RESUMEN
Fully restoring the lost population of cardiomyocytes and heart function remains the greatest challenge in cardiac repair post myocardial infarction. In this study, a pioneered highly ROS-eliminating hydrogel was designed to enhance miR-19a/b induced cardiomyocyte proliferation by lowering the oxidative stress and continuously releasing miR-19a/b in infarcted myocardium in situ. In vivo lineage tracing revealed that â¼20.47 % of adult cardiomyocytes at the injected sites underwent cell division in MI mice. In MI pig the infarcted size was significantly reduced from 40 % to 18 %, and thereby marked improvement of cardiac function and increased muscle mass. Most importantly, our treatment solved the challenge of animal death--all the treated pigs managed to live until their hearts were harvested at day 50. Therefore, our strategy provides clinical conversion advantages and safety for healing damaged hearts and restoring heart function post MI, which will be a powerful tool to battle cardiovascular diseases in patients.
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Proliferación Celular , MicroARNs , Infarto del Miocardio , Miocitos Cardíacos , Estrés Oxidativo , Animales , MicroARNs/metabolismo , MicroARNs/genética , Miocitos Cardíacos/metabolismo , Infarto del Miocardio/metabolismo , Infarto del Miocardio/patología , Estrés Oxidativo/efectos de los fármacos , Ratones , Porcinos , Hidrogeles/química , Ratones Endogámicos C57BL , Especies Reactivas de Oxígeno/metabolismoRESUMEN
ETHNOPHARMACOLOGICAL RELEVANCE: In accordance with the tenets of traditional Chinese medicine, sepsis is categorized into three distinct syndromes: heat syndrome, blood stasis syndrome, and deficiency syndrome. Xiaochaihu decoction (XCHD) has many functions, including the capacity to protect the liver, cholagogue, antipyretic, anti-inflammatory, and anti-pathogenic microorganisms. XCHD exerts the effect of clearing heat and reconciling Shaoyang. The XCHD contains many efficacious active ingredients, yet the mechanism of sepsis-induced cardiomyopathy (SIC) remains elusive. AIM OF THE STUDY: To investigate the molecular mechanisms underlying the protective effects of XCHD against SIC using an integrated approach combining network pharmacology and molecular biology techniques. MATERIALS AND METHODS: Network pharmacology methods identified the active ingredients, target proteins, and pathways affected by XCHD in the context of SIC. We conducted in vivo experiments using mice with lipopolysaccharide-induced SIC, evaluating cardiac function through echocardiography and histology. XCHD-containing serum was analyzed to determine its principal active components using ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The effects of XCHD-containing serum on SIC were further tested in vitro in LPS-treated H9c2 cardiac cells. Protein expression levels were quantified via Western blotting and enzyme-linked immunosorbent assay (ELISA). Additionally, molecular docking was performed between the active components and ZBP1, a potential target protein. Overexpression of ZBP1 in H9c2 cells allowed for a deeper exploration of its role in modulating SIC-associated gene expression. RESULTS: UPLC-MS/MS identified 31 shared XCHD and XCHD-containing serum components. These included organic acids, terpenoids, and flavonoids, which have been identified as the active components of XCHD. Our findings revealed that XCHD alleviated LPS-induced myocardial injury, improved cardiac function, and preserved cardiomyocyte morphology in mice. In vitro studies, we demonstrated that XCHD-containing serum significantly suppressed the expression of inflammatory cytokines (IL-6, IL-1ß, and TNF-α) in LPS-induced H9c2 cells. Mechanistic investigations showed that XCHD downregulated genes associated with PANoptosis, a novel cell death pathway, suggesting its protective role in sepsis-damaged hearts. Conversely, overexpression of ZBP1 abolished the protective effects of XCHD and amplified PANoptosis-related gene expression. CONCLUSIONS: Our study provides the first evidence supporting the protective effects of XCHD against SIC, both in vitro and in vivo. The underlying mechanism involves the inhibition of ZBP1-initiated PANoptosis, offering new insights into treating SIC using XCHD.
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Cardiomiopatías , Medicamentos Herbarios Chinos , Sepsis , Animales , Medicamentos Herbarios Chinos/farmacología , Sepsis/tratamiento farmacológico , Sepsis/complicaciones , Cardiomiopatías/tratamiento farmacológico , Cardiomiopatías/metabolismo , Ratones , Masculino , Línea Celular , Ratones Endogámicos C57BL , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Lipopolisacáridos/toxicidad , Farmacología en Red , Ratas , Modelos Animales de Enfermedad , Espectrometría de Masas en TándemRESUMEN
The heart relies on various defense mechanisms, including metabolic plasticity, to maintain its normal structure and function under high-altitude hypoxia. Pioglitazone, a peroxisome proliferator-activated receptor γ (PPARγ), sensitizes insulin, which in turn regulates blood glucose levels. However, its preventive effects against hypoxia-induced cardiac dysfunction at high altitudes have not been reported. In this study, pioglitazone effectively prevented cardiac dysfunction in hypoxic mice for 4 weeks, independent of its effects on insulin sensitivity. In vitro experiments demonstrated that pioglitazone enhanced the contractility of primary cardiomyocytes and reduced the risk of QT interval prolongation under hypoxic conditions. Additionally, pioglitazone promoted cardiac glucose metabolic reprogramming by increasing glycolytic capacity; enhancing glucose oxidation, electron transfer, and oxidative phosphorylation processes; and reducing mitochondrial reactive ROS production, which ultimately maintained mitochondrial membrane potential and ATP production in cardiomyocytes under hypoxic conditions. Notably, as a PPARγ agonist, pioglitazone promoted hypoxia-inducible factor 1α (HIF-1α) expression in hypoxic myocardium. Moreover, KC7F2, a HIF-1α inhibitor, disrupted the reprogramming of cardiac glucose metabolism and reduced cardiac function in pioglitazone-treated mice under hypoxic conditions. In conclusion, pioglitazone effectively prevented high-altitude hypoxia-induced cardiac dysfunction by reprogramming cardiac glucose metabolism.
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Glucosa , Hipoxia , Miocitos Cardíacos , PPAR gamma , Pioglitazona , Pioglitazona/farmacología , Pioglitazona/uso terapéutico , Animales , PPAR gamma/metabolismo , PPAR gamma/agonistas , Ratones , Glucosa/metabolismo , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Hipoxia/complicaciones , Hipoxia/metabolismo , Masculino , Ratones Endogámicos C57BL , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Tiazolidinedionas/farmacología , Tiazolidinedionas/uso terapéutico , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Background: Cellular senescence has emerged as a pivotal focus in cardiovascular research. This study investigates the previously unrecognized role of cellular senescence in septic cardiomyopathy (SCM) and evaluates senomorphic therapy using ruxolitinib (Rux) as a potential treatment option. Methods: We employed lipopolysaccharide (LPS)-induced neonatal rat cardiomyocytes (NRCMs) and two mouse models-LPS-induced and cecal ligation and puncture (CLP)-induced SCM models-to assess Rux's effects. RNA sequencing, western blotting (WB), quantitative polymerase chain reaction (qPCR), immunofluorescence, immunohistochemistry, senescence-associated ß-galactosidase (SA-ß-gal) assay, and other techniques were utilized to investigate underlying mechanisms. Results: Senescence-associated secretory phenotype (SASP) and cellular senescence markers were markedly elevated in LPS-induced NRCMs and SCM animal models, confirmed by the SA-ß-gal assay. Rux treatment attenuated SASP in vitro and in vivo, alongside downregulation of senescence markers. Moreover, Rux-based senomorphic therapy mitigated mitochondrial-mediated apoptosis, improved cardiac function in SCM mice, restored the balance of antioxidant system, and reduced reactive oxygen species (ROS) levels. Rux treatment restored mitochondrial membrane potential, mitigated mitochondrial morphological damage, and upregulated mitochondrial complex-related gene expression, thereby enhancing mitochondrial function. Additionally, Rux treatment ameliorated SCM-induced mitochondrial dynamic dysfunction and endoplasmic reticulum stress. Mechanistically, Rux inhibited JAK2-STAT3 signaling activation both in vitro and in vivo. Notably, low-dose Rux and ABT263 showed comparable efficacy in mitigating SCM. Conclusions: This study highlighted the potential significance of cellular senescence in SCM pathogenesis and suggested Rux-based senomorphic therapy as a promising therapeutic approach for SCM.
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Cardiomiopatías , Senescencia Celular , Janus Quinasa 2 , Miocitos Cardíacos , Nitrilos , Pirazoles , Pirimidinas , Factor de Transcripción STAT3 , Transducción de Señal , Animales , Janus Quinasa 2/metabolismo , Factor de Transcripción STAT3/metabolismo , Senescencia Celular/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Ratones , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Cardiomiopatías/metabolismo , Cardiomiopatías/tratamiento farmacológico , Nitrilos/uso terapéutico , Nitrilos/farmacología , Ratas , Pirimidinas/farmacología , Pirimidinas/uso terapéutico , Pirazoles/farmacología , Pirazoles/uso terapéutico , Masculino , Ratones Endogámicos C57BL , Sepsis/metabolismo , Sepsis/tratamiento farmacológico , Ratas Sprague-Dawley , Lipopolisacáridos , Modelos Animales de EnfermedadRESUMEN
Rationale: Regulatory processes of transcription factors (TFs) shape heart development and influence the adult heart's response to stress, contributing to cardiac disorders. Despite their significance, the precise mechanisms underpinning TF-mediated regulation remain elusive. Here, we identify that EBF1, as a TF, is highly expressed in human heart tissues. EBF1 is reported to be associated with human cardiovascular disease, but its roles are unclear in heart. In this study, we investigated EBF1 function in cardiac system. Methods: RNA-seq was utilized to profile EBF1 expression patterns. CRISPR/Cas9 was utilized to knock out EBF1 to investigate its effects. Human pluripotent stem cells (hPSCs) differentiated into cardiac lineages were used to mimic cardiac development. Cardiac function was evaluated on mouse model with Ebf1 knockout by using techniques such as echocardiography. RNA-seq was conducted to analyze transcriptional perturbations. ChIP-seq was employed to elucidate EBF1-bound genes and the underlying regulatory mechanisms. Results: EBF1 was expressed in some human and mouse cardiomyocyte. Knockout of EBF1 inhibited cardiac development. ChIP-seq indicated EBF1's binding on promoters of cardiogenic TFs pivotal to cardiac development, facilitating their transcriptional expression and promoting cardiac development. In mouse, Ebf1 depletion triggered transcriptional perturbations of genes, resulting in cardiac remodeling. Mechanistically, we found that EBF1 directly bound to upstream chromatin regions of cardiac hypertrophy-inducing genes, contributing to cardiac hypertrophy. Conclusions: We uncover the mechanisms underlying EBF1-mediated regulatory processes, shedding light on cardiac development, and the pathogenesis of cardiac remodeling. These findings emphasize EBF1's critical role in orchestrating diverse aspects of cardiac processes and provide a promising therapeutic intervention for cardiomyopathy.
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Perfilación de la Expresión Génica , Miocitos Cardíacos , Transactivadores , Animales , Humanos , Ratones , Transactivadores/genética , Transactivadores/metabolismo , Miocitos Cardíacos/metabolismo , Diferenciación Celular/genética , Corazón/fisiopatología , Ratones Noqueados , Células Madre Pluripotentes/metabolismo , Transcriptoma/genética , Sistemas CRISPR-Cas/genéticaRESUMEN
Myocardial ischemia-reperfusion (I/R) injury exacerbates cellular damage upon restoring blood flow to ischemic cardiac tissue, causing oxidative stress, inflammation, and apoptosis. This study investigates Nicotinamide Riboside (NR), a precursor of nicotinamide adenine dinucleotide (NAD+), for its cardioprotective effects. Administering NR to mice before I/R injury and evaluating heart function via echocardiography showed that NR significantly improved heart function, increased left ventricular ejection fraction (LVEF) and fractional shortening (FS), and reduced left ventricular end-diastolic (LVDd) and end-systolic diameters (LVSd). NR also restored E/A and E/e' ratios. It reduced cardiomyocyte apoptosis both in vivo and in vitro, inhibiting elevated caspase-3 activity and returning Bax protein levels to normal. In vitro, NR reduced the apoptotic rate in hydrogen peroxide (H2O2)-treated HL-1 cells from 30% to 10%. Mechanistically, NR modulated the SIRT3/mtROS/JNK pathway, reversing H2O2-induced SIRT3 downregulation, reducing mitochondrial reactive oxygen species (mtROS), and inhibiting JNK activation. Using SIRT3-knockout (SIRT3-KO) mice, we confirmed that NR's cardioprotective effects depend on SIRT3. Echocardiography showed that NR's benefits were abrogated in SIRT3-KO mice. In conclusion, NR provides significant cardioprotection against myocardial I/R injury by enhancing NAD+ levels and modulating the SIRT3/mtROS/JNK pathway, suggesting its potential as a novel therapeutic agent for ischemic heart diseases, meriting further clinical research.
Asunto(s)
Apoptosis , Ratones Noqueados , Daño por Reperfusión Miocárdica , Niacinamida , Compuestos de Piridinio , Especies Reactivas de Oxígeno , Sirtuina 3 , Animales , Sirtuina 3/metabolismo , Sirtuina 3/genética , Daño por Reperfusión Miocárdica/tratamiento farmacológico , Daño por Reperfusión Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/patología , Niacinamida/análogos & derivados , Niacinamida/farmacología , Niacinamida/uso terapéutico , Ratones , Compuestos de Piridinio/farmacología , Compuestos de Piridinio/administración & dosificación , Especies Reactivas de Oxígeno/metabolismo , Apoptosis/efectos de los fármacos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Masculino , Estrés Oxidativo/efectos de los fármacos , Humanos , Cardiotónicos/farmacología , Cardiotónicos/uso terapéutico , Modelos Animales de Enfermedad , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Ratones Endogámicos C57BL , Transducción de Señal/efectos de los fármacosRESUMEN
G protein-coupled receptors' conformational landscape can be affected by their local, microscopic interactions within the cell plasma membrane. We employ here a pleiotropic stimulus, namely osmotic swelling, to alter the cortical environment within intact cells and monitor the response in terms of receptor function and downstream signaling. We observe that in osmotically swollen cells the ß2-adrenergic receptor, a prototypical GPCR, favors an active conformation, resulting in cAMP transient responses to adrenergic stimulation that have increased amplitude. The results are validated in primary cell types such as adult cardiomyocytes, a model system where swelling occurs upon ischemia-reperfusion injury. Our results suggest that receptors' function is finely modulated by their biophysical context, and specifically that osmotic swelling acts as a potentiator of downstream signaling, not only for the ß2-adrenergic receptor, but also for other receptors, hinting at a more general regulatory mechanism.
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AMP Cíclico , Miocitos Cardíacos , Receptores Adrenérgicos beta 2 , Transducción de Señal , Receptores Adrenérgicos beta 2/metabolismo , Miocitos Cardíacos/metabolismo , Humanos , Animales , Ligandos , AMP Cíclico/metabolismo , Membrana Celular/metabolismo , Células HEK293 , RatonesRESUMEN
Ferroptosis is an important pathological mechanism of chronic heart failure (CHF). This study aimed to investigate the protective mechanism of Astragaloside IV (AS-IV) on CHF rats by integrating bioinformatics and ferroptosis. CHF-related targets and ferroptosis-related targets were collected. After the intersection, the common targets were obtained. The PPI network of the common targets was constructed, and topological analysis of the network was carried out. The target with the highest topological parameter values was selected as the key target. The key target p53 was obtained through bioinformatics analysis, and its molecular docking model with AS-IV was obtained, as well as molecular dynamics simulation analysis. The rat models of CHF after myocardial infarction were established by ligation of left coronary artery and treated with AS-IV for 4 weeks. AS-IV treatment significantly improved cardiac function in CHF rats, improved cardiomyocyte morphology and myocardial fibrosis, reduced mitochondrial damage, decreased myocardial MDA and Fe2+ content, increased GSH content, inhibited the expression of p53 and p-p53, and up-regulated the expression of SLC7A11 and GPX4. In conclusion, AS-IV improved cardiac function in CHF rats, presumably by regulating p53/SLC7A11/GPX4 signaling pathway and inhibiting myocardial ferroptosis.
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Biología Computacional , Ferroptosis , Insuficiencia Cardíaca , Saponinas , Triterpenos , Animales , Ferroptosis/efectos de los fármacos , Triterpenos/farmacología , Saponinas/farmacología , Insuficiencia Cardíaca/tratamiento farmacológico , Insuficiencia Cardíaca/metabolismo , Ratas , Biología Computacional/métodos , Masculino , Fosfolípido Hidroperóxido Glutatión Peroxidasa/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Simulación del Acoplamiento Molecular , Enfermedad Crónica , Modelos Animales de Enfermedad , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/patología , Infarto del Miocardio/tratamiento farmacológico , Infarto del Miocardio/metabolismo , Infarto del Miocardio/patología , Simulación de Dinámica Molecular , Miocardio/metabolismo , Miocardio/patologíaRESUMEN
Cardiac biological pacing (BP) is one of the future directions for bradyarrhythmias intervention. Currently, cardiac pacemaker cells (PCs) used for cardiac BP are mainly derived from pluripotent stem cells (PSCs). However, the production of high-quality cardiac PCs from PSCs remains a challenge. Here, we developed a cardiac PC differentiation strategy by adopting dual PC markers and simulating the developmental route of PCs. First, two PC markers, Shox2 and Hcn4, were selected to establish Shox2:EGFP; Hcn4:mCherry mouse PSC reporter line. Then, by stepwise guiding naïve PSCs to cardiac PCs following naïve to formative pluripotency transition and manipulating signaling pathways during cardiac PCs differentiation, we designed the FSK method that increased the yield of SHOX2+; HCN4+ cells with typical PC characteristics, which was 12 and 42 folds higher than that of the embryoid body (EB) and the monolayer M10 methods respectively. In addition, the in vitro cardiac PCs differentiation trajectory was mapped by single-cell RNA sequencing (scRNA-seq), which resembled in vivo PCs development, and ZFP503 was verified as a key regulator of cardiac PCs differentiation. These PSC-derived cardiac PCs have the potential to drive advances in cardiac BP technology, help with the understanding of PCs (patho)physiology, and benefit drug discovery for PC-related diseases as well.
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Diferenciación Celular , Miocitos Cardíacos , Células Madre Pluripotentes , Animales , Ratones , Diferenciación Celular/genética , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/citología , Células Madre Pluripotentes/metabolismo , Células Madre Pluripotentes/citología , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/genética , Canales Regulados por Nucleótidos Cíclicos Activados por Hiperpolarización/metabolismo , Cuerpos Embrioides/citología , Cuerpos Embrioides/metabolismoRESUMEN
Ferroptosis, triggered by iron overload and excessive lipid peroxidation, plays a pivotal role in the progression of DOX-induced cardiomyopathy (DIC), and thus limits the use of doxorubicin (DOX) in clinic. Here, we further showed that cardiac ferroptosis induced by DOX in mice was attributed to up-regulation of Hmox1, as knockdown of Hmox1 effectively inhibited cardiomyocyte ferroptosis. To targeted delivery of siRNA into cardiomyocytes, siRNA-encapsulated exosomes were injected followed by ultrasound microbubble targeted destruction (UTMD) in the heart region. UTMD greatly facilitated exosome delivery into heart. Consistently, UTMD assisted exosomal delivery of siHomox1 nearly blocked the ferroptosis and the subsequent cardiotoxicity induced by doxorubicin. In summary, our findings reveal that the upregulation of HMOX1 induces ferroptosis in cardiomyocytes and UTMD-assisted exosomal delivery of siHmox1 can be used as a potential therapeutic strategy for DIC.
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Doxorrubicina , Exosomas , Ferroptosis , Hemo-Oxigenasa 1 , Microburbujas , Miocitos Cardíacos , ARN Interferente Pequeño , Ferroptosis/efectos de los fármacos , Animales , Doxorrubicina/farmacología , Exosomas/metabolismo , Ratones , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Hemo-Oxigenasa 1/metabolismo , ARN Interferente Pequeño/farmacología , Ratones Endogámicos C57BL , Masculino , Sistemas de Liberación de Medicamentos , Cardiomiopatías/metabolismo , Proteínas de la MembranaRESUMEN
Tyrosine kinase inhibitors (TKIs) offer targeted therapy for cancers but can cause severe cardiotoxicities. Determining their dosedependent impact on cardiac function is required to optimize therapy and minimize adverse effects. The dosedependent cardiotoxic effects of two TKIs, imatinib and ponatinib, were assessed in vitro using H9c2 cardiomyoblasts and in vivo using zebrafish embryos. In vitro, H9c2 cardiomyocyte viability, apoptosis, size, and surface area were evaluated to assess the impact on cellular health. In vivo, zebrafish embryos were analyzed for heart rate, blood flow velocity, and morphological malformations to determine functional and structural changes. Additionally, reverse transcriptionquantitative PCR (RTqPCR) was employed to measure the gene expression of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), established markers of cardiac injury. This comprehensive approach, utilizing both in vitro and in vivo models alongside functional and molecular analyses, provides a robust assessment of the potential cardiotoxic effects. TKI exposure decreased viability and surface area in H9c2 cells in a dosedependent manner. Similarly, zebrafish embryos exposed to TKIs exhibited dosedependent heart malformation. Both TKIs upregulated ANP and BNP expression, indicating heart injury. The present study demonstrated dosedependent cardiotoxic effects of imatinib and ponatinib in H9c2 cells and zebrafish models. These findings emphasize the importance of tailoring TKI dosage to minimize cardiac risks while maintaining therapeutic efficacy. Future research should explore the underlying mechanisms and potential mitigation strategies of TKIinduced cardiotoxicities.
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Cardiotoxicidad , Mesilato de Imatinib , Imidazoles , Miocitos Cardíacos , Piridazinas , Pez Cebra , Animales , Pez Cebra/embriología , Imidazoles/toxicidad , Piridazinas/efectos adversos , Piridazinas/farmacología , Piridazinas/toxicidad , Mesilato de Imatinib/toxicidad , Mesilato de Imatinib/efectos adversos , Mesilato de Imatinib/farmacología , Cardiotoxicidad/etiología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Inhibidores de Proteínas Quinasas/efectos adversos , Inhibidores de Proteínas Quinasas/toxicidad , Inhibidores de Proteínas Quinasas/farmacología , Línea Celular , Péptido Natriurético Encefálico/metabolismo , Péptido Natriurético Encefálico/genética , Embrión no Mamífero/efectos de los fármacos , Embrión no Mamífero/metabolismo , Supervivencia Celular/efectos de los fármacos , Apoptosis/efectos de los fármacos , Mioblastos Cardíacos/efectos de los fármacos , Mioblastos Cardíacos/metabolismo , RatasRESUMEN
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 ß-adrenoceptor (ß-AR) and its effects on various in vivo and in vitro parameters, including electrocardiogram (ECG) measurements, left ventricular developed pressure (LVDP), the ß-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 ß1-AR). In vivo, s-Lim antagonized the effect of ISO (agonists ß1-AR), presenting a similar effect to propranolol (a non-selective blocker ß-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 ß-AR in the heart.
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Antiarrítmicos , Limoneno , Ratas Wistar , Receptores Adrenérgicos beta , Transducción de Señal , Animales , Ratas , Antiarrítmicos/farmacología , Masculino , Receptores Adrenérgicos beta/metabolismo , Limoneno/farmacología , Transducción de Señal/efectos de los fármacos , Terpenos/farmacología , Corazón/efectos de los fármacos , Frecuencia Cardíaca/efectos de los fármacos , Ciclohexenos/farmacología , Arritmias Cardíacas/tratamiento farmacológico , Arritmias Cardíacas/metabolismo , Arritmias Cardíacas/inducido químicamente , Arritmias Cardíacas/fisiopatología , Isoproterenol/farmacología , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismoRESUMEN
This study aimed to investigate the effects and possible mechanisms of adenylate cyclase 1 (ADCY1) on pirarubicin-induced cardiomyocyte injury. HL-1 cells were treated with pirarubicin (THP) to induce intracellular toxicity, and the extent of damage to mouse cardiomyocytes was assessed using CCK-8, Edu, flow cytometry, ROS, ELISA, RT-qPCR and western blotting. THP treatment reduced the viability of HL-1 cells, inhibited proliferation, induced apoptosis and triggered oxidative stress. In addition, the RT-qPCR results revealed that ADCY1 expression was significantly elevated in HL-1 cells, and molecular docking showed a direct interaction between ADCY1 and THP. Western blotting showed that ADCY1, phospho-protein kinase A and GRIN2D expression were also significantly elevated. Knockdown of ADCY1 attenuated THP-induced cardiotoxicity, possibly by regulating the ADCY1/PKA/GRIN2D pathway.
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Adenilil Ciclasas , Cardiotoxicidad , Doxorrubicina , Técnicas de Silenciamiento del Gen , Miocitos Cardíacos , Adenilil Ciclasas/metabolismo , Adenilil Ciclasas/genética , Animales , Ratones , Cardiotoxicidad/genética , Doxorrubicina/toxicidad , Doxorrubicina/farmacología , Doxorrubicina/análogos & derivados , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Línea Celular , Apoptosis/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/genética , Simulación del Acoplamiento Molecular , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , Antineoplásicos/farmacología , Antineoplásicos/toxicidadRESUMEN
Engineered heart tissues (EHTs) generated from human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) represent powerful platforms for human cardiac research, especially in drug testing and disease modeling. Here, we report a flexible, three-dimensional electronic framework that enables real-time, spatiotemporal analysis of electrophysiologic and mechanical signals in EHTs under physiological loading conditions for dynamic, noninvasive, longer-term assessments. These electromechanically monitored EHTs support multisite measurements throughout the tissue under baseline conditions and in response to stimuli. Demonstrations include uses in tracking physiological responses to pharmacologically active agents and in capturing electrophysiological characteristics of reentrant arrhythmias. This platform facilitates precise analysis of signal location and conduction velocity in human cardiomyocyte tissues, as the basis for a broad range of advanced cardiovascular studies.
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Células Madre Pluripotentes Inducidas , Miocitos Cardíacos , Ingeniería de Tejidos , Humanos , Ingeniería de Tejidos/métodos , Miocitos Cardíacos/fisiología , Miocitos Cardíacos/metabolismo , Células Madre Pluripotentes Inducidas/citología , Corazón/fisiología , Fenómenos ElectrofisiológicosRESUMEN
Drug-induced gene expression profiles can identify potential mechanisms of toxicity. We focus on obtaining signatures for cardiotoxicity of FDA-approved tyrosine kinase inhibitors (TKIs) in human induced-pluripotent-stem-cell-derived cardiomyocytes, using bulk transcriptomic profiles. We use singular value decomposition to identify drug-selective patterns across cell lines obtained from multiple healthy human subjects. Cellular pathways affected by cardiotoxic TKIs include energy metabolism, contractile, and extracellular matrix dynamics. Projecting these pathways to published single cell expression profiles indicates that TKI responses can be evoked in both cardiomyocytes and fibroblasts. Integration of transcriptomic outlier analysis with whole genomic sequencing of our six cell lines enables us to correctly reidentify a genomic variant causally linked to anthracycline-induced cardiotoxicity and predict genomic variants potentially associated with TKI-induced cardiotoxicity. We conclude that mRNA expression profiles when integrated with publicly available genomic, pathway, and single cell transcriptomic datasets, provide multiscale signatures for cardiotoxicity that could be used for drug development and patient stratification.
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Cardiotoxicidad , Perfilación de la Expresión Génica , Miocitos Cardíacos , Inhibidores de Proteínas Quinasas , Transcriptoma , Humanos , Miocitos Cardíacos/efectos de los fármacos , Miocitos Cardíacos/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/toxicidad , Perfilación de la Expresión Génica/métodos , Cardiotoxicidad/genética , Cardiotoxicidad/etiología , Células Madre Pluripotentes Inducidas/metabolismo , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Línea Celular , Análisis de la Célula Individual/métodos , Fibroblastos/efectos de los fármacos , Fibroblastos/metabolismoRESUMEN
Minocycline (Min), as an antibiotic, possesses various beneficial properties such as anti-inflammatory, antioxidant, and anti-apoptotic effects. Despite these known qualities, the precise cardioprotective effect and mechanism of Min in protecting against sepsis-induced cardiotoxicity (SIC) remain unspecified. To address this, our study sought to assess the protective effects of Min on the heart. Lipopolysaccharide (LPS) was utilized to establish a cardiotoxicity model both in vivo and in vitro. Min was pretreated in the models. In the in vivo setting, evaluation of heart tissue histopathological injury was performed using hematoxylin and eosin (H&E) staining and TUNEL. Immunohistochemistry (IHC) was employed to evaluate the expression levels of NLRP3 and Caspase-1 in the heart tissue of mice. During in vitro experiments, the viability of H9c2 cells was gauged utilizing the CCK8 assay kit. Intracellular ROS levels in H9c2 cells were quantified using a ROS assay kit. Both in vitro and in vivo settings were subjected to measurement of oxidative stress indexes, encompassing glutathione (GSH), malondialdehyde (MDA), and superoxide dismutase (SOD) levels. Additionglly, myocardial injury markers like lactate dehydrogenase (LDH) and creatine kinase MB (CK-MB) activity were quantified using appropriate assay kits. Western blotting (WB) analysis was conducted to detect the expression levels of NOD-like receptor protein-3 (NLRP3), caspase-1, IL-18, and IL-1ß, alongside apoptosis-related proteins such as Bcl-2 and Bax, and antioxidant proteins including superoxide dismutase-1 (SOD-1) and antioxidant proteins including superoxide dismutase-1 (SOD-2), both in H9c2 cells and mouse heart tissues. In vivo, Min was effective in reducing LPS-induced inflammation in cardiac tissue, preventing cell damage and apoptosis in cardiomyocytes. The levels of LDH and CK-MB were significantly reduced with Min treatment. In vitro studies showed that Min improved the viability of H9C2 cells, reduced apoptosis, and decreased ROS levels in these cells. Further analysis indicated that Min decreased the protein levels of NLRP3, Caspase-1, IL-18, and IL-1ß, while increasing the levels of SOD-1 and SOD-2 both in vivo and in vitro. Min alleviates LPS-induced SIC by suppressing the NLRP3/Caspase-1 signalling pathway in vivo and in vitro.
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Cardiotoxicidad , Caspasa 1 , Lipopolisacáridos , Minociclina , Proteína con Dominio Pirina 3 de la Familia NLR , Transducción de Señal , Animales , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Transducción de Señal/efectos de los fármacos , Lipopolisacáridos/toxicidad , Caspasa 1/metabolismo , Cardiotoxicidad/metabolismo , Cardiotoxicidad/tratamiento farmacológico , Ratones , Minociclina/farmacología , Masculino , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Línea Celular , Apoptosis/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismo , Ratones Endogámicos C57BL , RatasRESUMEN
Myocardial ischemia-reperfusion injury (MIRI) is a significant complication following reperfusion therapy after myocardial infarction. Mitochondrial oxidative stress is a critical factor in MIRI, and Sirtuin 3 (SIRT3), as a major mitochondrial deacetylase, plays a key protective role, with its activity potentially regulated by O-GlcNAcylation. This study used the H9C2 cell line to establish a simulated ischemia/reperfusion (SI/R) model, we utilized co-immunoprecipitated to validate the relationship between O-GlcNAc transferase (OGT) and SIRT3, demonstrated SIRT3 O-GlcNAcylation sites through LC-MS/MS, and performed site mutations using CRISPR/Cas9 technology. The results were validated using immunoblotting. SIRT3 and superoxide dismutase 2 (SOD2) activities were detected using a fluorometric assay, while mitochondrial reactive oxygen species (MROS) levels and cellular apoptosis were assessed using immunofluorescence. We have identified an interaction between SIRT3 and OGT, where SIRT3 undergoes dynamic O-GlcNAcylation at the S190 site, facilitating SIRT3 deacetylase activity. During SI/R, elevated levels of O-GlcNAcylation activate SOD2 by promoting SIRT3 enzyme activity, thereby inhibiting excessive MROS production. This significantly mitigates the occurrence of malignant autophagy in myocardial cells during reperfusion, promoting their survival. Conversely, blocking SIRT3 O-GlcNAcylation at the S190 site exacerbates SI/R injury. We demonstrate that O-GlcNAcylation is a crucial post-translational modification (PTM) of SIRT3 during SI/R, shedding light on a promising mechanism for future therapeutic approaches.
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Daño por Reperfusión Miocárdica , Estrés Oxidativo , Sirtuina 3 , Superóxido Dismutasa , Sirtuina 3/metabolismo , Daño por Reperfusión Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/patología , Animales , Superóxido Dismutasa/metabolismo , Línea Celular , Ratas , Especies Reactivas de Oxígeno/metabolismo , N-Acetilglucosaminiltransferasas/metabolismo , Mitocondrias/metabolismo , Apoptosis , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Humanos , SirtuinasRESUMEN
Rationale: Parkin (an E3 ubiquitin protein ligase) is an important regulator of mitophagy. However, the role of Parkin in viral myocarditis (VMC) remains unclear. Methods: Coxsackievirus B3 (CVB3) infection was induced in mice to create VMC. Cardiac function and inflammatory response were evaluated by echocardiography, histological assessment, and molecular analyses. AAV9 (adeno-associated virus 9), transmission electron microscopy (TEM) and western blotting were used to investigate the mechanisms by which Parkin regulates mitophagy and cardiac inflammation. Results: Our data indicated that Parkin- and BNIP3 (BCL2 interacting protein 3 like)-mediated mitophagy was activated in VMC mice and neonatal rat cardiac myocytes (NRCMs) infected with CVB3, which blocked autophagic flux by inhibiting autophagosome-lysosome fusion. Parkin silencing aggravated mortality and accelerated the development of cardiac dysfunction in CVB3-treated mice. While silencing of Parkin did not significantly increase inflammatory response through activating NF-κB pathway and production of inflammatory cytokines post-VMC, the mitophagy activity were reduced, which stimulated the accumulation of damaged mitochondria. Moreover, Parkin silencing exacerbated VMC-induced apoptosis. We consistently found that Parkin knockdown disrupted mitophagy activity and inflammatory response in NRCMs. Conclusion: This study elucidated the important role of Parkin in maintaining cardiac function and inflammatory response by regulating mitophagy activity and the NF-κB pathway during acute VMC. Although the functional impact of mitophagy remains unclear, our findings suggest that Parkin silencing may accelerate VMC development.
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Infecciones por Coxsackievirus , Mitofagia , Miocarditis , Miocitos Cardíacos , Ubiquitina-Proteína Ligasas , Animales , Ubiquitina-Proteína Ligasas/metabolismo , Ubiquitina-Proteína Ligasas/genética , Miocarditis/virología , Miocarditis/metabolismo , Ratones , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/virología , Infecciones por Coxsackievirus/metabolismo , Infecciones por Coxsackievirus/virología , Masculino , Ratas , Enterovirus Humano B/fisiología , Apoptosis , Modelos Animales de Enfermedad , Proteínas de la Membrana/metabolismo , Proteínas de la Membrana/genética , Proteínas Mitocondriales/metabolismo , Proteínas Mitocondriales/genética , FN-kappa B/metabolismo , Ratones Endogámicos BALB CRESUMEN
High mitochondrial DNA (mtDNA) amount has been reported to be beneficial for resistance and recovery of metabolic stress, while increased mtDNA synthesis activity can drive aging signs. The intriguing contrast of these two mtDNA boosting outcomes prompted us to jointly elevate mtDNA amount and frequency of replication in mice. We report that high activity of mtDNA synthesis inhibits perinatal metabolic maturation of the heart. The offspring of the asymptomatic parental lines are born healthy but manifest dilated cardiomyopathy and cardiac collapse during the first days of life. The pathogenesis, further enhanced by mtDNA mutagenesis, involves prenatal upregulation of mitochondrial integrated stress response and the ferroptosis-inducer MESH1, leading to cardiac fibrosis and cardiomyocyte death after birth. Our evidence indicates that the tight control of mtDNA replication is critical for early cardiac homeostasis. Importantly, ferroptosis sensitivity is a potential targetable mechanism for infantile-onset cardiomyopathy, a common manifestation of mitochondrial diseases.
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Replicación del ADN , ADN Mitocondrial , Miocitos Cardíacos , Animales , ADN Mitocondrial/genética , ADN Mitocondrial/metabolismo , Ratones , Miocitos Cardíacos/metabolismo , Femenino , Masculino , Cardiomiopatía Dilatada/genética , Cardiomiopatía Dilatada/metabolismo , Cardiomiopatía Dilatada/patología , Ferroptosis/genética , Miocardio/metabolismo , Miocardio/patología , Mitocondrias Cardíacas/metabolismo , Mitocondrias Cardíacas/genética , Ratones Endogámicos C57BL , Animales Recién Nacidos , Humanos , Corazón/fisiopatología , FibrosisRESUMEN
Introduction: Apigenin is a natural flavonoid compound with promising potential for the attenuation of myocardial hypertrophy (MH). The compound can also modulate the expression of miR-185-5p that both promote MH and suppress autophagy. The current attempts to explain the anti-MH effect of apigenin by focusing on changes in miR-185-5p-mediated autophagy. Methods: Hypertrophic symptoms were induced in rats using transverse aortic constriction (TAC) method and in cardiomyocytes using Ang II and then handled with apigenin. Changes in myocardial function and structure and cell viability and surface area were measured. The role of miR-185-5p in the anti-MH function of apigenin was explored by detecting changes in autophagic processes and miR-185-5p/SREBP2 axis. Results: TAC surgery induced weight increase, structure destruction, and collagen deposition in hearts of model rats. Ang II suppresses cardiomyocyte viability and increased cell surface area. All these impairments were attenuated by apigenin and were associated with the restored level of autophagy. At the molecular level, the expression of miR-185-5p was up-regulated by TAC, while the expression of SREBP2 was down-regulated, which was reserved by apigenin both in vivo and in vitro. The induction of miR-185-5p in cardiomyocytes could counteracted the protective effects of apigenin. Discussion: Collectively, the findings outlined in the current study highlighted that apigenin showed anti-MH effects. The effects were related to the inhibition of miR-185-5p and activation of SREBP, which contributed to the increased autophagy.