Calmodulin kinase II inhibition protects against structural heart disease

Beta-adrenergic receptor (betaAR) stimulation increases cytosolic Ca(2+) to physiologically augment cardiac contraction, whereas excessive betaAR activation causes adverse cardiac remodeling, including myocardial hypertrophy, dilation and dysfunction, in individuals with myocardial infarction. The Ca(2+)-calmodulin-dependent protein kinase II (CaMKII) is a recently identified downstream element of the betaAR-initiated signaling cascade that is linked to pathological myocardial remodeling and to regulation of key proteins involved in cardiac excitation-contraction coupling. We developed a genetic mouse model of cardiac CaMKII inhibition to test the role of CaMKII in betaAR signaling in vivo. Here we show CaMKII inhibition substantially prevented maladaptive remodeling from excessive betaAR stimulation and myocardial infarction, and induced balanced changes in excitation-contraction coupling that preserved baseline and betaAR-stimulated physiological increases in cardiac function. These findings mark CaMKII as a determinant of clinically important heart disease phenotypes, and suggest CaMKII inhibition can be a highly selective approach for targeting adverse myocardial remodeling linked to betaAR signaling.     

Erythropoietin protects against doxorubicin-induced heart failure

The hormone erythropoietin (EPO) has been demonstrated to have cardioprotective properties. The present study investigates the role of EPO to prevent heart failure following cancer treatment with doxorubicin [adriamycin (AD)]. Male Wistar rats (150 ± 10 g) were treated with saline (vehicle control group); with EPO, subcutaneously at 1,000 IU/kg body wt, three times per week for 4 wk (EPO group); with adriamycin, intraperitoneally at 2.5 mg/kg body wt, three times per week for 2 wk (AD group); and with adriamycin and EPO (EPO-AD group). Echocardiographic measurements showed that EPO-AD treatment prevented the AD-induced decline in cardiac function. Each of the hearts was then exposed to ischemia and reperfusion during Langendorff perfusion. The percentage of recovery after ischemia-reperfusion was significantly greater in EPO-AD than the AD-treated group for left ventricular developed pressure, maximal increase in pressure, and rate pressure product. The level of oxidative stress was significantly higher in AD (5 μM for 24 h)-exposed isolated cardiomyocytes; EPO (5 U/ml for 48 h) treatment prevented this. EPO treatment also decreased AD-induced cardiomyocyte apoptosis, which was associated with the decrease in the Bax-to-Bcl2 ratio and caspase-3 activation. Immunostaining of myocardial tissue for CD31 showed a significant decrease in the number of capillaries in AD-treated animals. EPO-AD treatment restored the number of capillaries. In conclusion, EPO treatment effectively prevented AD-induced heart failure. The protective effect of EPO was associated with a decreased level of oxidative stress and apoptosis in cardiomyocytes as well as improved myocardial angiogenesis.     

p13 protects against Parkinson's disease

Mitochondrial dysfunction is thought to contribute to Parkinson's disease progression, and factors that can overcome mitochondrial defects could potentially be used to combat the disease and prevent neuronal death. In this issue, Inoue et al 1 report that reduction of p13, a mitochondrial protein that inhibits complex I assembly, rescues the cellular and behavioral defects of Parkinson's disease models. This work suggests that stabilizing the mitochondrial electron transport chain may be beneficial in the context of Parkinson's disease.     

Cardiac mTOR protects the heart against ischemia-reperfusion injury

Cardiac mammalian target of rapamycin (mTOR) is necessary and sufficient to prevent cardiac dysfunction in pathological hypertrophy. However, the role of cardiac mTOR in heart failure after ischemic injury remains undefined. To address this question, we used transgenic (Tg) mice with cardiac-specific overexpression of mTOR (mTOR-Tg mice) to study ischemia-reperfusion (I/R) injury in two animal models: 1) in vivo I/R injury with transient coronary artery ligation and 2) ex vivo I/R injury in Langendorff-perfused hearts with transient global ischemia. At 28 days after I/R, mortality was lower in mTOR-Tg mice than littermate control mice [wild-type (WT) mice]. Echocardiography and MRI demonstrated that global cardiac function in mTOR-Tg mice was preserved, whereas WT mice exhibited significant cardiac dysfunction. Masson's trichrome staining showed that 28 days after I/R, the area of interstitial fibrosis was smaller in mTOR-Tg mice compared with WT mice, suggesting that adverse left ventricular remodeling is inhibited in mTOR-Tg mice. In the ex vivo I/R model, mTOR-Tg hearts demonstrated improved functional recovery compared with WT hearts. Perfusion with Evans blue after ex vivo I/R yielded less staining in mTOR-Tg hearts than WT hearts, indicating that mTOR overexpression inhibited necrosis during I/R injury. Expression of proinflammatory cytokines, including IL-6 and TNF-α, in mTOR-Tg hearts was lower than in WT hearts. Consistent with this, IL-6 in the effluent post-I/R injury was lower in mTOR-Tg hearts than in WT hearts. These findings suggest that cardiac mTOR overexpression in the heart is sufficient to provide substantial cardioprotection against I/R injury and suppress the inflammatory response.     

Ethanol protects the heart against the calcium paradox injury

Rat hearts were depleted of Ca2+ (less than 10(-9) M) for 10 min, followed by 15 min of Ca2+-repletion. The calcium paradox injury occurs during Ca2+-repletion, after a period of calcium depletion. The calcium paradox injury was assessed by percent recovery (hemodynamics, [Ca2+]i, and energy levels) during Ca2+-repletion. A decrease in Na+ concentration during Ca2(+)-depletion did not allow for recovery during Ca2(+)-repletion, however 2.5% and 5% ethanol during Ca2(+)-depletion allowed for an approximate 50% recovery during Ca2(+)-repletion. A combination of ethanol (2.5% or 5%) with a low extracellular Na+ concentration (88 mM) allowed for complete recovery. Ethanol prevented a depletion of diastolic [Ca2+]i during Ca2(+)-depletion, and allowed for a return of normal diastolic [Ca2+]i during Ca2(+)-repletion. Ethanol modulates the activity of the Na+/Ca2+ exchanger and protects against the Ca2(+)-paradox injury.     

CD46 protects against chronic obstructive pulmonary disease

Background

Chronic obstructive pulmonary disease and emphysema develops in 15% of ex-smokers despite sustained quitting, while 10% are free of emphysema or severe lung obstruction. The cause of the incapacity of the immune system to clear the inflammation in the first group remains unclear.

Methods and Findings

We searched genes that were protecting ex-smokers without emphysema, using microarrays on portions of human lungs surgically removed; we found that loss of lung function in patients with chronic obstructive pulmonary disease and emphysema was associated with a lower expression of CD46 and verified this finding by qRT-PCR and flow cytometry. Also, there was a significant association among decreased CD46⁺ cells with decreased CD4⁺T cells, apoptosis mediator CD95 and increased CD8⁺T cells that were protecting patients without emphysema or severe chronic obstructive pulmonary disease. CD46 not only regulates the production of T regulatory cells, which suppresses CD8⁺T cell proliferation, but also the complement cascade by degradation of C3b. These results were replicated in the murine smoking model, which showed increased C5a (produced by C3b) that suppressed IL12 mediated bias to T helper 1 cells and elastin co-precipitation with C3b, suggesting that elastin could be presented as an antigen. Thus, using ELISA from elastin peptides, we verified that 43% of the patients with severe early onset of chronic obstructive pulmonary disease tested positive for IgG to elastin in their serum compared to healthy controls.

Conclusions

These data suggest that higher expression of CD46 in the lungs of ex-smoker protects them from emphysema and chronic obstructive pulmonary disease by clearing the inflammation impeding the proliferation of CD8⁺ T cells and necrosis, achieved by production of T regulatory cells and degradation of C3b; restraining the complement cascade favors apoptosis over necrosis, protecting them from autoimmunity and chronic inflammation.     

Caffeic acid protects rat heart mitochondria against isoproterenol-induced oxidative damage

Cardiac mitochondrial dysfunction plays an important role in the pathology of myocardial infarction. The protective effects of caffeic acid on mitochondrial dysfunction in isoproterenol-induced myocardial infarction were studied in Wistar rats. Rats were pretreated with caffeic acid (15 mg/kg) for 10 days. After the pretreatment period, isoproterenol (100 mg/kg) was subcutaneously injected to rats at an interval of 24 h for 2 days to induce myocardial infarction. Isoproterenol-induced rats showed considerable increased levels of serum troponins and heart mitochondrial lipid peroxidation products and considerable decreased glutathione peroxidase and reduced glutathione. Also, considerably decreased activities of isocitrate, succinate, malate, α-ketoglutarate, and NADH dehydrogenases and cytochrome-C-oxidase were observed in the mitochondria of myocardial-infarcted rats. The mitochondrial calcium, cholesterol, free fatty acids, and triglycerides were considerably increased and adenosine triphosphate and phospholipids were considerably decreased in isoproterenol-induced rats. Caffeic acid pretreatment showed considerable protective effects on all the biochemical parameters studied. Myocardial infarct size was much reduced in caffeic acid pretreated isoproterenol-induced rats. Transmission electron microscopic findings also confirmed the protective effects of caffeic acid. The possible mechanisms of caffeic acid on cardiac mitochondria protection might be due to decreasing free radicals, increasing multienzyme activities, reduced glutathione, and adenosine triphosphate levels and maintaining lipids and calcium. In vitro studies also confirmed the free-radical-scavenging activity of caffeic acid. Thus, caffeic acid protected rat’s heart mitochondria against isoproterenol-induced damage. This study may have a significant impact on myocardial-infarcted patients.     

Epidermal growth factor protects the heart against low-flow ischemia-induced injury

The role of ErbB4 and ErbB2 in the heart of adult mammals is well established. The heart also expresses ErbB1 (the epidermal growth factor (EGF) receptor), but this receptor has received less attention. We studied the effect of EGF on the response of isolated mouse heart to low-flow ischemia and reperfusion. Reducing perfusate flow to 10% for 30 min resulted in an increase in anaerobic metabolism and the leakage of lactate dehydrogenase during reperfusion. In addition, left ventricle +dP/dt and developed pressure were depressed (20–25%) during reperfusion. The addition of EGF 5 min before and throughout the ischemic period prevented the increase in anaerobic metabolism and the leakage of intracellular lactate dehydrogenase during reperfusion. EGF improved both +dP/dt and developed pressure during ischemia and prevented the decrease in dP/dt during reperfusion. To determine whether the effect of EGF on cell integrity depends on its effect on contractility, we studied nonbeating isolated myocytes. In these cells, anoxia and reoxygenation reduced cell viability by nearly 25%. EGF prevented such a decrease. Our results indicate that, like ErbB4 and ErbB2, ErbB1 also has an important role in the heart of adult animals.     

Bcl-xL gene transfer protects the heart against ischemia/reperfusion injury

Ischemia and reperfusion (I/R) injury causes the progression of cardiac dysfunction. The prevention of cardiomyocyte-loss due to I/R injury is important for the treatment of heart failure. Therefore, we employed antiapoptotic Bcl-xL protein to prevent I/R injury in the heart and evaluated the cardioprotective effect of Bcl-xL transduction by adenoviral vector (Adv) after I/R injury. Adv with Bcl-xL gene was injected in the rat heart 4 days prior to I/R. The prevention of cardiac performance-loss and the reduction of cardiac apoptosis, after 30min ischemia and 30min reperfusion of global I/R, were demonstrated in the heart with adenoviral Bcl-xL transduction. Also, significant reductions of the infarct size and serum creatine kinase levels were observed in the heart transduced with Bcl-xL gene compared with control after 30min ischemia and 24h reperfusion of the left anterior coronary artery. Thus, Bcl-xL may serve as a potential therapeutic tool for cardioprotection.     

Selenium protects the immature rat heart against ischemia/reperfusion injury

The aim of the study was to find out whether administration of selenium (Se) will protect the immature heart against ischemia/reperfusion. The control pregnant rats were fed laboratory diet (0.237 mg Se/kg diet); experimental rats received 2 ppm Na₂SeO₃ in the drinking water from the first day of pregnancy until day 10 post partum. The concentration of Se in the serum and heart tissue was determined by activation analysis, the serum concentration of NO by chemiluminescence, cardiac concentration of lipofuscin-like pigment by fluorescence analysis. The 10 day-old hearts were perfused (Langendorff); recovery of developed force (DF) was measured after 40 min of global ischemia. In acute experiments, 10 day-old hearts were perfused with selenium (75 nmol/l) before or after global ischemia. Sensitivity to isoproterenol (ISO, pD₅₀) was assessed as a response of DF to increasing cumulative dose. Se supplementation elevated serum concentration of Se by 16%. Se increased ischemic tolerance (recovery of DF, 32.28 ± 2.37 vs. 41.82 ± 2.91%, P < 0.05). Similar results were obtained after acute administration of Se during post-ischemic reperfusion (32.28 ± 2.37 vs. 49.73 ± 4.40%, P < 0.01). The pre-ischemic treatment, however, attenuated the recovery (23.08 ± 3.04 vs. 32.28 ± 2.37%, P < 0.05). Moreover, Se supplementation increased the sensitivity to the inotropic effect of ISO, decreased cardiac concentration of lipofuscin-like pigment and serum concentration of NO. Our results suggest that Se protects the immature heart against ischemia/reperfusion injury. It seems therefore, that ROS may affect the function of the neonatal heart, similarly as in adults.     

Wild-type presenilin 1 protects against Alzheimer disease mutation-induced amyloid pathology

Mutations in presenilin 1 (PS1) lead to dominant inheritance of early onset familial Alzheimer disease (FAD). These mutations are known to alter the gamma-secretase cleavage of the amyloid precursor protein, resulting in increased ratio of Abeta42/Abeta40 and accelerated amyloid plaque pathology in transgenic mouse models. To investigate the factors that drive the Abeta42/Abeta40 ratio and amyloid pathogenesis and to investigate the possible interactions between wild-type and FAD mutant PS1, which are co-expressed in transgenic animals, we expressed the PS1 M146V knock-in allele either on wild-type PS1 (PS1M146V/+) or PS1 null (PS1M146V/-) background and crossed these alleles with the Tg2576 APP transgenic mice. Introduction of the PS1 M146V mutation on Tg2576 background resulted in earlier onset of plaque pathology. Surprisingly, removing the wild-type PS1 in the presence of the PS1 M146V mutation (PS1M146V/-) greatly exacerbated the amyloid burden; and this was attributed to a reduction of gamma-secretase activity rather than an increase in Abeta42. Our findings establish a protective role of the wild-type PS1 against the FAD mutation-induced amyloid pathology through a partial loss-of-function mechanism.     

Exercise protects the heart against myocardial infarction through upregulation of miR-1192

Myocardial infarction (MI) is known as a serious global problem, which has a high mortality rate and cause severe heart damage. Mounting evidence has suggested that exercise provides direct endogenous cardiac protection against various cardiovascular diseases including MI. However, the underlying mechanism of exercise’s cardioprotective effect against MI has not been fully understood. Here, we found that a 4-wk swim training exerted protective effects against MI in C57 mice, as evidenced by increased cardiac function and decreased cardiac apoptosis. A plasma miRNA profiling assay was then performed, and 10 differentially expressed miRNAs were detected. Among them, miR-1192 was increased after exercise, and it exerted significant protective effect against hypoxia in cultured neonatal cardiomyocytes. In addition, intramyocardially injection of agomiR-1192 exerted similar cardioprotective effect as exercise, and inhibition of miR-1192 using antgomiR-1192 abolished the cardioprotective effect of exercise in MI mice, suggesting that exercise exerted cardioprotection against MI through upregulation of miR-1192. Furthermore, we found that miR-1192 exerted cardioprotective effect via targeting caspase 3 in cardiomyocytes. These findings suggested that exercise protects the heart against MI through upregulation of miR-1192, and miR-1192 is a novel exerkine in exercise-induced cardioprotection against MI.     

Nuclear ATR lysine-tyrosylation protects against heart failure by activating DNA damage response

1. Download : Download high-res image (155KB)
2. Download : Download full-size image

     

Enhancing AMPK activation during ischemia protects the diabetic heart against reperfusion injury

AMPK activation during ischemia helps the myocardium to cope with the deficit of energy production. As AMPK activity is considered to be impaired in diabetes, we hypothesized that enhancing AMPK activation during ischemia above physiological levels would protect the ischemic diabetic heart through AMPK activation and subsequent inhibition of mitochondrial permeability transition pore (mPTP) opening. Isolated perfused hearts from normoglycemic Wistar or diabetic Goto-Kakizaki (GK) rats (n ≥ 6/group) were subjected to 35 min of ischemia in the presence of 10, 20, and 40 μM of A-769662, a known activator of AMPK, followed by 120 min of reperfusion with normal buffer. Myocardial infarction and AMPK phosphorylation were assessed. The effect of A-769662 on mPTP opening in adult cardiomyocytes isolated from both strains was also determined. A-769662 at 20 μM reduced infarct size in both Wistar (30.5 ± 2.7 vs. 51.8 ± 3.9% vehicle; P < 0.001) and GK hearts (22.7 ± 3.0 vs. 48.5 ± 4.7% vehicle; P < 0.001). This protection was accompanied by a significant increase in AMPK and GSK-3β phosphorylation. In addition, A-769662 significantly inhibited mPTP opening in both Wistar and GK cardiomyocytes subjected to oxidative stress. We demonstrate that AMPK activation during ischemia via A-769662 reduces myocardial infarct size in both the nondiabetic and diabetic rat heart. Furthermore, this cardioprotective effect appears to be mediated through inhibition of mPTP opening. Our findings suggest that improving AMPK activation during ischemia can be another mechanism for protecting the ischemic heart.     

Pretreatment with methylprednisolone protects the isolated rat heart against ischaemic and oxidative damage

In this study, we investigated whether a relationship exists between the levels of urate in vivo and lipid peroxidation during exercise. Seven healthy male subjects performed exhaustive cycling exercise under the following three conditions. The levels of urate, thiobarbituric acid reactive substances (TBARS) and allantoin in plasma and urine were examined before exercise and during a 3 h recovery period. (1) Benzbromarone administration experiment: benzbromarone (an uricosuric agent) was administered orally the day before exercise. (2) IMP administration experiment: inosine 5′-monophosphate disodium salt (a precursor of urate) was administered orally the day before exercise. (3) Control experiment: no test substance was administered. The main results obtained were as follows. Plasma urate levels and total peroxyl radical-trapping antioxidant parameter (TRAP) for deproteinized plasma in the resting period significantly decreased depending on the treatment, in the order of IMP > control > benzbromarone. A significant positive correlation was evident between plasma urate levels and TRAP values for deproteinized plasma. The increase in plasma levels of allantoin was observed only in the case of IMP treatment. A significant negative correlation between plasma levels of urate in the resting period and the amounts of urinary TBARS excreted during the recovery period was recognized. These results suggest that the urate level in vivo before exercise is a factor influencing lipid peroxidation during exhaustive exercise. Furthermore, these findings support the view that urate may serve as an important free-radical scavenger in vivo.     

Ghrelin protects heart against ERS-induced injury and apoptosis by activating AMP-activated protein kinase

Ghrelin, the endogenous ligand of growth hormone secretagogue receptor (GHS-R), is a cardioprotective peptide. In our previous work, we have revealed that ghrelin could protect heart against ischemia/reperfusion (I/R) injury by inhibiting endoplasmic reticulum stress (ERS), which contributes to many heart diseases. In current study, using both in vivo and in vitro models, we investigated how ghrelin inhibits myocardial ERS. In the in vivo rat heart injury model induced by isoproterenol (ISO), we found that exogenous ghrelin could alleviate heart dysfunction, reduce myocardial injury and apoptosis and inhibit the excessive myocardial ERS induced by ISO. More importantly, the activation of AMP-activated protein kinase (AMPK) was observed. To explore the role of AMPK activation in ERS inhibition by ghrelin, we set up two in vitro ERS models by exposing cultured rat cardiomyocytes to tunicamycin(Tm) or dithiothreitol (DTT). In both models, compared with Tm or DTT treatment alone, pre-incubation cardiomyocytes with ghrelin significantly activated AMPK, reversed the upregulation of the ERS markers, C/EBP-homologous protein (CHOP) and cleaved caspase-12, and reduced apoptosis of cardiomyocytes. Further, we found that the ERS inhibitory and anti-apoptotic actions induced by ghrelin were blocked by an AMPK inhibitor. To investigate how ghrelin activates AMPK, selective antagonist of GHS-R1a and inhibitor of Ca²⁺/Calmodulin-dependent protein kinase kinase (CaMKK) were added, respectively, before ghrelin pre-incubation, and we found that AMPK activation was prevented and the ERS inhibitory and anti-apoptotic actions of ghrelin were blocked. In conclusion, ghrelin could protect heart against ERS-induced injury and apoptosis, at least partially through a GHS-R1a/CaMKK/AMPK pathway.     

Voluntary exercise protects against acute doxorubicin cardiotoxicity in the isolated perfused rat heart

The clinical use of doxorubicin (DOX) is limited by a dose-dependent cardiotoxicity. The purpose of this study was to determine whether voluntary exercise training would confer protection against DOX cardiotoxicity in the isolated perfused rat heart. Female Sprague-Dawley rats were randomly assigned to standard holding cages or cages with running wheels for 8 wk. Twenty-four hours after the sedentary (SED) or voluntary exercise (VEX) running period, rats were anesthetized with pentobarbital sodium, and hearts were isolated and perfused with oxygenated Krebs-Henseleit (KH) buffer at a constant flow of 15 ml/min. After a 20-min stabilization period, hearts were paced at 300 beats per minute and perfused with KH buffer containing 10 microM DOX for 60 min. A set of control hearts from SED and VEX rats were perfused under identical conditions without DOX for the same period. DOX perfusion led to significant decreases in left ventricular developed pressure, +dP/dt, and -dP/dt, and significant increases in LV lipid peroxidation in sedentary rats compared with non-DOX controls (P < 0.05). Prior voluntary exercise training attenuated these DOX-induced effects and was associated with a significant increase (78%, P < 0.05) in heat shock protein (HSP72), but not mitochondrial isoform of SOD (MnSOD) or CuZnSOD protein expression in the hearts of wheel-run animals. These data indicate that chronic physical activity may provide resistance against the cardiac dysfunction and oxidative damage associated with DOX exposure and provide novel evidence of HSP72 induction in the heart after voluntary exercise.     

Pretreatment with methylprednisolone protects the isolated rat heart against ischaemic and oxidative damage

Methylprednisolone (MP), a synthetic glucocorticoid, is widely used clinically and experimentally as acute antiinflammatory treatment. The molecular actions of MP indicate that pretreatment with this drug may be cardioprotective. We investigated if giving rats MP prior to excising their hearts for Langendorff-perfusion protected cardiac function against oxidative stress, and if this was mediated by increasing antioxidant defence or influencing myocardial nitric oxide synthase (NOS). Rats (n=6-11 in each group) were injected with MP (40 mg/kg i.m.) or vehicle 24 and 12 h before Langendorff-perfusion with 30 min global ischaemia and 60 min reperfusion, or 10 min perfusion with 180 μmol/L hydrogen peroxide. Other hearts were exposed to 30 min global ischaemia 5 days after MP-injection. Additional hearts were sampled before, during, and after ischaemia for analyzing tissue activity of antioxidant enzymes. Tissue endothelial and inducible NOS (eNOS and iNOS) were investigated by immunoblotting and semiquantitative RT-PCR in a time-course after MP injection. Pretreatment with MP improved left ventricular function and increased coronary flow during postischaemic reperfusion, and this effect was sustained 5 days afterwards. When exposing hearts to hydrogen peroxide, MP improved coronary flow. Catalase, glutathione peroxidase, and oxidized glutathione were increased during reperfusion of MP-treated hearts compared to vehicle only. MP did not influence eNOS at protein or mRNA level. iNOS could not be detected by immunoblotting, indicating low cardiac enzyme content. Its mRNA initially increased the first hour after injection, thereafter decreased. In conclusions, pretreating rats with MP protects the heart against ischaemia-reperfusion dysfunction. This effect could be due to increase of tissue antioxidant activity during reperfusion. MP did not influence cardiac eNOS. mRNA for iNOS was influenced by MP, but the corresponding protein could not be detected.