Oxidative stress and mitochondrial dysfunction have major roles in the pathophysiology of cardiovascular disease. Mitochondria are the predominant source of energy production in the heart. Mitochondrial dysfunction decreases ATP production, while increasing reactive oxygen species (ROS) production. Early mitochondrial dysfunction leads to a vicious cycle, where excess ROS damages the mitochondria, leading to increased ROS production and impaired ATP production, decreasing the heart’s energetic capacity. Therefore, targeted mitochondrial antioxidants are a promising therapeutic target for cardiovascular disease and identification of novel endogenous mitochondrial antioxidants could lead to the discovery of new therapeutic targets. ABCB10 is a mitochondrial transporter with an unknown substrate. Our laboratory showed that hearts from mice with a heterozygous deletion of ABCB10 had decreased cardiac function and increased oxidative stress after ischemia reperfusion. Administration of an antioxidant ameliorated these effects, indicating that ABCB10 may have antioxidant effects. Therefore, we hypothesize that ABCB10 protects cardiac myocytes against oxidative injury through antioxidant effects in the cytosol and/or mitochondria. To test this hypothesis, we overexpressed ABCB10 in isolated adult rat ventricular myocytes (ARVMs) and exposed these cells to an oxidative challenge. ABCB10 overexpression protected cardiac myocytes against an oxidative insult, increasing cell viability and decreasing cytosolic ROS levels. ABCB10 overexpression also increased levels of HO-1, a cardioprotective and antioxidant-generating enzyme. We generated cardiac-specific ABCB10 knockout mice to explore the role of cardiac ABCB10 in vivo. We found that cardiac-specific deletion of ABCB10 in mice caused early death and mitochondrial dysfunction, evidenced by increased mitochondrial ROS production and decreased mitochondrial oxygen consumption. Additionally, microarray gene analysis revealed that hearts from ABCB10 knockout mice downregulated mitochondrial transcription and translation processes, indicating impaired mitochondrial proteostasis. These findings demonstrate that ABCB10 exerts an antioxidant effect, due, at least in part, to upregulation of the HO-1 antioxidant system. The ABCB10 mitochondrial transporter is involved in regulating mitochondrial oxidant levels and proteostasis and may be a novel therapeutic target in states associated with oxidative stress.

Background. This study examined the effect of microRNA-125b (miR-125b) on sepsis-induced cardiac dysfunction. Methods. Mouse hearts were transfected with lentivirus expressing miR-125b (LmiR-125b) 7 days before cecal ligation and puncture (CLP)-induced sepsis. Cardiac function was examined by echocardiography before and 6 hours after CLP (n = 6/group). Survival was monitored following CLP-induced sepsis (n = 12/group). Results. LmiR-125b transfection significantly attenuated cardiac dysfunction due to CLP-induced sepsis. Fractional shortening and ejection fraction values were significantly (P <.05) higher in the LmiR-125b-treated CLP group than in the untreated CLP group. Survival outcome in LmiR-125b-transfected septic mice was markedly improved, compared with mice with CLP-induced sepsis. Transfection of LmiR-125b into the heart significantly suppressed the expression of ICAM-1 and VCAM-1, decreased the accumulation of macrophages and neutrophils in the myocardium, and decreased serum levels of tumor necrosis factor a and interleukin 1β by targeting tumor necrosis factor receptor-associated factor 6 (TRAF6)-mediated nuclear factor κB (NF-κB) activation. In addition, sepsis-induced myocardial apoptosis was markedly attenuated by LmiR-125b transfection through suppression of p53, Bax, and Bak1 expression. In vitro transfection of endothelial cells with miR-125b mimics attenuate LPS-induced ICAM-1 and VCAM-1 expression by suppressing TRAF6 and NF-κB activation. Conclusions. Increased myocardial miR-125b expression attenuates sepsis-induced cardiac dysfunction and improves survival. miR-125b may be a target for septic cardiomyopathy.

Cardiac dysfunction is a major consequence that contributes to the high mortality of trauma-hemorrhage (TH) patients. Recent evidence suggests that innate immune and inflammatory responses mediated by Toll-like receptors (TLRs) play a critical role in the pathophysiologic mechanisms of acute organ dysfunction during TH. This study investigated the role of TLR4 in cardiac dysfunction following TH. Toll-like receptor 4-deficient (TLR4-/-, n = 7/group) and age-matched wild-type (WT, n = 8/group) mice were subjected to TH that was induced by soft tissue injury and blood withdrawal from the jugular vein to a mean arterial pressure of 35 ± 5 mmHg. Cardiac function and mean arterial pressure were measured with a Millar system before, during, and after blood withdrawal. Sham surgical-operated mice served as control (WT, n = 9/group; TLR4-/-, n = 10/group). Cardiac function in WT mice was significantly reduced following TH. However, cardiac function was well preserved in TLR4-/- mice. Administration of a TLR4 antagonist (3 mg/kg) to WT mice also significantly attenuated TH-induced cardiac dysfunction. Western blot showed that either TLR4-/- or TLR4 antagonist markedly attenuated TH-induced decreases in the levels of phosphorylated-Akt in myocardium. In addition, inhibition of TLR4 attenuated TH-induced myocardial nuclear factor κB-binding activity as well as lung myeloperoxidase activity and tumor necrosis factor α production. The data indicate that TLR4 plays a central role in TH-induced cardiac dysfunction. Toll-like receptor 4 deficiency or TLR4 inhibition attenuated cardiac dysfunction following TH, which may involve activation of the phosphoinositide 3-kinase/Akt signaling and decrease in nuclear factor κB-binding activity. Toll-like receptor 4 antagonism may be a new and novel approach for the treatment and management of cardiac dysfunction in TH patients.

Oxidative stress plays an important role in cardiac diseases, which has been well demonstrated, whereas the role of reductive stress has been poorly investigated. We and others have shown previously that heat shock protein 27 (Hsp27) plays a role as an antioxidant. To investigate whether overexpression of Hsp27 could lead to reductive stress and result in cardiomyopathy, we generated transgenic mice with different expression levels of Hsp27. We observed that transgenic mice with high levels of Hsp27 developed cardiomyopathy. The myopathic hearts were under reductive stress, which was evidenced by an increased ratio of reduced glutathione/oxidized glutathione and a decreased level of reactive oxygen species. In addition, upregulated glutathione peroxidase 1 and decreased iron content were revealed in the myopathic hearts. More importantly, inhibition of glutathione peroxidase 1 significantly attenuated the development of cardiomyopathy. The data indicate that the Hsp27-induced cardiomyopathy could be attributed to, at least in part, upregulation of glutathione peroxidase 1. Our findings suggest that reductive stress plays an important role in the development of cardiomyopathy and that Hsp27 may serve as a potential target for the treatment of patients with cardiomyopathy.

Indiana University-Purdue University Indianapolis (IUPUI) / Recent clinical outcome studies demonstrate that Sodium glucose cotransporter 2 inhibitors (SGLT2i) significantly reduce major adverse cardiovascular events and heart failure outcomes in subjects with type 2 diabetes mellitus. At present, several hypotheses have been proposed to explain the observed cardiovascular benefit of SGLT2i, however, the mechanisms responsible remain to be elucidated. This investigation tested the hypothesis that SGLT2i improves cardiac function and efficiency during acute, regional ischemia/reperfusion injury via preferential shifts in myocardial substrate selection and/or inhibition of cardiac sodium/hydrogen exchanger-1 (NHE-1). Our initial investigation evaluated the effects of 24 hour pretreatment of the SGLT2i canagliflozin on cardiac contractile function, substrate utilization, and efficiency before and during regional myocardial ischemia/reperfusion injury in healthy swine. At the onset of ischemia, canagliflozin increased left ventricular end diastolic and systolic volumes which returned to baseline with reperfusion. This increased end diastolic volume was directly associated with increased stroke volume and stroke work relative to controls during ischemia. Canagliflozin also increased cardiac work efficiency during ischemia relative to control swine. No differences in myocardial substrate uptake of glucose, lactate, fatty acids or ketones were detected between groups. In separate experiments using a longer 60 min coronary occlusion, canagliflozin significantly diminished myocardial infarct size. Subsequent studies investigated the effect of an acute administration (15-30 min pre-treatment) of canagliflozin and the NHE-1i cariporide on cardiac contractile function efficiency in response to myocardial ischemia/reperfusion injury. Similar to our initial studies, canagliflozin increased diastolic filling, stroke work and improved cardiac work efficiency relative to untreated control hearts during the ischemic period. In contrast, cariporide did not alter ventricular filling volume, cardiac output or work efficiency at any time point. Additional examination of AP-1 cells transfected with wild-type NHE-1 showed dose-dependent inhibition of NHE-1 activity by cariporide, while canagliflozin had minimal effect on overall activity. This investigation demonstrates that SGLT2i improves cardiac function and efficiency during acute, regional ischemia in healthy swine. However, the present data fail to support the hypothesis that these SGLT2i-mediated improvements involve either preferential alterations in myocardial substrate utilization or the inhibition of NHE-1 activity.

No description available.

Ischemic cardiomyopathies, such as myocardial infarction (MI), are a leading cause of heart failure in both men and women throughout the world. Despite timely intervention post-MI, the loss of viable myocardium can lead to global remodeling and loss of function in many patients due to the limited regenerative potential of heart tissue. Thus, there is a critical need to better understand the repair mechanisms involved and to develop new preventative and reparative therapies for treating MI and preventing progression to heart failure. Methylglyoxal (MG) is a highly reactive dicarbonyl metabolite of glycolysis and the main precursor of advanced glycation end-products (AGEs), which can cause oxidative stress and wound healing delay. MG was shown to play an important causative role in the cellular changes, adverse remodeling and functional loss of the infarcted heart. This suggests MG as a target for therapy to restore cell-ECM signaling, inhibit oxidative stress and improve cardiac function post-MI. The aim of this PhD project was to develop new biomaterial therapies that can reduce the effects of MG, decrease oxidative stress, enhance electrical conductivity and improve cardiac contractility and function post-MI. There were three primary objectives: 1) To develop an injectable antioxidant and hydrogel system for minimizing the effects of MG and promoting cardiac repair post-MI; 2) To synthesize a nanoparticle system for targeted delivery of Glyoxalase-1 (Glo1) enzyme to cardiac tissue for reducing the accumulation of MG, limiting adverse remodeling and preserving cardiac function following MI; and 3) To design a sprayable nano-therapeutic that uses surface engineered custom designed multi-armed peptide grafted nanogold for on-the-spot coating of infarcted myocardial surface for increasing contractility of the myocardium post-MI. In the first study, a fisetin-loaded collagen type I hydrogel (fisetin-HG) was injected intramyocardially in mice at 3h post-MI, and compared to fisetin-alone, hydrogel-alone, or saline treatment. The fisetin-HG treatment increased the level of glyoxalase-1 (the main MG-metabolizing enzyme), reduced MG-AGE accumulation, and decreased oxidative stress in the MI heart, which was associated with smaller scar size and improved cardiac function. Treatment with fisetin-HG also promoted neovascularization and increased the number of pro-healing macrophages in the infarct area, while reducing the number of pro-inflammatory macrophages. The second study revealed that when delivered intravenously at 3h post-MI, our Glo1-loaded nanoparticles specifically targeted the damaged cardiac tissue, led to improved cardiac function, protected cell viability and limited infarct expansion by reducing oxidative stress post-MI. Lastly, the third study showed that, when applied at 1-week post-MI, the sprayed nanogold treatment remained at the treatment site for at least 28 days with no significant off-target organ infiltration. Our results demonstrated a remarkable increase in cardiac function, muscle contractility, and myocardial electrical conductivity post-MI. Overall, these findings show that reducing MG levels through both increased activity of Glo1 and direct MG scavenging as well as increasing cardiac contractility may be a promising approach to limit adverse cardiac remodeling, prevent damage, and preserve the function of the infarcted heart

Honey bees are economically important pollinators of a wide variety of crops that have attracted the attention of both researchers and the public alike due to unusual declines in the numbers of managed colonies in some parts of the world. Viral infections are thought to be a significant factor contributing to these declines, along with exposure to agricultural and apicultural pesticides, but viruses have proven a challenging pathogen to study in a bee model and interactions between viruses and the bee antiviral immune response remain poorly understood. Recent studies have demonstrated an important role for inwardly-rectifying ATP-sensitive potassium (KATP) channels in the cardiac regulation of the fruit fly antiviral immune response, but no information is available on their role in the heart-specific regulation of bee immunity. The results of this work demonstrate that KATP channel modulators have an observable effect on honey bee heart rate that supports their expected physiological role in bee cardiac function. Here, it is also reported that the entomopathogenic flock house virus (FHV) infects adult bees, causing rapid onset of mortality and accumulation of viral RNA. Furthermore, infection-mediated mortality can be altered by pre-exposure to KATP channel modulators. Finally, this work shows that exposure to environmental stressors such as commonly used in-hive acaricides can impact bee cardiac physiology and tolerance to viral infection. These results suggest that KATP channels provide a significant link between cellular metabolism and the antiviral immune response in bees and highlight the significant impact of environmental stressors on pollinator health. / Ph. D.

Both hypertrophied and failing hearts are characterised by pathological left ventricular (LV) remodelling, impaired myocardial energy status and alteration in substrate metabolism. Cardiac magnetic resonance imaging (CMR) and magnetic resonance spectroscopy (MRS) are powerful tools in the characterisation of these disease conditions. More recent techniques have allowed assessment of myocardial steatosis using ¹H-MRS and tissue oxygenation using blood oxygen level dependent (BOLD) CMR. In hypertrophy and heart failure, studies on steatosis and the relationship with other parameters such as myocardial function and fibrosis, especially in humans are limited. I therefore investigated the presence of steatosis in severe aortic stenosis (AS) and dilated cardiomyopathy (DCM), and further assessed its relation to contractile function. This study found that myocardial triglyceride (TG) content is increased in both symptomatic and asymptomatic AS patients (lipid/water ratio 0.89±0.42% in symptomatic AS; 0.75±0.36% in asymptomatic AS vs. controls 0.45±0.17%, both p<0.05) and DCM patients (lipid/ratio 0.64±0.44% vs. controls 0.40±0.13%, p=0.03). Circumferential strain was lower in both AS (-16.4±2.5% in symptomatic AS; -18.9±2.9% in asymptomatic AS vs. controls 20.7±2.0%, both p<0.05) and DCM patients (-12.3±3.4% vs. controls -20.9±1.7%, p<0.001). In AS, myocardial contractility is related to the degree of steatosis, and were both reversible following aortic valve replacement (AVR), lipid/water ratio 0.92±0.41% vs. pre AVR 0.45±0.17%, p=0.04 and circumferential strain -17.2±2.0% vs. pre AVR -19.5±3.2%, p=0.04. A novel finding of this study was significant correlation of MRS-measured TG content with histological staining of TG of the myocardium, taken from endomyocardial biopsy during AVR. In DCM, myocardial TG was independently associated with LV dilatation and correlated significantly with hepatic TG, which suggests that both cardiac and hepatic steatosis might be a common feature in the failing heart. Additionally, although the hypertrophied heart is characterised by impaired perfusion, it is unknown if this is severe enough to translate into tissue deoxygenation and ischaemia. I assessed this by using adenosine vasodilator stress test and BOLD-CMR in patients with severe AS. It was found that AS patients had reduced perfusion (myocardial perfusion reserve index-MPRI 1.0±0.3 vs. controls 1.7±0.3, p<0.001), and blunted tissue oxygenation (blood-oxygen level dependent-BOLD signal intensity-SI change 4.8±9.6% vs. controls 18.2±11.6%, p=0.001) during stress. Importantly, there was a substantial improvement in perfusion and oxygenation towards normal after AVR, MPRI 1.5±0.4, p=0.005 vs. pre AVR and BOLD SI change 16.4±7.0%, p=0.014 vs. pre AVR. Overall, the work in this thesis supports the powerful role of CMR in assessing LV function and elucidating metabolic mechanisms in the hypertrophied and failing heart.

O presente estudo teve por objetivo investigar os efeitos do jejum de 48 horas sobre o metabolismo do músculo cardíaco e sobre a função cardíaca. Para isso, ratos machos com 60 dias foram separados em dois grupos: jejuados por 48 h (grupo experimental) e alimentados (grupo controle). Após eutanásia, sangue e coração foram coletados. O coração foi excisado e a aurícula do átrio direito (AAD), a aurícula do átrio esquerdo (AAE), parede do ventrículo direito (PVD), septo interventricular (SIV) e parede do ventrículo esquerdo (PVE) foram separadas e analisadas individualmente. Análises de parâmetros bioquímicos plasmáticos, dosagem de metabólitos, atividade máxima de enzimas, assim como expressão gênica e proteica foram realizadas. O jejum promoveu alterações metabólicas em todas as regiões, sendo mais intensas na PVE. Registros ventriculares e hemodinâmicos também foram obtidos. O jejum diminuiu a força de contração (dP/dt+), a força de relaxamento (dP/dt-) e a frequência cardíaca (FC), aumentou o tempo de enchimento diastólico e o hematócrito. Apesar de observamos aumento do potencial oxidativo e aumento da concentração disponível de ATP, é possível que 48h de jejum comprometa a volemia e por consequência a função cardíaca. / The present study aimed to investigate the effects of 48-hour fasting on cardiac muscle metabolism and cardiac function. For this, male rats with 60 days were separated into two groups: fasted for 48 h (experimental group) and fed (control group). After euthanasia, blood and heart were collected. The heart was excised and the right atrial atrium (RAA), left atrial atrium (LAA), right ventricular wall (RVW), interventricular septum (IVS) and left ventricular wall (LVW) were separated and analyzed individually. Analyzes of plasma biochemical parameters, dosage of metabolites, maximum activity of enzymes, as well as gene and protein expression were performed. Fasting promoted metabolic alterations in all regions, being more intense in PVE. Ventricular and hemodynamic records were also obtained. Fasting decreased contraction force (dP / dt +), relaxation force (dP / dt-) and heart rate (HR), increased diastolic filling time and hematocrit. Although we observed an increase in the oxidative potential and an increase in the available ATP concentration, it is possible that 48h-fasting compromises blood volume and, consequently, cardiac function.