Obesity is increasing in prevalence and is strongly associated with metabolic and cardiovascular disorders. The renin-angiotensin system (RAS) has emerged as a key pathogenic mechanism for these disorders; angiotensin (Ang)-converting enzyme 2 (ACE2) negatively regulates RAS by metabolizing Ang II into Ang 1-7. We studied the role of ACE2 in obesity-mediated cardiac dysfunction. ACE2 null (ACE2KO) and wild-type (WT) mice were fed a high-fat diet (HFD) or a control diet and studied at 6 months of age. Loss of ACE2 resulted in decreased weight gain but increased glucose intolerance, epicardial adipose tissue (EAT) inflammation, and polarization of macrophages into a proinflammatory phenotype in response to HFD. Similarly, human EAT in patients with obesity and heart failure displayed a proinflammatory macrophage phenotype. Exacerbated EAT inflammation in ACE2KO-HFD mice was associated with decreased myocardial adiponectin, decreased phosphorylation of AMPK, increased cardiac steatosis and lipotoxicity, and myocardial insulin resistance, which worsened heart function. Ang 1-7 (24 µg/kg/h) administered to ACE2KO-HFD mice resulted in ameliorated EAT inflammation and reduced cardiac steatosis and lipotoxicity, resulting in normalization of heart failure. In conclusion, ACE2 plays a novel role in heart disease associated with obesity wherein ACE2 negatively regulates obesity-induced EAT inflammation and cardiac insulin resistance.
Background Identifying new therapeutic targets for preventing the myocardial ischemia-reperfusion injury would have profound implications in cardiovascular medicine. Myocardial ischemia-reperfusion injury remains a major clinical burden in patients with coronary artery disease. Methods and Results We studied several key mechanistic pathways known to mediate cardioprotection in myocardial ischemia-reperfusion in 2 independent genetic models with reduced cardiac phosphoinositide 3-kinase-α (PI3Kα) activity. P3Kα-deficient genetic models (PI3KαDN and PI3Kα-Mer-Cre-Mer) showed profound resistance to myocardial ischemia-reperfusion injury. In an ex vivo reperfusion protocol, PI3Kα-deficient hearts had an 80% recovery of function compared with ≈10% recovery in the wild-type. Using an in vivo reperfusion protocol, PI3Kα-deficient hearts showed a 40% reduction in infarct size compared with wild-type hearts. Lack of PI3Kα increased late Na+ current, generating an influx of Na+, facilitating the lowering of mitochondrial Ca2+, thereby maintaining mitochondrial membrane potential and oxidative phosphorylation. Consistent with these functional differences, mitochondrial structure in PI3Kα-deficient hearts was preserved following ischemia-reperfusion injury. Computer modeling predicted that PIP3, the product of PI3Kα action, can interact with the murine and human NaV1.5 channels binding to the hydrophobic pocket below the selectivity filter and occluding the channel. Conclusions Loss of PI3Kα protects from global ischemic-reperfusion injury linked to improved mitochondrial structure and function associated with increased late Na+ current. Our results strongly support enhancement of mitochondrial function as a therapeutic strategy to minimize ischemia-reperfusion injury.
Coronary artery disease leading to myocardial ischemia is the most common cause of heart failure. Apelin (APLN), the endogenous peptide ligand of the APJ receptor, has emerged as a novel regulator of the cardiovascular system.Here we show a critical role of APLN in myocardial infarction (MI) and ischemia-reperfusion (IR) injury in patients and animal models. Myocardial APLN levels were reduced in patients with ischemic heart failure. Loss of APLN increased MI-related mortality, infarct size, and inflammation with drastic reductions in prosurvival pathways resulting in greater systolic dysfunction and heart failure. APLN deficiency decreased vascular sprouting, impaired sprouting of human endothelial progenitor cells, and compromised in vivo myocardial angiogenesis. Lack of APLN enhanced susceptibility to ischemic injury and compromised functional recovery following ex vivo and in vivo IR injury. We designed and synthesized two novel APLN analogues resistant to angiotensin converting enzyme 2 cleavage and identified one analogue, which mimicked the function of APLN, to be markedly protective against ex vivo and in vivo myocardial IR injury linked to greater activation of survival pathways and promotion of angiogenesis.APLN is a critical regulator of the myocardial response to infarction and ischemia and pharmacologically targeting this pathway is feasible and represents a new class of potential therapeutic agents.
Apelin peptides mediate beneficial effects on the cardiovascular system and are being targeted as potential new drugs. However, apelin peptides have extremely short biological half-lives, and improved understanding of apelin peptide metabolism may lead to the discovery of biologically stable analogues with therapeutic potential. We examined the ability of angiotensin-converting enzyme 2 (ACE2) to cleave and inactivate pyr-apelin 13 and apelin 17, the dominant apelin peptides. Computer-assisted modeling shows a conserved binding of pyr-apelin 13 and apelin 17 to the ACE2 catalytic site. In ACE2 knockout mice, hypotensive action of pyr-apelin 13 and apelin 17 was potentiated, with a corresponding greater elevation in plasma apelin levels. Similarly, pharmacological inhibition of ACE2 potentiated the vasodepressor action of apelin peptides. Biochemical analysis confirmed that recombinant human ACE2 can cleave pyr-apelin 13 and apelin 17 efficiently, and apelin peptides are degraded slower in ACE2-deficient plasma. The biological relevance of ACE2-mediated proteolytic processing of apelin peptides was further supported by the reduced potency of pyr-apelin 12 and apelin 16 on the activation of signaling pathways and nitric oxide production from endothelial cells. Importantly, although pyr-apelin 13 and apelin 17 rescued contractile function in a myocardial ischemia-reperfusion model, ACE2 cleavage products, pyr-apelin 12 and 16, were devoid of these cardioprotective effects. We designed and synthesized active apelin analogues that were resistant to ACE2-mediated degradation, thereby confirming that stable apelin analogues can be designed as potential drugs. We conclude that ACE2 represents a major negative regulator of apelin action in the vasculature and heart.
Introduction: PI3K inhibitors are a promising new therapy for cancer treatment now in clinical trials. However, loss of PI3K isoforms have negative consequences in some models of heart disease. Also, a genetic model with PI3Kα deletion using tamoxifen/Cre had reduced heart function, suggesting a role for PI3Kα in heart contractility; however, those findings may have been compounded by tamoxifen/Cre toxicity. Methods and Results: Cardiomyocyte specific deletion of PI3Kα was achieved with Mer-Cre-Mer (MCM) transgenic mice and tamoxifen administration (40 or 60 mg/kg/day*4days; LD (low dose), HD (high dose)) at 10-11weeks old to induce gene deletion, with function assessed by echocardiography. MCM mice developed systolic dysfunction and atrial dilation at 10 days after the start of HD tamoxifen treatment. Control hearts recovered at 28 days, but PI3Kα deletion hearts had sustained dysfunction and elevated expression of disease markers. Reduced tamoxifen treatment (LD) did not cause heart dysfunction or elevated disease markers but did cause increased ERK activation, as well as effective reduction of PI3Kα protein similar to HD tamoxifen. Deletion of PI3Kα using a constitutively active Cre (Cre) model was assessed for for heart function and insulin signaling as an indicator of PI3Kα activity. Cre PI3Kα knockouts had normal function and elevated ERK activation and blunted insulin signaling. Cardiomyocytes isolated from non-failing human donor hearts and adult mouse cardiomyocytes were treated with the PI3Kα inhibitor BYL-719 (100 nM). Insulin (10 mg/L) mediated Akt activation in both mouse and human cardiomyocytes was blocked by BYL-719. Mice treated with BYL-719 (30 mg/kg/day*2 wks) were assessed for heart function with echocardiography and invasive pressure-volume catheter as well as changes in insulin signaling. Treatment with BYL-719 in mice blocked insulin mediated Akt activation in the heart, caused weight loss and elevated fasting glucose levels, but did not affect heart function. Conclusions: PI3Kα is not essential for maintained function of the adult heart, but has an important role in metabolic signaling and is protective against Cre/tamoxifen toxicity. Our data justifies caution when using PI3Kα inhibitors in cancer patients.
Background: Biomechanical stress and cytoskeletal remodeling are key determinants in pressure overload-induced heart failure. Class Ia phosphoinositide 3-kinases (PI3Ks) mediate a variety of cellular activities, in response to agonist binding to cell-surface receptors, by generating the phosphatidylinositol (3,4,5)-trisphosphate (PIP 3 ) phosphoinositide lipid. Gelsolin is a Ca 2+ - and phosphoinositide-regulated actin filament severing and capping protein that is upregulated in failing human hearts and animal models of heart failure. Hypothesis: We hypothesize that PI3Kα regulates cytoskeletal remodeling through PIP 3 -mediated regulation of gelsolin. In addition, loss of gelsolin could attenuate the adverse cytoskeletal remodeling and result in increased resistance to the development of heart failure in response to pressure-overload. Methods and Results: Loss of p110α kinase activity, in two different transgenic models (PI3Kα dominant-negative (PI3KαDN) and cardiomyocyte-specific PI3Kα-null), resulted in dilated cardiomyopathy and markedly worsened cardiac dysfunction in response to transverse aortic constriction-induced pressure overload. Increased levels of mechanosensor proteins along with decreased F/G-actin ratio exhibited an uncoupling between cardiac mechanotransduction and cytoskeletal remodeling in p110α-null mice. Gelsolin activity was markedly increased in the p110α-null hearts in response to pressure-overload, whereas loss of gelsolin in PI3KαDN/gelsolin-null double mutant mice prevented the adverse cytoskeletal remodeling and preserved the cardiac function. In a murine model of chronic heart failure, loss of gelsolin prevented the pressure overload-induced cardiac dysfunction, fibrosis, and impaired cardiomyocyte contractility resulting in increased survival. Loss of gelsolin also mitigated the biomechanical stress-induced adverse cytoskeletal remodeling, via the attenuation of actin severing activity. Conclusions: We have identified a novel role of gelsolin as a mediator of adverse cytoskeletal remodeling leading to heart failure, where PI3Kα is a key regulator of gelsolin activity.
Iron-overload cardiomyopathy is a prevalent cause of heart failure on a world-wide basis and is a major cause of mortality and morbidity in patients with secondary iron-overload and genetic hemochromatosis. We investigated the therapeutic effects of resveratrol in acquired and genetic models of iron-overload cardiomyopathy. Murine iron-overload models showed cardiac iron-overload, increased oxidative stress, altered Ca(2+) homeostasis and myocardial fibrosis resulting in heart disease. Iron-overload increased nuclear and acetylated levels of FOXO1 with corresponding inverse changes in SIRT1 levels in the heart corrected by resveratrol therapy. Resveratrol, reduced the pathological remodeling and improved cardiac function in murine models of acquired and genetic iron-overload at varying stages of iron-overload. Echocardiography and hemodynamic analysis revealed a complete normalization of iron-overload mediated diastolic and systolic dysfunction in response to resveratrol therapy. Myocardial SERCA2a levels were reduced in iron-overloaded hearts and resveratrol therapy restored SERCA2a levels and corrected altered Ca(2+) homeostasis. Iron-mediated pro-oxidant and pro-fibrotic effects in human and murine cardiomyocytes and cardiofibroblasts were suppressed by resveratrol which correlated with reduction in iron-induced myocardial oxidative stress and myocardial fibrosis. Resveratrol represents a clinically and economically feasible therapeutic intervention to reduce the global burden from iron-overload cardiomyopathy at early and chronic stages of iron-overload.
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