and trained young adult groups had higher values in left ventricular thickness (P ≥ 0.01). The decrease in left ventricular internal diameter in the trained young adult group was significant compared to the control group (P ≥ 0.01). There was a significant increase in C-Kit gene expression in trained young adult and trained old groups (P ≥ 0.01). Conclusion. Aerobic-resistance training can be an effective stimulus for increasing the number of cardiac stem cells and creating structural adaptations of the heart, including increasing thickness and ventricular diameter in neonatal and young adult groups. Practical Implications. Participation in aerobic-resistance training programs leads to the improvement of the cardiac structure and an increase in cardiac stem cells. [ABSTRACT FROM AUTHOR]

The use of various model organisms has made a huge contribution to understanding the causes and mechanisms of disease development and the study of pathological processes occurring during the development of diseases of the human cardiovascular system, and, in particular, hypertrophic cardiomyopathy (HCM). The optimal solution in the study of primary molecular disturbances is the use of cellular models such as induced pluripotent stem cells (iPSCs), primary rodent cardiomyocytes (CMs), and immortalized lines. In this review, we have focused on the most commonly used cell models, including freshly isolated adult and neonatal rodent CMs, and on the commercially available immortalized cell lines (HL-1, AC16, and H9c2). In order to assess the adequacy of these lines as CM models for studying human myocardial pathologies, a comparative analysis of phenotypic characteristics (morphology, metabolism, calcium homeostasis, etc.) and the nuances of practical use (availability, response to hypertrophic inducers, transfection, etc.) was carried out. The latest published data on the use of these models to assess the pathogenicity of HCM-associated mutations, as well as to screen the effectiveness of developed therapeutic drugs, are also summarized. [ABSTRACT FROM AUTHOR]