Acute hyperglycemia exacerbates myocardial ischemia/reperfusion injury and blunts cardioprotective effect of GIK

There is a close association between hyperglycemia and increased risk of mortality after acute myocardial infarction (AMI). However, whether acute hyperglycemia exacerbates myocardial ischemia/reperfusion (MI/R) injury remains unclear. We observed the effects of acute hyperglycemia on MI/R injury and on the cardioprotective effect of glucose-insulin-potassium (GIK). Male rats were subjected to 30 min of myocardial ischemia and 6 h of reperfusion. Rats were randomly received one of the following treatments (at 4 ml.kg(-1).h(-1) iv): Vehicle, GIK (GIK during reperfusion; glucose: 200g/l, insulin: 60 U/l, KCL: 60 mmol/l), HG (high glucose during ischemia; glucose:500 g/l), GIK + HG (HG during I and GIK during R) or GIK + wortmannin (GIK during R and wortmannin 15 min before R). Blood glucose, plasma insulin concentration and left ventricular pressure (LVP) were monitored throughout the experiments. Hyperglycemia during ischemia not only significantly increased myocardial apoptosis (23.6 +/- 1.7% vs. 18.8 +/- 1.4%, P < 0.05 vs. vehicle), increased infarct size (IS) (45.6 +/- 3.0% vs. 37.6 +/- 2.0%, P < 0.05 vs. vehicle), decreased Akt and GSK-3beta phosphorylations (0.5 +/- 0.2 and 0.6 +/- 0.1% fold of vehicle, respectively, P < 0.05 vs. vehicle) following MI/R, but almost completely blocked the cardioprotective effect afforded by GIK, as evidenced by significantly increased apoptotic index (19.1 +/- 2.0 vs. 10.3 +/- 1.2%, P < 0.01 vs. GIK), increased myocardial IS (39.2 +/- 2.8 vs. 27.2 +/- 2.1%, P < 0.01 vs. GIK), decreased Akt phosphorylation (1.1 +/- 0.1 vs. 1.7 +/- 0.2%, P < 0.01 vs. GIK) and GSK-3beta phosphorylation (1.4 +/- 0.2 vs. 2.3 +/- 0.2%, P < 0.05 vs. GIK). Hyperglycemia significantly exacerbates MI/R injury and blocks the cardioprotective effect afforded by GIK, which is, at least in part, due to hyperglycemia-induced decrease of myocardial Akt activation.     

Ginsenoside Rg1 ameliorates diabetic cardiomyopathy by inhibiting endoplasmic reticulum stress‐induced apoptosis in a streptozotocin‐induced diabetes rat model

Ginsenoside Rg1 has been demonstrated to have cardiovascular protective effects. However, whether the cardioprotective effects of ginsenoside Rg1 are mediated by endoplasmic reticulum (ER) stress‐induced apoptosis remain unclear. In this study, among 80 male Wistar rats, 15 rats were randomly selected as controls; the remaining 65 rats received a diet rich in fat and sugar content for 4 weeks, followed by intraperitoneal injection of streptozotocin (STZ, 40 mg/kg) to establish a diabetes model. Seven days after STZ injection, 10 rats were randomly selected as diabetic model (DM) controls, 45 eligible diabetic rats were randomized to three treatment groups and administered ginsenoside Rg1 in a dosage of 10, 15 or 20 mg/kg/day, respectively. After 12 weeks of treatment, rats were killed and serum samples obtained to determine cardiac troponin (cTn)‐I. Myocardial tissues were harvested for morphological analysis to detect myocardial cell apoptosis, and to analyse protein expression of glucose‐regulated protein 78 (GRP78), C/EBP homologous protein (CHOP), and Caspase‐12. Treatment with ginsenoside Rg1 (10–20 mg/kg) significantly reduced serum cTnI levels compared with DM control group (all P < 0.01). Ginsenoside Rg1 (15 and 20 mg/kg) significantly reduced the percentage of apoptotic myocardial cells and improved the parameters of cardiac function. Haematoxylin and eosin and Masson staining indicated that ginsenoside Rg1 could attenuate myocardial lesions and myocardial collagen volume fraction. Additionally, ginsenoside Rg1 significantly reduced GRP78, CHOP, and cleaved Caspase‐12 protein expression in a dose‐dependent manner. These findings suggest that ginsenoside Rg1 appeared to ameliorate diabetic cardiomyopathy by inhibiting ER stress‐induced apoptosis in diabetic rats.     

AMPK-Regulated and Akt-Dependent Enhancement of Glucose Uptake Is Essential in Ischemic Preconditioning-Alleviated Reperfusion Injury

Aims

Ischemic preconditioning (IPC) is a potent form of endogenous protection. However, IPC-induced cardioprotective effect is significantly blunted in insulin resistance-related diseases and the underlying mechanism is unclear. This study aimed to determine the role of glucose metabolism in IPC-reduced reperfusion injury.

Methods

Normal or streptozotocin (STZ)-treated diabetic rats subjected to 2 cycles of 5 min ischemia/5 min reperfusion prior to myocardial ischemia (30 min)/reperfusion (3 h). Myocardial glucose uptake was determined by ¹⁸F-fluorodeoxyglucose-positron emission tomography (PET) scan and gamma-counter biodistribution assay.

Results

IPC exerted significant cardioprotection and markedly improved myocardial glucose uptake 1 h after reperfusion (P<0.01) as evidenced by PET images and gamma-counter biodistribution assay in ischemia/reperfused rats. Meanwhile, myocardial translocation of glucose transporter 4 (GLUT4) to plasma membrane together with myocardial Akt and AMPK phosphorylation were significantly enhanced in preconditioned hearts. Intramyocardial injection of GLUT4 siRNA markedly decreased GLUT4 expression and blocked the cardioprotection of IPC as evidence by increased myocardial infarct size. Moreover, the PI3K inhibitor wortmannin significantly inhibited activation of Akt and AMPK, reduced GLUT4 translocation, glucose uptake and ultimately, depressed IPC-induced cardioprotection. Furthermore, IPC-afforded antiapoptotic effect was markedly blunted in STZ-treated diabetic rats. Exogenous insulin supplementation significantly improved glucose uptake via co-activation of myocardial AMPK and Akt and alleviated ischemia/reperfusion injury as evidenced by reduced myocardial apoptosis and infarction size in STZ-treated rats (P<0.05).

Conclusions

The present study firstly examined the role of myocardial glucose metabolism during reperfusion in IPC using direct genetic modulation in vivo. Augmented glucose uptake via co-activation of myocardial AMPK and Akt in reperfused myocardium is essential to IPC-alleviated reperfusion injury. This intrinsic metabolic modulation and cardioprotective capacity are present in STZ-treated hearts and can be triggered by insulin.     

Reduction of SIRT1 blunts the protective effects of ischemic post-conditioning in diabetic mice by impairing the Akt signaling pathway

Ischemic post-conditioning (IPO) activates Akt signaling to confer cardioprotection. The responsiveness of diabetic hearts to IPO is impaired. We hypothesized that decreased cardiac SIRT1, a positive regulator of Akt, may be responsible for the impaired responsiveness of diabetic hearts to IPO-mediated cardioprotection. High-fat diet and streptozotocin-induced diabetic mice were subjected to myocardial ischemia/reperfusion (MI/R, 30 min ischemia and 180 min reperfusion) or IPO (three cycles of 10 s of reperfusion and ischemia at the onset of reperfusion). Adenoviral vectors encoding GFP or SIRT1 (Ad-SIRT1) were administered by direct injection into the left ventricular. Our results showed that IPO activated the Akt signaling pathway and reduced MI/R injury in non-diabetic hearts but not in diabetic hearts, in which reduced expression of SIRT1 and increased Akt acetylation were observed. Delivery of Ad-SIRT1 into the diabetic hearts reduced Akt acetylation and restored the cardioprotective effects of IPO by modulating Akt signaling pathway. In contrast, cardiac-specific SIRT1 knockout increased Akt acetylation and blunted the cardioprotective effects of IPO. In in vitro study, transfection with wild-type SIRT1 but not inactive mutant SIRT1 reduced the expression of Akt acetylation and restored the protective effects of hypoxic post-conditioning in high glucose-incubated cardiomyocytes. Moreover, the cardiomyocytes transfected with constitutive Akt acetylation showed repressed Akt phosphorylation and blunted protective effects against hypoxia/reoxygenation injury. These findings demonstrate that the reduction of SIRT1 blunts the protective effects of IPO by impairing Akt signaling pathway and that SIRT1 up-regulation restores IPO-mediated cardioprotection in diabetic mice via deacetylation-dependent activation of Akt signaling pathway.     

Ginsenoside Rd Attenuates Myocardial Ischemia/Reperfusion Injury via Akt/GSK-3β Signaling and Inhibition of the Mitochondria-Dependent Apoptotic Pathway

Evidence suggests Ginsenoside Rd (GSRd), a biologically active extract from the medical plant Panax Ginseng, exerts antioxidant effect, decreasing reactive oxygen species (ROS) formation. Current study determined the effect of GSRd on myocardial ischemia/reperfusion (MI/R) injury (a pathological condition where ROS production is significantly increased) and investigated the underlying mechanisms. The current study utilized an in vivo rat model of MI/R injury and an in vitro neonatal rat cardiomyocyte (NRC) model of simulated ischemia/reperfusion (SI/R) injury. Infarct size was measured by Evans blue/TTC double staining. NRC injury was determined by MTT and lactate dehydrogenase (LDH) leakage assay. ROS accumulation and apoptosis were assessed by flow cytometry. Mitochondrial membrane potential (MMP) was determined by 5, 5′, 6, 6′-tetrachloro-1, 1′, 3, 3′-tetrathylbenzimidazol carbocyanine iodide (JC-1). Cytosolic translocation of mitochondrial cytochrome c and expression of caspase-9, caspase-3, Bcl-2 family proteins, and phosphorylated Akt and GSK-3β were determined by western blot. Pretreatment with GSRd (50 mg/kg) significantly augmented rat cardiac function, as evidenced by increased left ventricular ejection fraction (LVEF) and ±dP/dt. GSRd reduced myocardial infarct size, apoptotic cell death, and blood creatine kinase/lactate dehydrogenase levels after MI/R. In NRCs, GSRd (10 µM) inhibited SI/R-induced ROS generation (P<0.01), decreased cellular apoptosis, stabilized the mitochondrial membrane potential (MMP), and attenuated cytosolic translocation of mitochondrial cytochrome c. GSRd inhibited activation of caspase-9 and caspase-3, increased the phosphorylated Akt and GSK-3β, and increased the Bcl-2/Bax ratio. Together, these data demonstrate GSRd mediated cardioprotective effect against MI/R–induced apoptosis via a mitochondrial-dependent apoptotic pathway.     

胰岛素保护缺血/再灌注心脏:PI3-K/Akt和JNKs信号通路间的交互作用

我们前期研究表明胰岛素可激活细胞内信号转导机制如磷脂酰肌醇3．激酶．蛋白激酶B．内皮型一氧化氮合酶．一氧化氮（P13-K-Akt-eNOS-NO）信号通路,减轻心肌缺血／再灌注（ischemia／reperfusion,I／R）损伤,改善缺血后心肌功能恢复。然而c-Jun氨基末端激酶（c-JunNH2-terminal kinase,JNK）信号通路在胰岛素保护I／R心肌中的作用尚不清楚,本研究旨在探讨JNK信号通路在胰岛素保护I／R心肌中的作用及其与P13．K／Akt信号通路间的相互关系。离体Sprague-Dawley大鼠心脏缺血30min后施行2h或4h的再灌注,缺血前用LY294002（15mmol／L）和SP600125（10mmol／L）灌注15min,分别阻断P13．K／Akt和磷酸化JNK（phosphorylated．JNK,p-JNK）活化,观测心脏功能、心肌梗死、细胞凋亡和蛋白磷酸化水平。与对照组相比,胰岛素再灌注2h后,心率、左心室发展压和左心室收缩／舒张最大速率均明显增加,梗死面积减少约16．1％[（28．9±2．0）％vs（45．0±4．0）％,n=6,P〈O．01],细胞凋亡指数从（27．6±113）％减少到（16．0±0．7）％（n=6,P〈O．01）,Akt的活性增加1．7倍（n=6,P〈0．05）,同时JNK活性增加1．5倍铆=6,P〈O．05）。用LY294002处理后,胰岛素对I／R心肌的保护作用消失;而用SP600125处理可增强胰岛素的保护作用,且可部分逆转LY294002的抑制作用。进一步观察发现SP600125减弱了Akt的磷酸化m=6,P〈0．05）。上述结果表明,在I／R心肌中,胰岛素可同时激活P13．K／Akt及JNK信号通路,且通过后者进一步增加Akt活化,从而减轻I／R损伤,改善心肌功能。这种P13．K／Akt与JNK信号通路交互机制对胰岛素保护I／R心肌有重要意义。     

Dynamic alteration of adiponectin/adiponectin receptor expression and its impact on myocardial ischemia/reperfusion in type 1 diabetic mice

The present study determined the dynamic change of adiponectin (APN, a cardioprotective adipokine), its receptor expression, and their impact upon myocardial ischemia/reperfusion (MI/R) injury during type 1 diabetes mellitus (T1DM) progression, and involved underlying mechanisms. Diabetic state was induced in mice via multiple intraperitoneal injections of low-dose streptozotocin. The dynamic change of plasma APN concentration and cardiac APN receptor-1 and -2 (AdipoR1/2) expression were assessed immediately after diabetes onset (0 wk) and 1, 3, 5, and 7 wk thereafter. Indicators of MI/R injury (infarct size, apoptosis, and LDH release) were determined at 0, 1, and 7 wk of DM duration. The effect of APN on MI/R injury was determined in mice subjected to different diabetic durations. Plasma APN levels (total and HMW form) increased, whereas cardiac AdipoR1 expression decreased early after T1DM onset. With T1DM progression, APN levels were reduced and cardiac AdipoR1 expression increased. MI/R injury was exacerbated with T1DM progression in a time-dependent manner. Administration of globular APN (gAD) failed to attenuate MI/R injury in 1-wk T1DM mice, while an AMP-activated protein kinase (AMPK) activator (AICAR) reduced MI/R injury. However, administration of gAD (and AICAR) reduced infarct size and cardiomyocyte apoptosis in 7-wk T1DM mice. In conclusion, our results demonstrate a dynamic dysfunction of APN/AdipoR1 during T1DM progression. Reduced cardiac AdipoR1 expression and APN concentration may be responsible for increased I/R injury susceptibility at early and late T1DM stages, respectively. Interventions bolstering AdipoR1 expression during early T1DM stages and APN supplementation during advanced T1DM stages may potentially reduce the myocardial ischemic injury in diabetic patients.     

Toll-like receptor 3 plays a role in myocardial infarction and ischemia/reperfusion injury

Innate immune and inflammatory responses mediated by Toll like receptors (TLRs) have been implicated in myocardial ischemia/reperfusion (I/R) injury. This study examined the role of TLR3 in myocardial injury induced by two models, namely, myocardial infarction (MI) and I/R. First, we examined the role of TLR3 in MI. TLR3 deficient (TLR3^−/−) and wild type (WT) mice were subjected to MI induced by permanent ligation of the left anterior descending (LAD) coronary artery for 21 days. Cardiac function was measured by echocardiography. Next, we examined whether TLR3 contributes to myocardial I/R injury. TLR3^−/− and WT mice were subjected to myocardial ischemia (45 min) followed by reperfusion for up to 3 days. Cardiac function and myocardial infarct size were examined. We also examined the effect of TLR3 deficiency on I/R-induced myocardial apoptosis and inflammatory cytokine production. TLR3^−/− mice showed significant attenuation of cardiac dysfunction after MI or I/R. Myocardial infarct size and myocardial apoptosis induced by I/R injury were significantly attenuated in TLR3^−/− mice. TLR3 deficiency increases B-cell lymphoma 2 (BCL2) levels and attenuates I/R-increased Fas, Fas ligand or CD95L (FasL), Fas-Associated protein with Death Domain (FADD), Bax and Bak levels in the myocardium. TLR3 deficiency also attenuates I/R-induced myocardial nuclear factor KappaB (NF-κB) binding activity, Tumor necrosis factor alpha (TNF-α) and Interleukin-1 beta (IL-1β) production as well as I/R-induced infiltration of neutrophils and macrophages into the myocardium. TLR3 plays an important role in myocardial injury induced by MI or I/R. The mechanisms involve activation of apoptotic signaling and NF-κB binding activity. Modulation of TLR3 may be an effective approach for ameliorating heart injury in heart attack patients.     

Post-infarct treatment with [Pyr1]-apelin-13 reduces myocardial damage through reduction of oxidative injury and nitric oxide enhancement in the rat model of myocardial infarction

Apelin is a newly discovered peptide that has been recently shown to have cardioprotective effects in the animal model of myocardial infarction (MI) and ischemia/reperfusion (I/R) injuries. The aim of the present study was to investigate the long term cardioprotective effect of [Pyr1]-apelin-13 in the rat model of MI. Male Wistar rats (n = 22) were randomly divided into three groups: (1) sham operated group (2) control MI group and (3) MI treated with apelin (MI-AP group). MI animals were subjected to 30 min of left anterior descending coronary artery (LAD) ligation and 14 days of reperfusion. 24 h after LAD ligation, apelin (10 nmol/kg/day) was administered i.p. for 5 days. Blood sampling was performed at days 1, 3, 5 and 7 after MI for determination of serum changes of lactate dehydrogenase (LDH), creatine kinase-MB (CK-MB), malondialdehyde (MDA) and nitric oxide (NO). Myocardial infarct size (IS) and hemodynamic function were also measured at the end of the study at day 14. We found out that post infarct treatment with apelin decreases infarct size, serum levels of LDH, CK-MB and MDA and increases heart rate and serum level of NO in the consecutive days, but there were no significant differences in blood pressure in the MI-AP group in comparison with MI. In conclusion, apelin has long term cardioprotective effects against myocardial infarction through attenuation of cardiac tissue injury and lipid peroxidation and enhancement of NO production.     

α-Lipoic Acid Reduces Infarct Size and Preserves Cardiac Function in Rat Myocardial Ischemia/Reperfusion Injury through Activation of PI3K/Akt/Nrf2 Pathway

Background

The present study investigates the effects and mechanisms of α-Lipoic acid (LA) on myocardial infarct size, cardiac function and cardiomyocyte apoptosis in rat hearts subjected to in vivo myocardial ischemia/reperfusion (MI/R) injury.

Methodology/Principal Findings

Male adult rats underwent 30 minutes of ischemia followed by 3, 24, or 72 h of reperfusion. Animals were pretreated with LA or vehicle before coronary artery ligation. The level of MI/R- induced LDH and CK release, infarct size, cardiomyocyte apoptosis and cardiac functional impairment were examined and compared. Western blot analysis was performed to elucidate the mechanism of LA pretreatment. The level of inflammatory cytokine TNF-α released to serum and accumulated in injured myocardium as well as neutrophil accumulation in injured myocardium were also examined after MI/R injury. Our results reveal that LA administration significantly reduced LDH and CK release, attenuated myocardial infarct size, decreased cardiomyocytes apoptosis, and partially preserved heart function. Western blot analysis showed that LA pretreatment up-regulated Akt phosphorylation and Nrf2 nuclear translocation while producing no impact on p38MAPK activation or nitric oxide (NO) production. LA pretreatment also increased expression of HO-1, a major target of Nrf2. LA treatment inhibited neutrophil accumulation and release of TNF-α. Moreover, PI3K inhibition abolished the beneficial effects of LA.

Conclusions/Significance

This study indicates that LA attenuates cardiac dysfunction by reducing cardiomyoctyes necrosis, apoptosis and inflammation after MI/R. LA exerts its action by activating the PI3K/Akt pathway as well as subsequent Nrf2 nuclear translocation and induction of cytoprotective genes such as HO-1.     

Long-term aerobic exercise protects the heart against ischemia/reperfusion injury via PI3 kinase-dependent and Akt-mediated mechanism

Objective Physical activity has been shown to improve cardiovascular function and to be beneficial to type 2 diabetic patients. However, the effects of aerobic exercise (AE) on myocardial ischemia/reperfusion (MI/R) are largely unclear. Therefore, the aims of the present study were to determine whether long-term AE can protect the heart against I/R injury, and if so, to investigate the underlying mechanism. Methods Adult male Sprague–Dawley rats were randomly subjected to 8 weeks of either sedentary or free-loading swimming exercise (3 h/day, 5 d/week). Then the animals were subjected to 30 min MI followed by 4 h R. Arterial blood pressure and left ventricular pressure (LVP) were monitored throughout the whole MI/R procedure. Plasma creatine kinase (CK) and lactate dehydrogenase (LDH) activities were measured spectrophotometrically. Myocardial infarction and myocardial apoptosis (TUNEL analysis) were determined in a blinded manner. Results MI/R caused significant cardiac dysfunction and myocardial apoptosis (strong TUNEL-positive staining). Compared with sedentary group, rats subjected to 8 weeks of AE showed protection against MI/R as evidenced by reduced myocardial infarction (26.8 ± 1.5% vs. 35.3 ± 2.4%, n = 8, P < 0.05), inhibited cardiomyocyte apoptosis (decreased apoptotic index (12.4 ± 1.1% vs. 21.0 ± 1.7%, n = 8, P < 0.01) and decreased myocardial caspase-3 activity), decreased plasma CK and LDH activities and improved recovery of cardiac systolic/diastolic function (including LVSP and ±LVdP/dt) at the end of R. Moreover, exercise resulted in 1.7-fold, 2.5-fold and 2.5-fold increases in Akt expression, Akt phosphorylation and glycogen synthase kinase-3β phosphorylation in I/R myocardium, respectively (n = 3, all P < 0.05). More importantly, treatment with wortmannin, a PI3 kinase inhibitor, 15 min before R not only significantly blocked Akt phosphorylation (P < 0.05) in exercise rats, but also abolished long-term AE-induced cardioprotection for the I/R heart as manifested by increased apoptosis and myocardial infarction, and reduced cardiac function. Conclusion Long-term AE exerts cardioprotective effect against MI/R injury, including anti-cardiomyocyte apoptosis, which is at least partly via PI3 kinase-dependent and Akt-mediated mechanism.     

Sulodexide attenuates endoplasmic reticulum stress induced by myocardial ischaemia/reperfusion by activating the PI3K/Akt pathway

Acute myocardial ischaemia/reperfusion (MI/R) injury causes severe arrhythmias with a high rate of lethality. Extensive research focus on endoplasmic reticulum (ER) stress and its dysfunction which leads to cardiac injury in MI/R Our study evaluated the effects of sulodexide (SDX) on MI/R by establishing MI/R mice models and in vitro oxidative stress models in H9C2 cells. We found that SDX decreases cardiac injury during ischaemia reperfusion and decreased myocardial apoptosis and infarct area, which was paralleled by increased superoxide dismutase and reduced malondialdehyde in mice plasm, increased Bcl‐2 expression, decreased BAX expression in a mouse model of MI/R. In vitro, SDX exerted a protective effect by the suppression of the ER stress which induced by tert‐butyl hydroperoxide (TBHP) treatment. Both of the in vivo and in vitro effects were involved in the phosphatidylinositol 3‐kinase (PI3K)/Akt signalling pathway. Inhibition of PI3K/Akt pathway by specific inhibitor, LY294002, partially reduced the protective effect of SDX. In short, our results suggested that the cardioprotective role of SDX was related to the suppression of ER stress in mice MI/R models and TBHP‐induced H9C2 cell injury which was through the PI3K/Akt signalling pathway.     

TNF-α inhibitor protects against myocardial ischemia/reperfusion injury via Notch1-mediated suppression of oxidative/nitrative stress

TNF-α inhibitor reportedly protects against myocardial ischemia/reperfusion (MI/R) injury. It can also increase Notch1 expression in inflammatory bowel disease, revealing the regulation of Notch1 signaling by TNF-α inhibitor. However, the interaction between TNF-α inhibitor and Notch1 signaling in MI/R remains unclear. This study aimed to determine the involvement of TNF-α inhibitor with Notch1 in MI/R and delineate the related mechanism. Notch1-specific small interfering RNA (20 μg) or Jagged1 (a Notch ligand, 12 μg) was delivered through intramyocardial injection. Forty-eight hours after injection, mice received 30 min of myocardial ischemia followed by 3 h (for cell apoptosis and oxidative/nitrative stress) or 24 h (for infarct size and cardiac function) of reperfusion. Ten minutes before reperfusion, mice randomly received an intraperitoneal injection of vehicle, etanercept, diphenyleneiodonium, 1400W, or EUK134. Finally, downregulation of Notch1 significantly reversed the alleviation of MI/R injury induced by etanercept, as evidenced by enlarged myocardial infarct size, suppressed cardiac function, and increased myocardial apoptosis. Moreover, Notch1 blockade increased the expression of inducible NO synthase (iNOS) and gp^91phox, enhanced NO and superoxide production, and accelerated their cytotoxic reaction product, peroxynitrite. Furthermore, NADPH inhibition with diphenyleneiodonium or iNOS suppression with 1400W mitigated the aggravation of MI/R injury induced by Notch1 downregulation in mice treated with etanercept. Additionally, either Notch1 activation with Jagged1 or peroxynitrite decomposition with EUK134 reduced nitrotyrosine content and attenuated MI/R injury. These data indicate that MI/R injury can be attenuated by TNF-α inhibitor, partly via Notch1 signaling-mediated suppression of oxidative/nitrative stress.     

Acute hyperglycaemia enhances oxidative stress and aggravates myocardial ischaemia/reperfusion injury: role of thioredoxin‐interacting protein

Hyperglycaemia during acute myocardial infarction is common and associated with increased mortality. Thioredoxin‐interacting protein (Txnip) is a modulator of cellular redox state and contributes to cell apoptosis. This study aimed to investigate whether or not hyperglycaemia enhances Txnip expression in myocardial ischaemia/reperfusion (MI/R) and consequently exacerbates MI/R injury. Rats were subjected to 30 min. of left coronary artery ligation followed by 4 hrs of reperfusion and treated with saline or high glucose (HG, 500 g/l, 4 ml/kg/h intravenously). In vitro study was performed on cultured rat cardiomyocytes subjected to simulated ischaemia/reperfusion (SI/R) and incubated with HG (25 mM) or normal glucose (5.6 mM) medium. In vivo HG infusion during MI/R significantly impaired cardiac function, aggravated myocardial injury and increased cardiac oxidative stress. Meanwhile, Txnip expression was enhanced whereas thioredoxin activity was inhibited following HG treatment in ischaemia/reperfusion (I/R) hearts. In addition, HG activated p38 MAPK and inhibited Akt in I/R hearts. In cultured cardiomyocytes subjected to SI/R, HG incubation stimulated Txnip expression and reduced thioredoxin activity. Overexpression of Txnip enhanced HG‐induced superoxide generation and aggravated cardiomyocyte apoptosis, whereas Txnip RNAi significantly blunted the deleterious effects of HG. Moreover, inhibition of p38 MAPK or activation of Akt markedly blocked HG‐induced Txnip expression in I/R cardiomyocytes. Most importantly, intramyocardial injection of Txnip siRNA markedly decreased Txnip expression and alleviated MI/R injury in HG‐treated rats. Hyperglycaemia enhances myocardial Txnip expression, possibly through reciprocally modulating p38 MAPK and Akt activation, leading to aggravated oxidative stress and subsequently, amplification of cardiac injury following MI/R.     

Luteolin limits infarct size and improves cardiac function after myocardium ischemia/reperfusion injury in diabetic rats

Background

The present study was to investigate the effects and mechanism of Luteolin on myocardial infarct size, cardiac function and cardiomyocyte apoptosis in diabetic rats with myocardial ischemia/reperfusion (I/R) injury.

Methodology/Principal Findings

Diabetic rats underwent 30 minutes of ischemia followed by 3 h of reperfusion. Animals were pretreated with or without Luteolin before coronary artery ligation. The severity of myocardial I/R induced LDH release, arrhythmia, infarct size, cardiac function impairment, cardiomyocyte apoptosis were compared. Western blot analysis was performed to elucidate the target proteins of Luteolin. The inflammatory cytokine production were also examined in ischemic myocardium underwent I/R injury. Our results revealed that Luteolin administration significantly reduced LDH release, decreased the incidence of arrhythmia, attenuated myocardial infarct size, enhanced left ventricular ejection fraction and decreased myocardial apoptotic death compared with I/R group. Western blot analysis showed that Luteolin treatment up-regulated anti-apoptotic proteins FGFR2 and LIF expression, increased BAD phosphorylation while decreased the ratio of Bax to Bcl-2. Luteolin treatment also inhibited MPO expression and inflammatory cytokine production including IL-6, IL-1a and TNF-a. Moreover, co-administration of wortmannin and Luteolin abolished the beneficial effects of Luteolin.

Conclusions/Significance

This study indicates that Luteolin preserves cardiac function, reduces infarct size and cardiomyocyte apoptotic rate after I/R injury in diabetic rats. Luteolin exerts its action by up-regulating of anti-apoptotic proteins FGFR2 and LIF expression, activating PI3K/Akt pathway while increasing BAD phosphorylation and decreasing ratio of Bax to Bcl-2.     

CpG-ODN,the TLR9 agonist,attenuates myocardial ischemia/reperfusion injury: Involving activation of PI3K/Akt signaling

BackgroundToll-like receptors (TLRs) have been implicated in myocardial ischemia/reperfusion (I/R) injury. The TLR9 ligand, CpG-ODN has been reported to improve cell survival. We examined effect of CpG-ODN on myocardial I/R injury.MethodsMale C57BL/6 mice were treated with either CpG-ODN, control-ODN, or inhibitory CpG-ODN (iCpG-ODN) 1 h prior to myocardial ischemia (60 min) followed by reperfusion. Untreated mice served as I/R control (n = 10/each group). Infarct size was determined by TTC straining. Cardiac function was examined by echocardiography before and after myocardial I/R up to 14 days.ResultsCpG-ODN administration significantly decreased infarct size by 31.4% and improved cardiac function after myocardial I/R up to 14 days. Neither control-ODN nor iCpG-ODN altered I/R-induced myocardial infarction and cardiac dysfunction. CpG-ODN attenuated I/R-induced myocardial apoptosis and prevented I/R-induced decrease in Bcl2 and increase in Bax levels in the myocardium. CpG-ODN increased Akt and GSK-3β phosphorylation in the myocardium. In vitro data suggested that CpG-ODN treatment induced TLR9 tyrosine phosphorylation and promoted an association between TLR9 and the p85 subunit of PI3K. Importantly, PI3K/Akt inhibition and Akt kinase deficiency abolished CpG-ODN-induced cardioprotection.ConclusionCpG-ODN, the TLR9 ligand, induces protection against myocardial I/R injury. The mechanisms involve activation of the PI3K/Akt signaling pathway.     

κ-Opioid receptor stimulation modulates TLR4/NF-κB signaling in the rat heart subjected to ischemia–reperfusion

It is well documented that the Toll-like receptor 4 (TLR4)/NF-κB signaling mediates early inflammation during myocardial ischemia and reperfusion. Our previous study has demonstrated that κ-opioid receptor stimulation with U50,488H produces cardioprotective and anti-inflammatory effects. The aim of the present study was to investigate whether κ-opioid receptor stimulation could modulate the TLR4/NF-κB signaling and reduce neutrophil accumulation and TNF-α induction in an ischemia–reperfusion injured rat heart model. Rats were randomly exposed to sham operation, myocardial ischemia and reperfusion (MI/R), and MI/R + U50,488H in the absence or presence of Nor-BNI, a selective κ-opioid receptor antagonist. The results demonstrated that after MI/R, the expressions of myocardial TLR4 and NF-κB increased significantly both in ischemia area and risking area. Compared with MI/R, κ-opioid receptor stimulation with U50,488H significantly attenuated the expressions of TLR4 and NF-κB. At the mean time, it also reduced myeloperoxidase (MPO) levels, both serum and myocardial TNF-α production, myocardial infarct sizes (INF/AAR%) and myocardial apoptosis induced by MI/R, all the effects of U50,488H were abolished by Nor-BNI. These data provide evidence for the first time that κ-opioid receptor stimulation inhibits TLR4/NF-κB signaling in the rat heart subjected to MI/R.     

Niacin bound chromium treatment induces myocardial Glut-4 translocation and caveolar interaction via Akt,AMPK and eNOS phosphorylation in streptozotocin induced diabetic rats after ischemia-reperfusion injury

Diabetes, one of the major risk factors of metabolic syndrome culminates in the development of Ischemic Heart Disease (IHD). Refined diets that lack micronutrients, mainly trivalent chromium (Cr³⁺) have been identified as the contributor in the rising incidence of diabetes. We investigated the effect of niacin-bound chromium (NBC) during ischemia/reperfusion (IR) injury in streptozotocin induced diabetic rats. Rats were randomized into: Control (Con); Diabetic (Dia) and Diabetic rats fed with NBC (Dia + NBC). After 30 days of treatment, the isolated hearts were subjected to 30 min of global ischemia followed by 2 h of reperfusion. NBC treatment demonstrated significant increase in left ventricular functions and significant reduction in infarct size and cardiomyocyte apoptosis in Dia + NBC compared with Dia. Increased Glut-4 translocation to the lipid raft fractions was also observed in Dia + NBC compared to Dia. Reduced Cav-1 and increased Cav-3 expression along with phosphorylation of Akt, eNOS and AMPK might have resulted in increased Glut-4 translocation in Dia + NBC. Our results indicate that the cardioprotective effect of NBC is mediated by increased activation of AMPK, Akt and eNOS resulting in increased translocation of Glut-4 to the caveolar raft fractions thereby alleviating the effects of IR injury in the diabetic myocardium.