Expression Of Sarcoplasmic Reticulum Ca2 Research Articles

Estrogen inhibits endoplasmic reticulum stress and ameliorates myocardial ischemia/reperfusion injury in rats by upregulating SERCA2a

BackgroundThe incidence of coronary heart disease (CHD) in premenopausal women is significantly lower than that of men of the same age, suggesting protective roles of estrogen for the cardiovascular system against CHD. This study aimed to confirm the protective effect of estrogen on myocardium during myocardial ischemia/reperfusion (MI/R) injury and explore the underlying mechanisms.MethodsNeonatal rat cardiomyocytes and Sprague–Dawley rats were used in this study. Different groups were treated by bilateral ovariectomy, 17β-estradiol (E2), adenoviral infection, or siRNA transfection. The expression of sarcoplasmic reticulum Ca2+ ATPase pump (SERCA2a) and endoplasmic reticulum (ER) stress-related proteins were measured in each group to examine the effect of different E2 levels and determine the relationship between SERCA2a and ER stress. The cell apoptosis, myocardial infarction size, levels of apoptosis and serum cardiac troponin I, ejection fraction, calcium transient, and morphology changes of the myocardium and ER were examined to verify the effects of E2 on the myocardium.ResultsBilateral ovariectomy resulted in reduced SERCA2a levels and more severe MI/R injury. E2 treatment increased SERCA2a expression. Both E2 treatment and exogenous SERCA2a overexpression decreased levels of ER stress-related proteins and alleviated myocardial damage. In contrast, SERCA2a knockdown exacerbated ER stress and myocardial damage. Addition of E2 after SERCA2a knockdown did not effectively inhibit ER stress or reduce myocardial injury.ConclusionsOur data demonstrate that estrogen inhibits ER stress and attenuates MI/R injury by upregulating SERCA2a. These results provide a new potential target for therapeutic intervention and drug discovery in CHD.BAgGUtqRXuc394yJDQeuxtVideo

Gallic acid protects against isoproterenol-induced cardiotoxicity in rats.

Gallic acid (GA) is a polyphenolic agent with interesting pharmacological impacts on the cardiovascular system. The present study purposed to study the protective effects of GA at 25 and 50mg/kg against isoproterenol (ISO)-induced cardiac damage in ischemia/reperfusion (I/R) in rats. Male Wistar rats were randomly assigned into six groups: Control, Control treated with GA at 25mg/kg (GA25), Control treated with GA at 50mg/kg (GA50), Hypertrophic rats induced by ISO (ISO), Hypertrophic rats treated with GA at 25mg/kg (ISO+GA25), and Hypertrophic rats treated with GA at 50mg/kg (ISO+GA50). Heart isolation was performed to induce a cardiac I/R injury model. Cardiac hemodynamic parameters were recorded. Serum Lactate Dehydrogenase (LDH) and Creatine Kinase-MB (CK-MB) and cardiac Superoxide dismutases (SOD) levels were evaluated. The gene expression of Sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) was assessed. We found that GA at 50mg/kg was significantly increased cardiac function at post I/R period in ISO-induced hypertrophic hearts. Moreover, it suppressed cardiac hypertrophy, the serum LDH and CK-MB levels in ISO injected rats. Administration of GA at 50mg/kg was significantly increased SOD level and SERCA2a gene expression in the hypertrophic hearts. GA at 50mg/kg could improve cardiac performance possibly by increasing antioxidant defense enzymes, reducing cell damage, and enhancing SERCA2a gene expression in hypertrophic heart induced by ISO in I/R injury conditions.

Maximal strength training-induced increase in efferent neural drive is not reflected in relative protein expression of SERCA

IntroductionMaximal strength training (MST), performed with heavy loads (~ 90% of one repetition maximum; 1RM) and few repetitions, yields large improvements in efferent neural drive, skeletal muscle force production, and skeletal muscle efficiency. However, it is elusive whether neural adaptations following such high intensity strength training may be accompanied by alterations in energy-demanding muscular factors.MethodsSixteen healthy young males (24 ± 4 years) were randomized to MST 3 times per week for 8 weeks (n = 8), or a control group (CG; n = 8). Measurements included 1RM and rate of force development (RFD), and evoked potentials recordings (V-wave and H-reflex normalized to M-wave (M) in the soleus muscle) applied to assess efferent neural drive to maximally contracting skeletal muscle. Biopsies were obtained from vastus lateralis and analyzed by western blots and real-time PCR to investigate the relative protein expression and mRNA expression of Sarcoplasmic Reticulum Ca2+ ATPase (SERCA) 1 and SERCA2.ResultsSignificant improvements in 1RM (17 ± 9%; p < 0.001) and early (0–100 ms), late (0–200 ms) and maximal RFD (31–53%; p < 0.01) were observed after MST, accompanied by increased maximal Vmax/Msup-ratio (9 ± 14%; p = 0.046), with no change in H-reflex to M-wave ratio. No changes were observed in the CG. No pre- to post-training differences were found in mRNA or protein expressions of SERCA1 and SERCA2 in either group.ConclusionMST increased efferent neural drive to maximally contracting skeletal muscle, causing improved force production. No change was observed in SERCA expression, indicating that responses to high intensity strength training may predominantly be governed by neural adaptations.

In vivo effects of nitrosyl hydrogen on cardiac function and sarcoplasmic reticulum calcium pump (SERCA2a) in rats with heart failure after myocardial infarction.

Abnormal Ca2+ circulation in cardiomyocytes is an important cause of decreased myocardial contractility in failing hearts. Nitroxyl hydrogen (HNO) can oxidize Ca2+ cycle-related proteins, alter their biological functions, promote Ca2+ recovery as well as release, and enhance myocardial contractility. In this study, we aim to observe the effect of nitrosyl hydrogen (HNO) on the cardiac function of rats with heart failure and elucidate the underlying mechanism. Twenty six male Wistar rats were randomly divided into heart failure group (HF group), Angeli's salt treatment group (HF + AS group) and sham operation group (Sham group). The HF + AS group rats were treated with HNO donor Angeli's salt by intraperitoneal injection of 1 mg/kg/d, and the rats in the HF group and the Sham group were given the same amount of normal saline. Cardiac function was measured by echocardiography before and after treatment. NT-proBNP was measured by enzyme immunoassay (ELISA) kit after treatment. Western blot were used to measure the expression of sarcoplasmic reticulum Ca2+-ATPase (SERCA) in protein levels in rats. The activities of SERCA2a were detected by the biochemical kit finally. We found that Nitrosyl hydrogen could significantly increase LVEF, +dp/dt, -dp/dt (P<0.05), significantly decrease NT-ProBNP and LVEDP (P<0.01), and significantly enhance the activities of SERCA2a protein (P<0.05). These findings suggest that Nitrosyl hydrogen could improve the cardiac function possibly by increasing protein activities of SERCA2a in rats.

Neural And Muscular Responses To Maximal Strength Training

PURPOSE: Maximal strength training (MST), performed with heavy loads (~90% of one repetition maximum; 1RM) and few repetitions (4-5), yields large improvements in efferent neural drive and skeletal muscle force generating capacity. However, it is elusive how MST-induced neural adaptations may translate to muscular factors regulating excitation-contraction coupling. MEHODS: Sixteen healthy young males (24±4 years) were randomized to MST 3 times per week for 8 weeks (n=8), or a control group (CG; n=8). Measurements were taken of 1RM and rate of force development (RFD), and evoked potentials recordings (V-wave and H-reflex normalized to M-wave (M) in musculus soleus) applied to assess efferent neural drive to maximally contracting skeletal muscle. Biopsies were obtained from m. vastus lateralis and analyzed by western blot and mRNA isolation to investigate the protein expression of Sarcoplasmic Reticulum Ca2+ ATPase (SERCA) and mRNA expression of SERCA1 and SERCA2, myostatin, MuRF1 and Ryanodine receptor (RyR1). RESULTS: 1RM (17±9%; p<0.05) and early (0-100ms), late (100-200ms) and maximal RFD increased (31-53%; p<0.01) in the MST group, accompanied by increased maximal V-M wave ratio (9±14%; p<0.05), with no change in H-reflex to M-wave ratio. No changes were observed in the CG. No pre- to post-training differences were found in mRNA or protein expressions in either group (p>0.05). CONCLUSION: MST increased efferent neural drive to maximally contracting skeletal muscle, and resulted in improved force generating capacity. The neural adaptations were not reflected in key muscular factors involved in excitation-contraction coupling, indicating that responses to high intensity strength training may predominantly be governed by neural adaptations.

Differential isoform expression of SERCA and myosin heavy chain in hypopharyngeal muscles.

SUMMARYComposition of slow, fast and hybrid fibres of pharyngeal muscles, associated with pharyngeal movements and regulation, has been rarely studied. The present study aimed to identify expression of sarcoplasmic reticulum Ca2+ ATPase (SERCA) and myosin heavy chain (MHC) and hybrid isoforms in different pharyngeal muscles of young and aged rats as well as humans. Isoform expression profiles of SERCA, MHC and hybrid isoforms among six components of pharyngeal muscle were immunohistochemically evaluated in rat and human. The result showed that pharyngeal muscles predominantly expressed fast fibres (SERCA1 and MHCII), whereas expression of slow fibres (SERCA2 and MHCI) was low, but different depending on muscle components. Inner layer of pharyngeal muscles expressed more SERCA2 and hybrid fibres than the outer layer. Pharyngeal muscles in aged rats showed increased hybrid fibers and SERCA2. Human thyropharyngeus also showed a higher portion of fast fibres compared to cricopharyngeus. Thus, in contrast to abundance of fast fibres, slow and hybrid fibres are differentially expressed depending on muscle components and layers as well as aging. These results lead to further understanding of coordinated regulation for speech and swallowing. The unique data presented in this study on SERCA isoform expressions in both rats and human suggest an ability to handle calcium changes according functional demands.

Asprosin attenuates insulin signaling pathway through PKCδ-activated ER stress and inflammation in skeletal muscle.

It has been reported that asprosin is a novel adipokine which is augmented in mice and humans with type 2 diabetes (T2DM). Asprosin stimulates hepatic gluconeogenesis under fasting conditions. However, the roles of asprosin in inflammation, endoplasmic reticulum (ER) stress, and insulin resistance in skeletal muscle has not been studied. In the currents study, elevated levels of asprosin expression were observed in adipocytes under hyperlipidemic conditions. Treatment of C2C12 myocytes with asprosin-induced ER stress markers (phosphorylated inositol-requiring enzyme 1 and eukaryotic initiation factor 2, and CHOP expression) as well as inflammation markers (interleukin-6 expression, phosphorylated IκB, and nuclear translocated nuclear factor-κβ). Finally, asprosin treatment promoted exacerbation of insulin sensitivity as determined by levels of insulin receptor substrate 1 and Akt phosphorylation as well as glucose uptake. Moreover, treatment of asprosin augmented protein kinase C-δ (PKCδ) phosphorylation and nuclear translocation, but suppressed messenger RNA expression of sarcoplasmic reticulum Ca2+ ATPase 2b in both C2C12 myocytes and in mouse soleus skeletal muscle. These asprosin-induced effects were markedly decreased in small interfering (si) RNA-mediated PKCδ-knockdown in C2C12 myocytes. These results suggest that asprosin results in impairment of insulin sensitivity in skeletal muscle through PKCδ-associated ER stress/inflammation pathways and may be a valuable strategy for management of insulin resistance and T2DM.

The Formin, DIAPH1, is a Key Modulator of Myocardial Ischemia/Reperfusion Injury

The biochemical, ionic, and signaling changes that occur within cardiomyocytes subjected to ischemia are exacerbated by reperfusion; however, the precise mechanisms mediating myocardial ischemia/reperfusion (I/R) injury have not been fully elucidated. The receptor for advanced glycation end-products (RAGE) regulates the cellular response to cardiac tissue damage in I/R, an effect potentially mediated by the binding of the RAGE cytoplasmic domain to the diaphanous-related formin, DIAPH1. The aim of this study was to investigate the role of DIAPH1 in the physiological response to experimental myocardial I/R in mice. After subjecting wild-type mice to experimental I/R, myocardial DIAPH1 expression was increased, an effect that was echoed following hypoxia/reoxygenation (H/R) in H9C2 and AC16 cells. Further, compared to wild-type mice, genetic deletion of Diaph1 reduced infarct size and improved contractile function after I/R. Silencing Diaph1 in H9C2 cells subjected to H/R downregulated actin polymerization and serum response factor-regulated gene expression. Importantly, these changes led to increased expression of sarcoplasmic reticulum Ca2+ ATPase and reduced expression of the sodium calcium exchanger. This work demonstrates that DIAPH1 is required for the myocardial response to I/R, and that targeting DIAPH1 may represent an adjunctive approach for myocardial salvage after acute infarction.

Transgenic overexpression of GTP cyclohydrolase 1 in cardiomyocytes ameliorates post-infarction cardiac remodeling

GTP cyclohydrolase 1 (GCH1) and its product tetrahydrobiopterin play crucial roles in cardiovascular health and disease, yet the exact regulation and role of GCH1 in adverse cardiac remodeling after myocardial infarction are still enigmatic. Here we report that cardiac GCH1 is degraded in remodeled hearts after myocardial infarction, concomitant with increases in the thickness of interventricular septum, interstitial fibrosis, and phosphorylated p38 mitogen-activated protein kinase and decreases in left ventricular anterior wall thickness, cardiac contractility, tetrahydrobiopterin, the dimers of nitric oxide synthase, sarcoplasmic reticulum Ca2+ release, and the expression of sarcoplasmic reticulum Ca2+ handling proteins. Intriguingly, transgenic overexpression of GCH1 in cardiomyocytes reduces the thickness of interventricular septum and interstitial fibrosis and increases anterior wall thickness and cardiac contractility after infarction. Moreover, we show that GCH1 overexpression decreases phosphorylated p38 mitogen-activated protein kinase and elevates tetrahydrobiopterin levels, the dimerization and phosphorylation of neuronal nitric oxide synthase, sarcoplasmic reticulum Ca2+ release, and sarcoplasmic reticulum Ca2+ handling proteins in post-infarction remodeled hearts. Our results indicate that the pivotal role of GCH1 overexpression in post-infarction cardiac remodeling is attributable to preservation of neuronal nitric oxide synthase and sarcoplasmic reticulum Ca2+ handling proteins, and identify a new therapeutic target for cardiac remodeling after infarction.

Exendin-4 inhibits structural remodeling and improves Ca2+ homeostasis in rats with heart failure via the GLP-1 receptor through the eNOS/cGMP/PKG pathway

The glucagon-like peptide-1 receptor (GLP-1R) agonist exendin-4 is a long-acting analog of GLP-1, which stimulates insulin secretion and is clinically used in the treatment of type 2 diabetes. Previous studies have demonstrated that GLP-1 agonists and analogs serve as cardioprotective factors in various conditions. Disturbances in calcium cycling are characteristic of heart failure (HF); therefore, the aim of this study was to investigate the effect of exendin-4 (a GLP-1 mimetic) on the regulation of calcium handling and to identify the underlying mechanisms in an HF rat model after myocardial infarction (MI). Rats underwent surgical ligation of the left anterior descending coronary artery or sham surgery prior to infusion with vehicle, exendin-4, or exendin-4 and exendin9-39 for 4 weeks. Exendin-4 treatment decreased MI size, suppressed chamber dilation, myocyte hypertrophy, and fibrosis and improved in vivo heart function in the rats subjected to MI. Exendin-4 resulted in an increase in circulating GLP-1 and GLP-1R in ventricular tissues. Additionally, exendin-4 activated the eNOS/cGMP/PKG signaling pathway and inhibited the Ca2+/calmodulin-dependent kinase II (CaMKII) pathways. Myocytes isolated from exendin-4-treated hearts displayed higher Ca2+ transients, higher sarcoplasmic reticulum Ca2+ content, and higher l-type Ca2+ current densities than MI hearts. Exendin-4 treatment restored the protein expression of sarcoplasmic reticulum Ca2+ uptake ATPase (SERCA2a), phosphorylated phospholamban (PLB) and Cav1.2 and decreased the levels of phosphorylated ryanodine receptor (RyR). Moreover, the favorable effects of exendin-4 were significantly inhibited by exendin9-39 (a GLP-1 receptor antagonist). Exendin-4 treatment of an HF rat model after MI inhibited cardiac and cardiomyocytes progressive remodeling. In addition, Ca2+ handling and its molecular modulation were also improved by exendin-4 treatment. The beneficial effects of exendin-4 on cardiac remodeling may be mediated through activation of the eNOS/cGMP/PKG pathway.

Gene therapy in heart failure: the unexpected results from the CUPID 2 trial

Heart failure is still a major cause of morbidity and mortality in Europe and North America. In the last three decades, gene therapy emerged as a target in the molecular mechanisms implicated in heart failure encouraging preclinical gene therapy studies in small and large animal models. Prior studies documented a decreased expression of sarcoplasmic reticulum Ca2+-ATPase protein (SERCA2a), a major cardiac calcium cycling protein, in heart failure. These results paved the way to preliminary studies based on gene transfer strategies of SERCA2a. The encouraging results in terms of safety and surrogate clinical endpoints led to a large randomized clinical study, the CUPID trial, including patients with heart failure and reduced ejection fraction. This phase IIb trial enrolling 250 patients randomized to intracoronary delivery of adeno-associated virus 1 (AAV1)/SERCA2a or placebo concluded with a non-superiority of the studied therapy, thus not confirming the results of the previous experiences with the same approach. In the present manuscript, we provide an overview of the scientific experiences that preceded the design of this major trial, then critically revising its structure and results. Finally we tried to understand the reasons of this unexpected failure and which are the future perspectives of gene therapy for heart failure.

Vasostatin-1 Stops Structural Remodeling and Improves Calcium Handling via the eNOS-NO-PKG Pathway in Rat Hearts Subjected to Chronic β-Adrenergic Receptor Activation

Chronically elevated catecholamine levels activate cardiac β-adrenergic receptors, which play a vital role in the pathogenesis of heart failure. Evidence suggests that vasostatin-1 (VS-1) exerts anti-adrenergic effects on isolated and perfused hearts in vitro. Whether VS-1 ameliorates hypertrophy/remodeling by inducing the chronic activation of β-adrenergic receptors is unknown. The present study aims to test the efficacy of using VS-1 to treat the advanced hypertrophy/remodeling that result from chronic β-adrenergic receptor activation and to determine the cellular and molecular mechanisms that underlie this response. Rats were subjected to infusion with either isoprenaline (ISO, 5mg/kg/d), ISO plus VS-1 (30mg/kg/d) or placebo for 2weeks. VS-1 suppressed chamber dilation, myocyte hypertrophy and fibrosis and improved in vivo heart function in the rats subjected to ISO infusion. VS-1 increased phosphorylated nitric oxide synthase levels and induced the activation of protein kinase G. VS-1 also deactivated multiple hypertrophy signaling pathways that were triggered by the chronic activation of β-adrenergic receptors, such as the phosphoinositide-3 kinase (PI3K)/Akt and Ca2+/calmodulin-dependent kinase (CaMK-II) pathways. Myocytes isolated from ISO+VS-1 hearts displayed higher Ca2+ transients, shorter Ca2+ decays, higher sarcoplasmic reticulum Ca2+ levels and higher L-type Ca2+ current densities than the ISO rat hearts. VS-1 treatment restored the protein expression of sarcoplasmic reticulum Ca2+ uptake ATPase, phospholamban and Cav1.2, indicating improved calcium handling. Chronic VS-1 treatment inhibited the progression of hypertrophy, fibrosis, and chamber remodeling, and improved cardiac function in a rat model of ISO infusion. In addition, Ca2+ handling and its molecular modulation were also improved by VS-1. The beneficial effects of VS-1 on cardiac remodeling may be mediated by the enhanced activation of the eNOS-cGMP-PKG pathway.

Shen-Fu Injection (参附注射液) alleviates post-resuscitation myocardial dysfunction by up-regulating expression of sarcoplasmic reticulum Ca2+-ATPase

To compare the effect of Shen-Fu Injection (SFI) and epinephrine on the expression of sarcoplasmic reticulum Ca(2+) ATPase 2a (SERCA2a) in a pig model with post-resuscitation myocardial dysfunction. Ventricular fibrillation (VF) was electrically induced in Wu-zhi-shan miniature pigs. After 8 min of untreated VF and 2 min of cardiopulmonary resuscitation (CPR), all animals were randomly administered a bolus injection of saline placebo (SA group, n=10), SFI (0.8 mg/kg, SFI group, n=10) or epinephrine (20 μg/kg, EPI group, n=10). After 4 min of CPR, a 100-J shock was delivered. If the defibrillation attempt failed to attain restoration of spontaneous circulation (ROSC), manual chest compressions were rapidly resumed for a further 2 min followed by a second defibrillation attempt. Hemodynamic variables were recorded, and plasma concentrations of catecholamines were measured. Adenylate cyclase (AC), cyclic adenosine monophosphate (cAMP) and the expressions of β1-adrenoceptor (AR) and SERCA 2a were determined. Cardiac output, left ventricular dp/dtmax and negative dp/dtmax were significantly higher in the SFI group than in the SA and EPI groups at 4 and 6 h after ROSC. The expression of β1-AR and SERCA2a at 24 h after ROSC were significantly higher in the SFI group than in the SA and EPI groups (P<0.05 or P<0.01). The administration of epinephrine during CPR decreased the expression of SERCA2a and aggravated postresuscitation myocardial function (P<0.01). SFI attenuated post-resuscitation myocardial dysfunction, and the mechanism might be related to the up-regulation of SERCA2a expression.

Intermittent cardiac overload results in adaptive hypertrophy and provides protection against left ventricular acute pressure overload insult.

The present study aimed to test whether a chronic intermittent workload could induce an adaptive cardiac phenotype Chronic intermittent workload induced features of adaptive hypertrophy This was paralleled by protection against acute pressure overload insult The heart may adapt favourably to balanced demands, regardless of the nature of the stimuli. The present study aimed to test whether submitting the healthy heart to intermittent and tolerable amounts of workload, independently of its nature, could result in an adaptive cardiac phenotype. Male Wistar rats were subjected to treadmill running (Ex) (n = 20), intermittent cardiac overload with dobutamine (ITO) (2 mg kg(-1) , s.c.; n = 20) or placebo administration (Cont) (n = 20) for 5 days week(-1) for 8 weeks. Animals were then killed for histological and biochemical analysis or subjected to left ventricular haemodynamic evaluation under baseline conditions, in response to isovolumetric contractions and to sustained LV acute pressure overload (35% increase in peak systolic pressure maintained for 2 h). Baseline cardiac function was enhanced only in Ex, whereas the response to isovolumetric heartbeats was improved in both ITO and Ex. By contrast to the Cont group, in which rats developed diastolic dysfunction with sustained acute pressure overload, ITO and Ex showed increased tolerance to this stress test. Both ITO and Ex developed cardiomyocyte hypertrophy without fibrosis, no overexpression of osteopontin-1 or β-myosin heavy chain, and increased expression of sarcoplasmic reticulum Ca(2+) protein. Regarding hypertrophic pathways, ITO and Ex showed activation of the protein kinase B/mammalian target of rapamycin pathway but not calcineurin. Mitochondrial complex IV and V activities were also increased in ITO and Ex. Chronic submission to controlled intermittent cardiac overload, independently of its nature, results in an adaptive cardiac phenotype. Features of the cardiac overload, such as the duration and magnitude of the stimuli, may play a role in the development of an adaptive or maladaptive phenotype.

Effect of oral administration of DHA on the remodeling of calcium handling proteins in congestive heart failure in male rats

Calcium (Ca2+) cycling is essential in cardiac contractile function, and remodeling of calcium handling is thought to be one of the factors contributing to the dysfunction in heart failure (HF). This study investigated the effect of DHA administration on remodeling of Ca2+handling proteins in congestive heart failure (CHF). Male rats were divided into 4 groups (n=8 for each group): C (normal, administered saline); D (normal, administered DHA‐ethyl ester (DHA‐Et)); H (CHF, administered saline); HD (CHF, administered DHA‐Et). CHF was produced by injection of monocrotaline (MCT) into 5‐week‐old rats. Starting from 3 days before MCT injection, saline or DHA‐Et wasorally administered(1g/kg body weight) daily. The right ventriclewas removed on day 24, and quantitative PCR was used to measure the expression of transcripts.The expression of sarcoplasmic reticulum Ca2+‐ATPase (SERCA) 2a, which is responsible to transport back Ca2+ to sarcoplasmic reticulum (SR), was significantly decreased in group H compared to groups C, D and HD (P 蠄 0.05). The expression of ryanodine receptor (RYR) 3, which releases Ca2+ from SR, in group H was significantly increased compared to groups C and D (P 蠄 0.05), and also tended to be increased in group H compared to group HD (P=0.06).The expression of Cav1.2, a component of L‐type Ca2+ channel,was significantly decreased in group H compared to groups C and D (P 蠄 0.05). Remodeling of intracellular Ca2+ handling may be caused by these changes in gene expression in CHF. Administration of DHA‐Et may improve abnormal Ca2+ handling through recovery of the expression of these proteins.

Running performance at high running velocities is impaired but V'O(₂max) and peripheral endothelial function are preserved in IL-6⁻/⁻ mice.

It has been reported that IL-6 knockout mice (IL-6−/−) possess lower endurance capacity than wild type mice (WT), however the underlying mechanism is poorly understood. The aim of the present work was to examine whether reduced endurance running capacity in IL-6−/− mice is linked to impaired maximal oxygen uptake (V′O2max), decreased glucose tolerance, endothelial dysfunction or other mechanisms. Maximal running velocity during incremental running to exhaustion was significantly lower in IL-6−/− mice than in WT mice (13.00±0.97 m.min−1 vs. 16.89±1.15 m.min−1, P<0.02, respectively). Moreover, the time to exhaustion during running at 12 m.min−1 in IL-6−/− mice was significantly shorter (P<0.05) than in WT mice. V′O2max in IL-6−/− (n = 20) amounting to 108.3±2.8 ml.kg−1.min−1 was similar as in WT mice (n = 22) amounting to 113.0±1.8 ml.kg−1.min−1, (P = 0.16). No difference in maximal COX activity between the IL-6−/− and WT mice in m. soleus and m. gastrocnemius was found. Moreover, no impairment of peripheral endothelial function or glucose tolerance was found in IL-6−/− mice. Surprisingly, plasma lactate concentration during running at 8 m.min−1 as well at maximal running velocity in IL-6−/− mice was significantly lower (P<0.01) than in WT mice. Interestingly, IL-6−/− mice displayed important adaptive mechanisms including significantly lower oxygen cost of running at a given speed accompanied by lower expression of sarcoplasmic reticulum Ca2+-ATPase and lower plasma lactate concentrations during running at submaximal and maximal running velocities. In conclusion, impaired endurance running capacity in IL-6−/− mice could not be explained by reduced V′O2max, endothelial dysfunction or impaired muscle oxidative capacity. Therefore, our results indicate that IL-6 cannot be regarded as a major regulator of exercise capacity but rather as a modulator of endurance performance. Furthermore, we identified important compensatory mechanism limiting reduced exercise performance in IL-6−/− mice.

Sarcoplasmic Phospholamban Protein Is Involved in the Mechanisms of Postresuscitation Myocardial Dysfunction and the Cardioprotective Effect of Nitrite during Resuscitation

ObjectivesSarcoplasmic reticulum (SR) Ca2+-handling proteins play an important role in myocardial dysfunction after acute ischemia/reperfusion injury. We hypothesized that nitrite would improve postresuscitation myocardial dysfunction by increasing nitric oxide (NO) generation and that the mechanism of this protection is related to the modulation of SR Ca2+-handling proteins.MethodsWe conducted a randomized prospective animal study using male Sprague-Dawley rats. Cardiac arrest was induced by intravenous bolus of potassium chloride (40 µg/g). Nitrite (1.2 nmol/g) or placebo was administered when chest compression was started. No cardiac arrest was induced in the sham group. Hemodynamic parameters were monitored invasively for 90 minutes after the return of spontaneous circulation (ROSC). Echocardiogram was performed to evaluate cardiac function. Myocardial samples were harvested 5 minutes and 1 hour after ROSC.ResultsMyocardial function was significantly impaired in the nitrite and placebo groups after resuscitation, whereas cardiac function (i.e., ejection fraction and fractional shortening) was significantly greater in the nitrite group than in the placebo group. Nitrite administration increased the level of nitric oxide in the myocardium 5 min after resuscitation compared to the other two groups. The levels of phosphorylated phospholamban (PLB) were decreased after resuscitation, and nitrite increased the phosphorylation of phospholamban compared to the placebo. No significant differences were found in the expression of sarcoplasmic reticulum Ca2+ ATPase (SERCA2a) and ryanodine receptors (RyRs).Conclusionspostresuscitation myocardial dysfunction is associated with the impairment of PLB phosphorylation. Nitrite administered during resuscitation improves postresuscitation myocardial dysfunction by preserving phosphorylated PLB protein during resuscitation.

Abstract 354: Nitrate Improves Postresuscitation Myocardial Dysfunction by Preserving Myocardial Nitric Oxide and Phospholamban Protein

Objectives: Sarcoplasmic reticulum (SR) Ca 2+ handling proteins have been shown to play an important role in myocardial dysfunction after acute ischemic/reperfusion injury. We assessed the hypothesis that nitrite would improve postresuscitation myocardial dysfunction by increasing NO generation, and the mechanism is related to the modulation of SR Ca 2+ handling proteins. Methods: We performed a randomized prospective animal study using male Sprague-Dawley rats. Cardiac arrest was induced by intravenous bolus of potassium chloride (40 μg/g). Nitrite (1.2nmol/g) or placebo was administered when chest compression was started. No cardiac arrest was induced in sham group. Hemodynamic parameters were monitored invasively for one hour after ROSC. Echocardiogram was performed to evaluate cardiac function. Myocardial sample was harvested 5 minutes and 1 hour after ROSC respectively. The expression of sarcoplasmic reticulum Ca 2+ ATPase (SERCA2a), phospholamban (PLB) and rynodine receptors (RyRs) were analyzed by Western blot analysis. Myocardial NO five minutes after ROSC were measured using Nitric Oxide Assay Kit. Results: Myocardial function measured as the maximal rate of LV pressure increase (dP/dtmax), the maximal rate of LV pressure decline (-dP/dtmax), ejection fraction, fractional shortening was significantly impaired in nitrite and placebo groups after resuscitation, and cardiac function in nitrite group was significantly greater than placebo group. Nitrite administration increased the level of nitric oxide in myocardium 5 min after resuscitation when compared to the other two groups. The levels of phosphorylated phospholamban were decreased after resuscitation, and nitrite preserved the phosphorylation of phospholamban in myocardium after resuscitation when compared to placebo group. No significant differences were found in the expressions of myocardial sarcoplasmic reticulum Ca 2+ ATPase and rynodine receptors after resuscitation. Conclusions: Nitrite administered during resuscitation improves postresusctation myocardial dysfunction as an exogenous donor of nitric oxide, and the modulation of phosphorylated phospholamban, an important SR Ca 2+ uptake protein, is involved in the mechanism.

Decreasing Tropomyosin Phosphorylation Rescues Tropomyosin-induced Familial Hypertrophic Cardiomyopathy

Studies indicate that tropomyosin (Tm) phosphorylation status varies in different mouse models of cardiac disease. Investigation of basal and acute cardiac function utilizing a mouse model expressing an α-Tm protein that cannot be phosphorylated (S283A) shows a compensated hypertrophic phenotype with significant increases in SERCA2a expression and phosphorylation of phospholamban Ser-16 (Schulz, E. M., Correll, R. N., Sheikh, H. N., Lofrano-Alves, M. S., Engel, P. L., Newman, G., Schultz Jel, J., Molkentin, J. D., Wolska, B. M., Solaro, R. J., and Wieczorek, D. F. (2012) J. Biol. Chem. 287, 44478-44489). With these results, we hypothesized that decreasing α-Tm phosphorylation may be beneficial in the context of a chronic, intrinsic stressor. To test this hypothesis, we utilized the familial hypertrophic cardiomyopathy (FHC) α-Tm E180G model (Prabhakar, R., Boivin, G. P., Grupp, I. L., Hoit, B., Arteaga, G., Solaro, R. J., and Wieczorek, D. F. (2001) J. Mol. Cell. Cardiol. 33, 1815-1828). These FHC hearts are characterized by increased heart:body weight ratios, fibrosis, increased myofilament Ca(2+) sensitivity, and contractile defects. The FHC mice die by 6-8 months of age. We generated mice expressing both the E180G and S283A mutations and found that the hypertrophic phenotype was rescued in the α-Tm E180G/S283A double mutant transgenic animals; these mice exhibited no signs of cardiac hypertrophy and displayed improved cardiac function. These double mutant transgenic hearts showed increased phosphorylation of phospholamban Ser-16 and Thr-17 compared with the α-Tm E180G mice. This is the first study to demonstrate that decreasing phosphorylation of tropomyosin can rescue a hypertrophic cardiomyopathic phenotype.

GW24-e1015 Nitrate improves postresuscitation myocardial dysfunction and the involvement of nitric oxide and phospholamban protein

ObjectivesThe abnormality of Sarcoplasmic reticulum (SR) Ca2+ handling proteins was shown to be essential in myocardial dysfunction after acute reperfusion injury, which might be attenuated by NO therapy. Cardiac arrest...