This work deals with the electrophysiology of cardiac cells, their electro-stimulation and design of stimulation device to extend their life. The first part is focused on the action potential in general, in muscle cells, followed by an analysis of the process and characteristics of the action potential in cardiomyocytes. The next section provides a procedure for isolating and process of changes that lead to cell death and of a method that the process would be eliminated. Part of this work is a program for processing and evaluation of pictures obtained during electro-stimulation process.

Sodium-Glucose Linked Transporter 2 (SGLT2) inhibitors block renal glucose reabsorption and have shown marked cardiac protection in type 2 diabetics, and surprisingly, also in non-diabetics. However, the mechanism by which these drugs improve cardiovascular outcomes is unknown. Metabolic heart disease, which is characterized by cardiac hypertrophy and diastolic dysfunction, is associated with obesity and insulin resistance and leads to adverse cardiovascular outcomes including heart failure with a preserved ejection fraction. A high fat, high sucrose “Western” diet can induce metabolic syndrome, an aggregate of obesity-driven clinical phenotypes including insulin resistance, elevated triglycerides, hypertension, and abnormal cholesterol. Using a mouse model of metabolic syndrome and adult rat ventricular myocytes (ARVMs) in vitro, we aim to determine if the SGLT2 inhibitor Ertugliflozin (ERTU) can prevent metabolic syndrome-induced cardiac pathophysiology and whether ERTU can exert a direct action on cardiomyocytes, a cell type lacking SGLT2. SGLT2 inhibitors have been proposed to act directly on the Sodium-Hydrogen Exchanger 1 (NHE1) and thus could have direct action on cardiomyocytes that may mediate cardioprotective effects. Mice were fed either a control diet (CD) or a high fat high sucrose (HFHS) diet ± ERTU for 16 weeks. Echocardiography was performed and heart weights were obtained. ARVMs were used to assess ERTU’s effect on insulin sensitivity in vitro in a high-palmitate, insulin resistance model, and to test the efficacy of the known NHE1 inhibitor Cariporide (NHEi). A NHE1 activity ammonium chloride pulse assay was performed in HEK293 cells over-expressing either wild-type (WT) or a known NHEi-insensitive point mutant NHE1 ± NHEi or ERTU. In HFHS-fed mice, ERTU attenuated weight gain and restored blood glucose, insulin, hemoglobin A1c, and HOMA-IR to CD levels. HFHS-induced cardiac hypertrophy and diastolic dysfunction were prevented with ERTU. In vitro, high palmitate media decreased insulin stimulated AKT signaling compared to low palmitate media and was rescued by either ERTU or NHEi treatment. ERTU inhibited WT NHE1 activity in HEK293 cells by 67%, whereas activity of the NHEi-insensitive mutant NHE1 was unaffected by ERTU treatment. ERTU prevented the hallmarks of diet-induced metabolic heart disease (cardiac hypertrophy and diastolic dysfunction) in mice. These benefits exceed the expected consequences of glucose control alone. The actions of ERTU on ARVMs in vitro suggest the favorable effects on cardiac structure and function may be due, at least in part, to the direct action of the drug on cardiomyocytes. Furthermore, mutational overexpression studies show that ERTU can directly affect NHE1 in cardiac myocytes. Taken together, this thesis provides evidence that the direct cardioprotective effects of ERTU could be via inhibition of NHE1, a critical modulator of intracellular pH and sodium in the cardiomyocyte, with known implications in the pathophysiology of diabetes and heart failure. / 2022-06-04T00:00:00Z

Background: The canonical Wnt/β-catenin pathway is activated in a variety of heart diseases, such as myocardial infarction and cardiac hypertrophy, that are associated with altered ion channel expressions and increased risk of cardiac arrhythmias. Previous work from our lab has demonstrated that the Wnt/β-catenin signalling (Wnt signalling) inhibits sodium (Na+) current in rat cardiomyocytes. In this project, we aim to investigate the mechanisms that underlie the inhibition of Na+ current by Wnt signalling in both rat and human cardiomyocytes. Results: In both neonatal rat ventricular myocytes (NRVMs) and human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), activation of the Wnt/β-catenin signalling led to reduced level of Na+ channel gene transcript (Scn5a), channel protein (Nav1.5) and channel current density. This suggests that reduced Scn5a expression is likely the primary mechanism for reduced Na+ current. In addition, we found that activation of the Wnt/β-catenin signalling in both NRVMs and iPSC-CMs upregulated Tbx3 transcript and protein levels, which is a transcription factor that is known to suppress Scn5a transcription. In NRVMs, siRNA-mediated Tbx3 knockdown attenuated (by ~30%) Wnt-induced reductions in Scn5a and Nav1.5 levels. Conclusions: Our findings are consistent with the conclusion that Wnt/β-catenin signalling inhibits Na+ current in both rat and human cardiomyocytes by reducing Scn5a levels, with Tbx3 as one of the mediators.

Elevated circulatory free fatty acids (FFAs) especially saturated FFAs, such as palmitate (PA), are detrimental to the heart. However, mechanisms responsible for this phenomenon remain unknown. Here, the role of JAK2/STAT3 in PA-induced cytotoxicity was investigated in cardiomyocytes. We demonstrate that PA suppressed the JAK2/STAT3 pathway by dephosphorylation of JAK2 (Y1007/1008) and STAT3 (Y705), and thus blocked the translocation of STAT3 into the nucleus. Conversely, phosphorylation of S727, another phosphorylated site of STAT3, was increased in response to PA treatment. Pretreatment of JNK inhibitor, but not p38 MAPK inhibitor, inhibited STAT3 (S727) activation induced by PA and rescued the phosphorylation of STAT3 (Y705). The data suggested that JNK may be another upstream factor regulating STAT3, and verified the important function of P-STAT3 (Y705) in PA-induced cardiomyocyte apoptosis. Sodium orthovanadate (SOV), a protein tyrosine phosphatase inhibitor, obviously inhibited PA-induced apoptosis by restoring JAK2/STAT3 pathways. This effect was diminished by STAT3 inhibitor Stattic. Collectively, our data suggested a novel mechanism that the inhibition of JAK2/STAT3 activation was responsible for palmitic lipotoxicity and SOV may act as a potential therapeutic agent by targeting JAK2/STAT3 in lipotoxic cardiomyopathy treatment.

Background: Erythropoietin (EPO) can reduce myocardial ischemia/reperfusion (I/R) injury. However, the cellular mechanisms have not been elucidated entirely. The present study was to investigate whether transcription factor GATA-4 could be involved in EPO-induced cardioprotection when it was administered after ischemia, immediately before reperfusion. Methods and results: Male Balb/c mice treated with or without EPO were subjected to ischemia (45 min) followed by reperfusion (4 h). TTC staining showed that the infarct size in EPO-treated mice was significantly reduced compared with untreated I/R mice (P < 0.05). Echocardiography examination suggested that EPO administration significantly improved cardiac function following I/R. TUNEL assay indicated that EPO treatment decreased apoptosis. EPO administration also significantly increased the level of nuclear GATA-4 phosphorylation in the myocardium which was positively correlated with the reduction of myocardial infarction. In vitro hypoxia/re-oxygenation study showed that EPO treatment increased the levels of phospho-GATA-4 and decreased cardiomyocyte apoptosis. More significantly, blocking GATA-4 by transfection of a dominant-negative form of GATA-4 (dnGATA-4) abolished EPO-induced cardioprotective effects. Conclusion: EPO administration after ischemia, just before reperfusion induced cardioprotection and stimulated GATA-4 phosphorylation. Activation of GATA-4 may be one of the mechanisms by which EPO induced protection against myocardial I/R injury.

Objective Plasma fibrinogen has been defined as a risk factor of cardiovascular disease and may play a role in the development of cardiac hypertrophy. We have previously demonstrated that the Toll-like receptor 4 (TLR4)-mediated myeloid differentiation primary response protein 88 (MyD88)-dependent nuclear factor-κB (NF-κB) pathway is involved in cardiac hypertrophy. The present study aimed to investigate whether fibrinogen will stimulate the hypertrophic response of cardiac myocytes and to examine the role of the TLR4/MyD88/NF-κB pathway in fibrinogen-induced cardiac hypertrophy. Methods and Results Cardiac hypertrophy was induced by transverse aortic banding for 5 weeks in Sprague-Dawley rats. The deposition of fibrinogen in the left ventricle, as determined by immunohistochemistry and immunoblotting, was increased. Aortic banding also significantly enhanced the association of TLR4 with MyD88 and increased NF-κB activity. In-vitro studies showed that fibrinogen induced a dose-dependent, hypertrophic response of neonatal cardiomyocytes. Fibrinogen stimulation significantly increased myocyte size, 3H-leucine incorporation and mRNA levels of atrial natriuretic peptide (ANP); fibrinogen challenge also significantly increased associations of TLR4 with MyD88 and NF-κB binding activity. Transient transfection of cardiomyocytes with a dominant-negative MyD88 plasmid significantly attenuated the fibrinogen-induced hypertrophic response of neonatal cardiac myocytes and blunted fibrinogen-increased activation of the TLR4/MyD88/NF-κB signaling pathway. Conclusion Our results suggest that fibrinogen induces hypertrophic response of cardiomyocytes partially through a TLR4-mediated, MyD88-dependent NF-κB pathway.

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No description available.

Regulation of striated muscle differentiation and development are complex processes coordinated by an array of transcription factors. MEF2 is a crucial transcription factor required for muscle differentiation, but the roles of the individual MEF2 family members, MEF2A-D, have not been extensively evaluated. Acute ablation of Mef2 expression in skeletal myoblasts revealed a required role for MEF2A activity in myoblast differentiation that was not shared with the other MEF2 factors. We hypothesized that a transcriptomic level analysis of Mef2-deficient skeletal myoblasts would reveal distinct regulatory roles for each MEF2 isoform. Comparative microarray analysis supported our hypothesis and we observed distinct gene programs preferentially-sensitive to individual MEF2 isoforms. While there was no variance in the consensus binding site associated with regulation by individual MEF2 isoforms, we did observe uniquely enriched binding sites for candidate co-regulatory proteins that mediate these complex regulatory patterns. Based on our observations in skeletal myoblasts, we performed a series of acute Mef2 knockdowns in neonatal cardiomyocytes and uncovered a requirement for MEF2A and -D, but not MEF2C in cardiomyocyte survival. Comparative microarray analysis confirmed that, similar to skeletal myoblasts, the MEF2 family regulated distinct but overlapping gene programs in cardiomyocytes. Additionally, this analysis uncovered a previously uncharacterized antagonistic regulation of a subset of cell cycle and sarcomere genes. Interestingly, Mef2a and -d knockdowns caused an upregulation of cell cycle markers and downregulation of sarcomere genes, with the opposite regulatory pattern in Mef2c knockdown. Further investigation of the proximal promoter region of these genes revealed enriched binding sites for transcription factors associated with key signaling pathways in the developing embryo, Hedgehog and Notch. Overexpression of constitutively active components of these signaling pathways revealed that Notch function requires the presence of MEF2A and -D, while Hedgehog does not appear to interact with these two isoforms. We have shown through our studies that MEF2, a core muscle transcription factor, takes part in complex regulatory interactions that are critical for the appropriate development of striated muscle tissues. / 2018-08-09T00:00:00Z

L’ischémie/reperfusion (I/R) induit des lésions impliquées dans la physiopathologie de la transplantation cardiaque où elles contribuent à augmenter le rejet de greffe. Le stress induit par l’ischémie entraîne la libération d’ATP conduisant à l’activation de récepteurs purinergiques (P2R) dont l’expression est établie au niveau cardiaque et immunitaire. L’objectif de ce travail a été d’explorer l’effet de la signalisation P2R sur le phénotype des cellules dendritiques (DCs) et la réponse des cardiomyocytes (CM) à l’I/R. Nous avons montré que la récepteur P2Y11 (P2Y11R) avait une action immunomodulatrice sur les DCs en diminuant la sécrétion d’IL-6 et IL-12 et en inhibant la polarisation de la réponse adaptative vers Th1. Le post-conditionnement pharmacologique ciblant P2Y11R a apporté une protection efficace sur les CM en limitant le stress oxydant et en activant la PKCe connue pour inhiber l’ouverture du mPTP. Les effets protecteurs et immunomodulateurs de P2Y11R se sont confirmés in vivo en diminuant le rejet allogénique dans un modèle murin de transplantation cardiaque hétérotopique. Nos résultats suggèrent que P2Y11R pourrait être une cible thérapeutique apportant des effets bénéfiques en transplantation cardiaque. / Ischemia/reperfusion (I/R) injuries are involved in the pathophysiology of heart transplantation where they will increase graft rejection. Ischemia generates cellular stress leading to ATP release in the extracellular medium that may activate purinergic receptors (P2R) expressed by cardiomyocytes and immune cells. Therefore, these receptors may play important regulatory roles. The aim of this study was to investigate the effect of P2R signaling on dendritic cells phenotype (DCs) and cardiomyocyte (CM) response to I/R. We showed that P2Y11 receptor (P2Y11R) exhibited an immunomodulatory role in DCs by decreasing release of IL-6 and IL-12 and inhibiting polarization of the adaptive response towards Th1. Pharmacological post-conditioning targeting P2Y11R provided effective protection to CM by limiting oxidative stress and activating PKCe known to inhibit the opening of the mPTP. The protective and immunomodulatory effects of P2Y11R stimulation were confirmed in vivo by the decrease of allogeneic acute rejection in a murine model of heterotopic heart transplantation. In conclusion, our results strongly suggest that P2Y11R may be a promising therapeutic target providing beneficial effects in cardiac transplantation.