Canagliflozin mediates mitophagy through the AMPK/PINK1/PARKIN pathway to alleviate isoprenaline-induced cardiac remodelling.

Heart failure has always been a prevalent, disabling, and potentially life-threatening disease. For the treatment of heart failure, controlling cardiac remodeling is very important. In recent years, clinical trials have shown that SGLT-2 inhibitors not only excel in lowering glucose levels but also demonstrate favorable cardiovascular protective effects. However, the precise mechanisms behind the cardiovascular benefits of SGLT-2 inhibitors remain elusive. In our current research, we assessed the impact of canagliflozin (CANA, an SGLT-2 inhibitor) on cardiac remodeling progression in mice, and preliminarily elucidated the possible mechanism of action of SGLT-2 inhibitor. Our results indicate that the administration of canagliflozin significantly attenuates myocardial hypertrophy and fibrosis and enhances cardiac ejection function in mice with isoprenaline (ISO)-induced cardiac remodeling. Notably, excessive mitophagy, along with mitochondrial structural abnormalities observed in ISO-induced cardiac remodeling, were mitigated by canagliflozin treatment, thereby attenuating cardiac remodeling progression. Furthermore, the differential expression of AMPK/PINK1/Parkin pathway-related proteins in ISO-induced cardiac remodeling was effectively reversed by canagliflozin, suggesting the therapeutic potential of targeting this pathway with the drug. Thus, our study indicates that canagliflozin holds promise in mitigating cardiac injury, enhancing cardiac function, and potentially exerting cardioprotective effects by modulating mitochondrial function and mitophagy through the AMPK/PINK1/Parkin pathway.

Fibrillatory wave amplitude on transesophageal ECG as a marker of left atrial low-voltage areas in patients with persistent atrial fibrillation.

BACKGROUND: Low-voltage areas (LVAs) are frequently observed in patients with persistent atrial fibrillation (PeAF) and may represent adverse atrial remodeling. However, noninvasive method of evaluating LAVs is not well established. METHODS: In a cohort of 68 patients with PeAF, endocardial voltage maps of left atrium (LA) were created during sinus rhythm after pulmonary vein isolation (PVI). LVAs were defined as areas with electrogram amplitudes <0.5 mV. LA-LVAs were correlated with clinical, echocardiographic, surface, and transesophageal electrocardiography (TE-ECG) variables. RESULTS: LA voltage mapping revealed any degree of LA-LVAs in 50 (73.5%) patients. Patients with LA-LVAs were older, had a longer history of AF, and lower fibrillatory wave (F wave) amplitude on TE-ECG (0.27 ± 0.06 vs 0.39 ± 0.08 mv, p < .01) as compared to patients without LA-LVAs. The extent of LA-LVAs was weakly correlated with age (R = 0.36, p = .03) and AF duration (R = 0.26, p = .02), but significantly correlated with F-wave amplitude on TE-ECG (R = -0.57, p < .01). Only F-wave amplitude on TE-ECG was found as independent predictor for the presence of LA-LVAs (OR = 1.53, 95% CI = 1.09-2.96, p = .03). A receiver operating characteristic (ROC) curve identified an F-wave amplitude of 0.29 mV (AUC = 0.788; sensitivity = 68.4%; specificity = 73.2%) on TE-ECG as the optimal cutoff value for predicting LA-LVAs. CONCLUSIONS: As a noninvasive investigation, F-wave amplitude on TE-ECG may be used as an indicator for the presence of LA-LVAs.

Predictive value of exercise-induced atrial natriuretic peptide secretion for the presence of left atrial low-voltage areas in patients with persistent atrial fibrillation.

Objectives Left atrial (LA) low-voltage areas (LVAs) are a strong predictor of atrial fibrillation (AF) recurrence after pulmonary vein isolation (PVI). However, a non-invasive method for evaluating LA-LAVs has not been established yet. The objective of our study was to assess the predictive value of the plasma atrial natriuretic peptide (ANP) level for the presence of LA-LVAs in patients with persistent AF (PeAF). Methods Seventy-two PeAF patients underwent an exercise stress test preprocedurally. LA voltage maps were created after PVI. Demographic, clinical and echocardiographic data were recorded. Plasma levels of ANP at baseline (ANP0) and increase induced by exercise (ΔANP) were also measured. Results Compared with patients without LA-LVAs, patients with LA-LVAs had a longer history of AF, higher CHADS2 score and higher ANP0 and lower ΔANP. LA-LVAs extent correlated with duration of AF history, CHADS2 score and ΔANP (R = -0.76, P < 0.01). Only ΔANP independently predicted the presence of LA-LVAs (OR =1.63, P = 0.02). Derived from the ROC curve, ΔANP <55 pg/mL predicted the presence of LA-LVAs with high accuracy (AUC =0.78; 95% CI =0.57-0.87, P < 0.01). Conclusions Exercise-induced secretion of ANP may be used to predict the presence of LA-LVAs in patients with PeAF before catheter ablation.

ACTN3 R577X Gene Variant Is Associated With Muscle-Related Phenotypes in Elite Chinese Sprint/Power Athletes.

Yang, R, Shen, X, Wang, Y, Voisin, S, Cai, G, Fu, Y, Xu, W, Eynon, N, Bishop, DJ, and Yan, X. ACTN3 R577X gene variant is associated with muscle-related phenotypes in elite Chinese sprint/power athletes. J Strength Cond Res 31(4): 1107-1115, 2017-The ACTN3 R577X polymorphism (rs1815739) has been shown to influence athletic performance. The aim of this study was to investigate the prevalence of this polymorphism in elite Chinese track and field athletes, and to explore its effects on athletes' level of competition and lower-extremity power. We compared the ACTN3 R577X genotypes and allele frequencies in 59 elite sprint/power athletes, 44 elite endurance athletes, and 50 healthy controls from Chinese Han origin. We then subcategorized the athletes into international level and national level and investigated the effects of ACTN3 genotype on lower-extremity power. Genotype distribution of the sprint/power athletes was significantly different from endurance athletes (p = 0.001) and controls (p < 0.001). The frequency of the RR genotype was significantly higher in international-level than that in the national-level sprint/power athletes (p = 0.004), with no international-level sprint/power athletes with XX genotype. The best standing long jump and standing vertical jump results of sprint/power athletes were better in the RR than those in the RX + XX genotypes (p = 0.004 and p = 0.001, respectively). In conclusion, the ACTN3 R577X polymorphism influences the level of competition and lower-extremity power of elite Chinese sprint/power athletes. Including relevant phenotypes such as muscle performance in future studies is important to further understand the effects of gene variants on elite athletic performance.

Angiotensin II-induced calcium overload affects mitochondrial functions in cardiac hypertrophy by targeting the USP2/MFN2 axis.

Ubiquitination, a common type of post-translational modification, is known to affect various diseases, including cardiac hypertrophy. Ubiquitin-specific peptidase 2 (USP2) plays a crucial role in regulating cell functions, but its role in cardiac functions remains elusive. The present study aims to investigate the mechanism of USP2 in cardiac hypertrophy. Animal and cell models of cardiac hypertrophy were established using Angiotensin II (Ang II) induction. Our experiments revealed that Ang II induced USP2 downregulation in the in vitro and in vivo models. USP2 overexpression suppressed the degree of cardiac hypertrophy (decreased ANP, BNP, and ß-MHC mRNA levels, cell surface area, and ratio of protein/DNA), calcium overload (decreased Ca2+ concentration and t-CaMKⅡ and p-CaMKⅡ, and increased SERCA2), and mitochondrial dysfunction (decreased MDA and ROS and increased MFN1, ATP, MMP, and complex Ⅰ and II) both in vitro and in vivo. Mechanically, USP2 interacted with MFN2 and improved the protein level of MFN2 through deubiquitination. Rescue experiments confirmed that MFN2 downregulation neutralized the protective role of USP2 overexpression in cardiac hypertrophy. Overall, our findings suggested that USP2 overexpression mediated deubiquitination to upregulate MFN2, thus alleviating calcium overload-induced mitochondrial dysfunction and cardiac hypertrophy.

Single-molecule imaging revealed enhanced dimerization of transforming growth factor ß type II receptors in hypertrophic cardiomyocytes.

Transforming growth factor ß (TGF-ß) signaling plays an important role in the pathogenesis of cardiac hypertrophy. However, the molecular mechanism of TGF-ß signaling during the process of cardiac remodeling remains poorly understood. In the present study, by employing single-molecule fluorescence imaging approach, we demonstrated that in neonatal rat cardiomyocytes, TGF-ß type II receptors (TßRII) existed as monomers at the low expression level, and dimerized upon TGF-ß1 stimulation. Importantly, for the first time, we found the increased dimerization of TßRII in hypertrophic cardiomyocytes comparing to the normal cardiomyocytes. The enhanced TßRII dimerization was correlated with the enhanced Smad3 phosphorylation levels. These results provide new information on the mechanism of TGF-ß signaling in cardiac remodeling.

Metformin attenuates pressure overload-induced cardiac hypertrophy via AMPK activation.

AIM: To identify the role of metformin in cardiac hypertrophy and investigate the possible mechanism underlying this effect. METHODS: Wild type and AMPKα2 knockout (AMPKα2⁻/⁻) littermates were subjected to left ventricular pressure overload caused by transverse aortic constriction. After administration of metformin (200 mg·kg⁻¹·d⁻¹) for 6 weeks, the degree of cardiac hypertrophy was evaluated using echocardiography and anatomic and histological methods. The antihypertrophic mechanism of metformin was analyzed using Western blotting. RESULTS: Metformin significantly attenuated cardiac hypertrophy induced by pressure overload in wild type mice, but the antihypertrophic actions of metformin were ablated in AMPKα2⁻/⁻ mice. Furthermore, metformin suppressed the phosphorylation of Akt/protein kinase B (AKT) and mammalian target of rapamycin (mTOR) in response to pressure overload in wild type mice, but not in AMPKα2⁻/⁻ mice. CONCLUSION: Long-term administration of metformin may attenuate cardiac hypertrophy induced by pressure overload in nondiabetic mice, and this attenuation is highly dependent on AMPK activation. These findings may provide a potential therapy for patients at risk of developing pathological cardiac hypertrophy.

Knockdown of dual oxidase 1 suppresses activin A-induced fibrosis in cardiomyocytes via the reactive oxygen species-dependent pyroptotic pathway.

Fibrotic diseases account for more than 8 million deaths worldwide annually. Reactive oxygen species (ROS) has been shown to activate pyroptosis and promote the production of interleukin (IL)-1ß and IL-18, leading to fibrosis development. However, the role of dual oxidase 1 (DUOX1)-induced ROS production and pyroptosis in cardiac fibrosis remains largely unknown. Activin A was used to induce ROS and pyroptosis in cardiomyocytes. ROS level, pyroptosis, and cytokine production were detected using Active Oxygen Detection Kit, flow cytometry, and enzyme-linked immunosorbent assay, respectively. Western blotting analysis was used to measure expression changes of proteins. DUOX1 was silenced or overexpressed to investigate its role in fibrosis. We found that activin A induced ROS production and pyroptosis in cardiomyocytes, which was blocked by the ROS scavenger, N-acetyl-L-cysteine (NAC). Knockdown of DUOX1 reversed activin A-induced ROS production, pyroptosis, cytokine release, and the upregulation of proinflammatory proteins. Overexpression of DUOX1 resulted in opposite effects of knockdown DUOX1. Administration of an ROS scavenger blocked the effect of DUOX1 overexpression. Supplementation of IL-1ß and IL-18 caused significant fibrosis in human cardiac fibroblasts (hCFs). The knockdown of DUOX1 protected cardiomyocytes against activin A-induced fibrosis via the inhibition of ROS, cytokine release, and pyroptosis.

Metformin reduces proteinuria in spontaneously hypertensive rats by activating the HIF-2α-VEGF-A pathway.

Metformin has protective effects on diabetic nephropathy. However, the mechanism underlying the renoprotective action of metformin in spontaneously hypertensive rats (SHR) is not completely understood. We determined the role of metformin in proteinuria and investigated the mechanism. We measured the urinary protein concentration (mg/day) in 48-week-old SHR. Matched control animals were of the same genetic strain, Wistar-Kyoto (WKY). The rats received metformin (100 mg/kg/day) or vehicle for 10 months. Metformin improved renal function and reduced the proteinuria (urine protein: 48.4 ± 3.7 vs. 25.4 ± 1.8 mg, P < 0.01) induced by long-term high blood pressure. Metformin increased the production of vascular endothelial growth factor (VEGF)-A in rat kidneys and cultured rat podocytes. Metformin activated hypoxia-inducible factor-2α (Hif-2α) in response to VEGF but did not affect Hif-1α in rat kidneys and cultured rat podocytes. Metformin reduced the proteinuria induced by long-term high blood pressure in vivo and increased the VEGF-A production in rat kidneys and cultured rat podocytes, probably by activating the Hif-2α-VEGF signaling pathway. These findings provide a new mechanism for the renoprotective effects of metformin.

The carboxyl terminus of heat shock protein 70-interacting protein (CHIP) participates in high glucose-induced cardiac injury.

The carboxyl terminus of heat shock protein 70-interacting protein (CHIP) is confirmed to have a protective effect on the myocardium, but its effect on diabetic cardiomyopathy is unclear. Small interfering RNA (siRNA) was used for knockdown experiments in neonatal rat cardiomyocytes to examine the function of CHIP in high glucose-induced injury. High glucose stimulated the production of reactive oxygen species (ROS), nicotinamide adenine dinucleotide phosphate oxidase (NOX), interleukin-1ß (IL-1ß), interleukin-6 (IL-6), and monocyte chemoattractant protein-1 (MCP-1) production. However, cardiomyocytes lacking CHIP suffered from increased oxidative stress and inflammatory responses. High glucose increased the expression of Bax and caspase-3 mRNAs, decreased the expression of Bcl-2 mRNA, and up-regulated the expression of the nuclear factor kappa B (NF-κB) and mitogen-activated protein kinase (MAPK) proteins. However, upon CHIP knockdown, the expression of Bax and caspase-3 mRNAs increased even further, and the expression of Bcl-2 mRNA was further suppressed. The expression of the phosphorylated p65 and p38 proteins (p-p65 and p-p38) was also further enhanced. Thus, CHIP is a potent cardioprotective molecule.

Molecular mechanisms in microRNA-mediated TRB3 gene and hypertension left ventricular hypertrophy.

The present study investigated the relationship between microRNA-mediated TRB3 gene and hypertension left ventricular hypertrophy at the molecular level. Polymorphic site in TRB3 gene was identified by direct PCR method, and the correlation between the SNP site and ventricular hypertrophy was determined. MicroRNAs target gene sequence interacting on the TRB3 polymorphic site was screened by bioinformatics, and the effect of microRNAs on the TRB3 polymorphic site was finally verified by luciferase test. Two polymorphic sites rs6186912 and rs6186923 were found in the TRB3 gene, and the direct relationship between rs6186923 polymorphic site and the hypertension left ventricular hypertrophy in patients with myocardial hypertrophy was compared and analyzed. Pictar software was used to analyze the effect of miR-100 on rs6186923, and the argumentation was verified by luciferase test. In conclusion, the study showed that the TRB3 gene polymorphism rs6186923 was able to affect the TRB3 gene by affecting the binding of miR-100, which indirectly caused the formation of hypertension left ventricular hypertrophy.

Cardiac Fibrosis Alleviated by Exercise Training Is AMPK-Dependent.

Regular exercise can protect the heart against external stimuli, but the mechanism is not well understood. We determined the role of adenosine monophosphate-activated protein kinase (AMPK) in regulating swimming exercise-mediated cardiac protection against ß-adrenergic receptor overstimulation with isoproterenol (ISO) in mice. Ten-week-old AMPKα2+/+ and AMPKα2-knockout (AMPKα2-/-) littermates were subjected to 4 weeks of swimming training (50 min daily, 6 days a week) or housed under sedentary conditions. The mice received daily subcutaneous injection of ISO (5 mg/kg/d), a nonselective ß-adrenergic receptor agonist, during the last 2 weeks of swimming training. Swimming training alleviated ISO-induced cardiac fibrosis in AMPKα2+/+ mice but not AMPKα2-/- mice. Swimming training activated cardiac AMPK in AMPKα2+/+ mice. Furthermore, swimming training attenuated ISO-induced production of reactive oxygen species (ROS) and expression of NADPH oxidase and promoted the expression of antioxidant enzymes in AMPKα2+/+ mice but not AMPKα2-/- mice. In conclusion, swimming training attenuates ISO-induced cardiac fibrosis by inhibiting the NADPH oxidase-ROS pathway mediated by AMPK activation. Our findings provide a new mechanism for the cardioprotective effects of exercise.

Beta-adrenoceptor signaling pathways mediate cardiac pathological remodeling.

Beta-adrenoceptors (ARs), members of the G protein-coupled receptor (GPCR) superfamily, play a key role in the rapid regulation of myocardial function. Meanwhile, chronic catecholamine stimulation of adrenoceptors has been proved to be involved in the adverse myocardial remodeling, including cardiac hypertrophy, fibrosis, and apoptosis, which finally develop into heart failure. In the clinical situation, sympathetic hyperactivity is a key factor in the development of heart failure, and beta-blockers greatly improve the outcome of the disease. However, heart failure is still one of the leading causes of death. Therefore, a full understanding of the mechanism of beta-AR-mediated cardiac remodeling could indicate more targets for treating heart failure. This review summarizes a number of important signaling pathways involved in the process of cardiac pathological remodeling under chronic adrenergic stimulation.

Distinct actions of intermittent and sustained ß-adrenoceptor stimulation on cardiac remodeling.

Heart disease is associated with increased sympathetic nerve activity and elevated levels of circulating catecholamines, resulting in chronic stimulation of the ß-adrenergic receptors (ß-AR) and consequent pathological cardiac remodeling. Experimentally, chronic administration of the ß-AR agonist isoproterenol (ISO) has been most commonly used to model ß-AR-induced cardiac remodeling. However, it remains unclear whether ß-AR-mediated cardiac remodeling and dysfunction differs between sustained versus pulsatile (intermittent) exposure to a ß-agonist. Here, we compare the effects of intermittent versus sustained administration of ISO on cardiac remodeling and function in mice. Animals were administered 5 mg (kg d)(-1) ISO for 2 weeks either by daily subcutaneous injection, or continuous infusion via an implanted osmotic minipump. Cardiac function and remodeling were determined by echocardiography, micromanometry and histology. Moreover, Western blotting and quantitative real-time polymerase chain reaction (qRT-PCR) were utilized to define the proteins and genes involved. Both sustained and intermittent administration of ISO resulted in a similar degree of cardiac hypertrophy (16% and 19%, respectively). However, mice receiving ISO by daily injection developed more severe ventricular systolic and diastolic dysfunction and myocardial fibrosis compared with mice receiving ISO via the osmotic minipump. The disparity in results between the delivery methods is suggested to be due, at least in part, to increased expression of fibrogenic factors, including connective tissue growth factor (CTGF) and NADPH oxidase (NOX4), in mice receiving intermittent application of ISO. In summary, compared with sustained exposure to a ß-AR agonist, intermittent ß-AR stimulation leads to more severe cardiac dysfunction and fibrosis. These findings not only further our understanding of ß-AR function in the setting of cardiac pathophysiology, but also highlight that significant differences can result dependent upon the mode of experimental ß-AR stimulation in inducing cardiomyopathy.

Metformin attenuates cardiac fibrosis by inhibiting the TGFbeta1-Smad3 signalling pathway.

AIMS: The mechanism of the cardioprotective action of metformin is incompletely understood. We determined the role of metformin in cardiac fibrosis and investigated the mechanism. METHODS AND RESULTS: Ten-week-old male mice (C57BL/6) were subjected to left ventricular pressure overload by transverse aortic constriction. Mice received metformin (200 mg/kg/day) or normal saline for 6 weeks. Metformin inhibited cardiac fibrosis (fibrosis area/total heart area: 0.6 +/- 0.3 vs. 3.6 +/- 0.9%, P < 0.01) induced by pressure overload and improved cardiac diastolic function (left ventricular end-diastolic pressure: 5.2 +/- 0.9 vs. 11.0 +/- 1.6 mmHg, P < 0.05). Metformin inhibited the pressure overload-induced transforming growth factor (TGF)-beta(1) production in mouse hearts and the TGF-beta(1)-induced collagen synthesis in cultured adult mouse cardiac fibroblasts (CFs). Metformin suppressed the phosphorylation of Smad3 in response to TGF-beta(1) in CFs. Metformin also inhibited the nuclear translocation and transcriptional activity of Smad3 in CFs. CONCLUSION: Metformin inhibited cardiac fibrosis induced by pressure overload in vivo and inhibited collagen synthesis in CFs probably via inhibition of the TGF-beta(1)-Smad3 signalling pathway. These findings provide a new mechanism for the cardioprotective effects of metformin.