Effects of radical scavengers,TA248 and TA276, on stunned myocardium in dogs: involvement of K ATP channels

TA248 (7-(beta-D-glucopyranosyloxy)-4-hydroxy-3-octyloxy-2H-1-benzopyran-2-one) and TA276 (sodium 7-hydroxy-3-octyloxy-2H-1-benzopyran-2-one-4-oxide) were newly developed as radical scavengers. In vitro, TA276 scavenged both superoxide anions (. O(2)(-)) and hydroxyl radicals (. OH). TA248 also trapped. O(2)(-), but had less activity on. OH. In vivo, left ventricular contractile functions were determined in pentobarbital-anesthetized open-chest dogs. A regional portion of the left ventricular wall was made ischemic for 20 min by ligating the left anterior descending coronary artery and then reperfused for 60 min. TA248 (3 mg/kg) and TA276 (3 mg/kg) injected i.v. 10 min before occlusion significantly improved myocardial stunning that is contractile dysfunction observed after reperfusion following brief ischemia. Glibenclamide (1 mg/kg) injected i.v. 20 min before occlusion significantly worsened the myocardial stunning. Pretreatment with glibenclamide completely abolished the beneficial effect of TA276 on myocardial stunning, whereas it only partially attenuated that of TA248, showing some improvement even in the presence of glibenclamide. Because of the incomplete scavenging activity of TA248, residual. OH may play some roles in improvement of myocardial stunning with TA248 in the presence of glibenclamide. We speculate that the. OH may eject glibenclamide from its binding site on K(ATP) channels, leading to opening of the channels.     

Beneficial effects of nafazatrom on ischemic reperfused myocardium

The effect of nafazatrom, a new antithrombotic agent, was studied in a canine model of regional myocardial ischemia. Nafazatrom was administered 1 mg/kg intravenously every 6 h for 48 h. After 24 h of drug or placebo administration, animals underwent 90 min of occlusion of the proximal left circumflex coronary artery followed by gradual reperfusion over a period of 30 min. Twenty-four hours later, the animals were sacrificed and infarct size was determined by histochemical staining with triphenyltetrazolium chloride. Nafazatrom-treated animals had a significant reduction in infarct size expressed as a percent of the anatomical area at risk for infarction: 21 +/- 5% in the treated group vs. 41 +/- 5% in the control group (X +/- S.E.M., P less than 0.05). Histological examination confirmed the gross results of postmortem histochemical staining. Salvage of ischemically jeopardized tissue appeared to be unrelated to myocardial oxygen demand as there were no hemodynamic differences between groups. The beneficial effects of nafazatrom are presumably related to a limitation of autolytic processes on the heart during and after ischemia as a result of the drug's ability to inhibit lipoxygenase and to prevent the enzymatic degradation of prostacyclin.     

Radical scavenging properties of adenosine and derivatives in vitro

The radical scavenging properties of adenosine and related nucleosides were determined. Various protocols were used to assess the reactivity of the compounds toward hydroxyl radicals, superoxide ions, and hypochlorous acid. In the 2-deoxy-D-ribose assay (hydroxyl radicals) all nucleosides tested were equally effective in the presence of EDTA, with rate constants between 10⁹ and 10¹⁰ M⁻¹·sec⁻¹. However, in the absence of EDTA, resembling in vivo conditions, large differences were observed. An intact cis diol function in the sugar moiety of the compounds appeared important to retain full scavenging activity, probably due to firm complexation of iron ions. The reactivity toward hydroxyl radicals was confirmed in electron spin resonance (ESR) experiments with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin-trapping agent. Superoxide ions and hypochlorous acid did not react with the nucleosides. From the results it is hypothesized that the radical scavenging properties of adenosine play a role in its established cardioprotective actions in vivo. © 1996 Wiley-Liss, Inc.     

赤土茯苓苷对离体大鼠心脏缺血再灌注损伤的保护作用

采用 Langendorff离体大鼠非循环灌流模型测定赤土茯苓苷对离体大鼠缺血再灌注损伤时心肌收缩力、冠脉阻力、心率的影响及其抗脂质过氧化作用。结果 2 mg/L,1 0 mg/L,50 mg/L赤土茯苓苷可保护缺血再灌注心肌超氧化物歧化酶与硒谷胱甘肽过氧化物酶 ,降低脂质过氧化产物丙二醛的含量 ,能增加再灌后冠脉流量、冠脉阻力 ,促进心肌收缩幅度的恢复 ,减轻心脏水肿 ,但对心率的影响未见统计学差异     

The administration of alpha-melanocyte-stimulating hormone protects the ischemic/reperfused myocardium

The contribution of alpha-melanocyte-stimulating hormone (alpha-MSH) treatment, an active fragment of adrenocorticotropic hormone (ACTH), to the recovery of postischemic cardiac function, infarct size, the incidence of reperfusion-induced ventricular fibrillation and apoptotic cell death was studied in ischemic/reperfused isolated rat hearts. Rats were subcutaneously injected with 40, 200 and 400 microg/kg of alpha-MSH, and 12 h later, hearts were isolated, perfused and subjected to 30 min of ischemia followed by 120 min of reperfusion. Thus, after 120 min of reperfusion, with the concentration of 200 microg/kg alpha-MSH, coronary flow, aortic flow and left ventricular developed pressure were significantly improved from their control values of 14.6+/-0.6 ml/min, 7.5+/-0.5 ml/min and 9.1+/-0.4 kPa to 20.2+/-0.4 ml/min (p<0.05), 31.5+/-0.9 ml/min (p<0.05) and 15.9+/-0.6 (p<0.05) kPa, respectively. With the doses of 40, 200 and 400 microg/kg of alpha-MSH, infarct size was reduced from its control value of 38+/-5% to 33+/-6% (NS), 17+/-3% (p<0.05) and 19+/-4% (p<0.05), respectively. The reduction in the incidence of reperfusion-induced ventricular fibrillation followed the same pattern. It is reasonable to assume that a reduction in infarct size, in the alpha-MSH-treated myocardium, resulted in a reduction as well in apoptotic cell death. Although we did not specifically study the exact mechanism(s) of alpha-MSH-afforded postischemic protection, we assume that this protection may be related to alpha-MSH-induced corticosterone release and corticosterone-induced de novo protein synthesis, which reflected in the recovery of postischemic cardiac function in isolated hearts. Thus, interventions that are able to increase plasma corticosterone or glucocorticoid release may prevent the development of ischemia/reperfusion-induced damage.     

NHE1抑制剂对缺血及再灌流心肌的保护作用

心肌缺血时 ,由于细胞内 pH值下降 ,激活了Na+/H+交换泵 (NHE) ,细胞内Na+浓度升高 ,促进Na+ Ca2 +交换 ,最终导致细胞内Ca2 +超负荷 ,造成细胞损伤。NHE1抑制剂可通过抑制Na+/H+交换 ,防止细胞内Ca2 +超负荷及其他相关作用 ,对缺血再灌注心脏起保护作用 ,改善心肌功能。目前部分NHE1抑制剂已进行临床研究。     

Calpain inhibitor-1 reduces infarct size and DNA fragmentation of myocardium in ischemic/reperfused rat heart

Myocardial ischemia/reperfusion activates a calcium-dependent protease, calpain, in the ischemic myocytes. It is not known whether calpain is involved in the mechanism of ischemia/reperfusion injury in hearts. Thus the purpose of this study was to clarify the effect of a selective calpain inhibitor (CAI) on infarct size and the extent of DNA damage in ischemic/reperfused rat hearts. Rats were divided in four groups (n = 7 each). In saline group, 0.3 ml of saline was administered (i.v.) 10 min before 30-min coronary occlusion followed by 6-h reperfusion. In vehicle group, 0.3 ml of 10% dimethyl sulfoxide (DMSO) was administered 10 min before the 30-min ischemia. CAI (0.5 mg/kg) was administered 10 min before the 30-min ischemia (CAI-A group) and 10 min before the 6-h reperfusion period (CAI-B group). Infarct size was detected with triphenyl tetrazolium chloride, and DNA fragmentation was detected by agarose gel electrophoresis and by in situ nick end labeling (ISEL). Infarct size was significantly smaller in the CAI-A group compared with the vehicle group (13+/-9% vs. 48+/-12%; p < 0.01), and the incidence of ISEL-positive myocyte nuclei in the subendocardial region was significantly reduced in the CAI-A group compared with the vehicle group (26+/-3% vs. 59+/-6%; p < 0.01). However, the effects of CAI in CAI-B group were not significant. Activation of calpain is involved in the mechanism of ischemia/reperfusion injury, and the preischemic administration of CAI was effective in reducing myocardial infarct size and the DNA damage of the myocytes in ischemic/reperfused rat heart.     

Anti-apoptotic effect and the mechanism of orientin on ischaemic/reperfused myocardium

We investigated the anti-apoptotic effect of orientin, from bamboo leaves (Phyllostachys nigra), on rat heart after treatment with ischemia/reperfusion (I/R), and on rat cardiomyocytes injured by hypoxia/reoxygenation (H/R). I/R injury was induced by occluding the left anterior descending coronary artery for 45 min and restoring perfusion for 240 min. Orientin (0.5, 1.0 and 2.0 mg kg^- 1) or its vehicle was injected i.v. 10 min prior to ischemia. Cultured cardiomyocytes were subjected to hypoxia for 120 min, then reoxygenated for 60 min to induce H/R. Vehicle or orientin (3, 10, 30 μmol l^- 1) was added 10 min before hypoxia and reoxygenated. TUNEL assay and DNA fragmentation assay demonstrated that myocardium apoptosis was attenuated by pretreatment with orientin (0.5, 1.0 and 2.0 mg kg^- 1). Flow cytometric analysis also showed that apoptosis of cardiomyocytes was reduced by pretreatment with orientin (3, 10 and 30 μmol l^- 1). In addition, results of immunohistochemistry and Western blot analysis showed that orientin increased the expression of bcl-2 and reduced Bax expression, resulting in up-regulation of the bcl-2/Bax ratio. Cytochrome c (Cyt-c) and caspase-3 expression was also reduced in myocardium and cardiomyocytes injured by I/R and H/R. These observations indicate that orientin exerts a potent cardioprotective effect on I/R- and H/R-treated myocardium and cardiomyocytes, and inhibits apoptosis by preventing activation of the mitochondrial apoptotic pathway (cytochrome c-caspase-3).     

Antioxidant properties of calcium dobesilate in ischemic/reperfused diabetic rat retina.

Calcium dobesilate possesses antioxidant properties and protects against capillary permeability by reactive oxygen species in the rat peritoneal cavity, but whether a similar action can take place in the diabetic rat retina is unknown. We investigated the oral treatment of diabetic rats with calcium dobesilate on the prevention of free radical-mediated retinal injury induced by ischemia/reperfusion (90 min ischemia followed by 3 min and/or 24 h of reperfusion). Streptozotocin-induced diabetic rats were orally treated with 50 and 100 mg/kg of calcium dobesilate for 10 days (n=12 in each group). In the first series of studies, calcium dobesilate was found to significantly reduce the maldistribution of ion content in diabetic ischemic/reperfused rat retina. Thus, in diabetic rats treated with 100 mg/kg/day calcium dobesilate, ischemia/reperfusion provoked: (i) 27.5% increase in retinal Na(+) content compared to 51.8% in the vehicle-treated group (P<0.05), and (ii) 59.6% increase in retinal Ca(2+) content compared to 107.1% in vehicle-treated animals (P<0.05). In the second series of studies, calcium dobesilate was found to significantly protect diabetic rat retina against inhibition of Na(+)/K(+)-ATPase and Ca(2+)/Mg(2+)-ATPase activities by ischemia/reperfusion (54% and 41% reduction, respectively, with 100 mg/kg of calcium dobesilate) and also against changes in retinal ATP, reduced glutathione (GSH), and oxidized glutathione (GSSG) contents. In the third series of experiments, rats treated with 100 mg/kg of calcium dobesilate reduced the hydroxyl radical signal intensity to 41% (measured by electron paramagnetic resonance), induced by ischemia/reperfusion in diabetic rat retina. Finally, 100 mg/kg calcium dobesilate significantly reduced retinal edema (measured by the thickness of the inner plexiform layer) in diabetic rats. In conclusion, oral treatment with calcium dobesilate significantly protected diabetic rat retina against oxidative stress induced by ischemia/reperfusion. Whether the antioxidant properties of calcium dobesilate explain, at least in part, its beneficial therapeutic effects in diabetic retinopathy deserves further investigation.     

Physiologically tolerable insulin reduces myocardial injury and improves cardiac functional recovery in myocardial ischemic/reperfused dogs

This study was designed to examine whether physiologically tolerable insulin, which maintains lower blood glucose, can protect the myocardium against ischemia/reperfusion (I/R) injury in a preclinical large animal model. Adult dogs were subjected to 50 minutes of myocardial ischemia (80% reduction in coronary blood flow) followed by 4 hours of reperfusion and treated with vehicle, glucose-insulin-potassium (GIK; glucose, 250 g/L; insulin, 60 U/L; potassium, 80 mmol/L), GK, or low-dose insulin (30 U/L) 10 minutes before reperfusion. Treatment with GIK exerted significant cardioprotective effects as evidenced by improved cardiac function, improved coronary blood flow, reduced infarct size, and myocardial apoptosis. In contrast, treatment with GK increased blood glucose level and aggravated myocardial I/R injury. It is interesting that treatment with insulin alone at the dose that reduced blood glucose to a clinically tolerable level exerted significant cardioprotective effects that were comparable to that seen in the GIK-treated group. This low-dose insulin had no effect on coronary blood flow after reperfusion but markedly reduced coronary reactive hyperemia and switched myocardial substrate uptake from fat to carbohydrate. Our results suggest that lower glucose supply to the ischemic myocardium at early reperfusion may create a "metabolic postconditioning" and thus reduce myocardial ischemia/reperfusion injury after prolonged reperfusion.     

Antiapoptotic mechanisms of benidipine in the ischemic/reperfused heart

1. Considerable evidence indicates that calcium plays a critical role in apoptosis. We have previously shown that benidipine, a vasodilatory calcium channel blocker, attenuates postischemia myocardial apoptosis. The present study was designed to determine the mechanisms by which benidipine exerts its antiapoptotic effect. 2. Adult male rats were subjected to 30 min of ischemia followed by 3 h of reperfusion. Rats were randomized to receive either vehicle or benidipine (10 microg x kg(-1), i.v.) 10 min before reperfusion. 3. Compared with rats receiving vehicle, those rats treated with benidipine had reduced postischemic myocardial apoptosis as evidenced by decreased TUNEL-positive staining (8.4+/-1.2 vs 15.3+/-1.3%, P<0.01) and caspase-3 activity (1.94+/-0.25 vs 3.43+/-0.29, P<0.01). 4. Benidipine treatment significantly reduced mitochondrial cytochrome c release and caspase-9 activation, but had no effect on caspase-8 activation, suggesting that benidipine exerts its antiapoptotic effect by inhibiting the mitochondrial-mediated, but not death receptor-mediated, apoptotic pathway. 4. 5. Benidipine treatment not only increased the maximal activity of ERK1/2 at 10 min after reperfusion, but also prolonged the duration of ERK1/2 activation. Benidipine treatment had no significant effect on other apoptotic regulating molecules, such as p38 MAPK. 6. Taken together, our present study demonstrated for the first time the differential regulation of a calcium channel blocker. Benidipine tilted the balance between ERK1/2 and p38 MAPK toward an antiapoptotic state, decreased mitochondrial cytochrome c release, reduced caspase-9 activation, and attenuated subsequent caspase-3 activation and postischemic myocardial apoptosis.     

A microdialysis study of effects of gastrodin on neurochemical changes in the ischemic/reperfused rat cerebral hippocampus

Gastrodin is a component extracted from the rhizome of Gastrodia elata, and has been shown to possess protective effects against neuron damage induced by simulated cerebral ischemia in previous studies. But its neurochemical effects on the ischemic brain had not been well studied. The present study aimed at evaluating the effects of gastrodin on the changes of transmitter amino acids in rat hippocampus during cerebral ischemia/reperfusion. Microdialysis sampling was performed during ischemia and early reperfusion periods in rats, and the glutamate and gamma-aminobutyric acid (GABA) in the dialysate were measured using high-performance liquid chromatography (HPLC). Administration of gastrodin (100 mg/kg) before ischemia significantly reduced the ischemia-induced elevation of glutamate levels during the postischemic period, increased the rise of extracellular GABA during the reperfusion periods, thus decreased the glutamate/GABA ratios during ischemia and reperfusion. These results provide insights to explain the neurochemical effects of gastrodin when applied prior to an ischemic event.     

Antiarrhythmic properties of benzyl-triamterene derivatives in the coronary artery ligated and reperfused rat

Triamterene (CAS 396-01-0) and a series of benzyl-triamterene derivatives were evaluated for their antiarrhythmic properties in the coronary artery ligated and reperfused (CAL-R) rat. The effects were compared with the antiarrhythmic activity of the potassium sparing diuretic amiloride and drugs out of the class-I (lidocaine) and class-III (amiodarone and sotalol). Triamterene and sotalol revealed at high doses antifibrillator activity, while the benzyl-triamterenes 2, 3, 5 and 6 could also depress ventricular extrasystoles (VES) and ventricular tachycardia (VT). At low doses the most benzyltriamterenes protected significantly against ventricular fibrillation (VF) and so they were equieffective or more effective than amiodarone or lidocaine. Amiloride showed in the CAL-R rat no antiarrhythmic activity, so that we conclude different mechanisms responsible for antikaliuretic and antiarrhythmic properties of amiloride and triamterenes. Taking into account the results of recently reported in vitro studies, where we could demonstrate antiarrhythmic activity combined with positive inotropic properties for triamterenes, the antiarrhythmic profile of these compounds may offer new possibilities for the treatment of ventricular arrhythmias.     

Cardioprotective effects of 2-octynyladenosine (YT-146) in ischemic/reperfused rat hearts

The present study was aimed at investigating the cardiac receptor subtypes involved in the cardioprotective effects of 2-octynyladenosine (YT-146), a novel adenosine receptor (AR) agonist. Isolated rat hearts were perfused in the Langendorff manner, and the hearts were exposed to 30 minute of ischemia followed by 60 minutes of reperfusion. YT-146 was infused for 10 minutes just before ischemia, and selective antagonists for AR subtypes were coadministered with YT-146. YT-146 (0.03–0.3 μM) dose dependently improved postischemic recovery of the left ventricular developed pressure (LVDP) of the ischemic/reperfused rat heart (maximum 59.7% ± 2.3% of the preischemic value). Coadministration of 8-(3-chlorostyryl) caffeine (A(2A) AR antagonist), alloxazine (A(2B)AR antagonist), or MRS-1191 (A(3) AR antagonist) with YT-146 failed to alter the cardioprotective effects of YT-146, and their LVDP recoveries were 55.9% ± 5.1%, 52.1% ± 1.9%, and 47.5% ± 1.7%, respectively, at the end of the reperfusion. On the other hand, coadministration of 8-cyclopentyl-1,3-dipropylxanthine (A(1) AR antagonist) abolished the YT-146–induced enhancement of postischemic LVDP recovery (31.7% ± 4.6%). The protein kinase C inhibitor chelerythrine also abolished the YT-146–induced enhancement of postischemic LVDP recovery (22.2% ± 4.5%). YT-146 has been known as an A(2) AR agonist, but our findings suggest that the cardioprotective effects of YT-146 are exerted via cardiac A(1) AR, not A(2) AR, stimulation and the activation of protein kinase C by preischemic treatment in isolated and crystalloid-perfused rat hearts.     

呋喃丙吡啶对再灌注心肌顿抑兔血小板聚集和血压的影响

目的：探讨心肌顿抑发生后血流动力学和血小板聚集率的变化，以及钙通道拮抗剂呋喃丙吡啶对心肌顿抑兔的保护作用。方法：经冠状动脉结扎法建立兔心肌顿抑模型。用药组缺血前用呋喃丙吡啶静脉给药。结果：对照组再灌注后１５ｍｉｎ血浆最大血小板聚集率达高峰（６１．７６％±９．２２％），用药组同一时间点血浆最大血小板聚集率较对照组低（４０．２３％±５．３９％，Ｐ＜０．０１）。对照组血压较缺血前明显下降（Ｐ＜０．０５），用药组：Ⅰ组（１ｍｇ·ｋｇ－１）血压变化与对照组无统计学差异，Ⅱ组（３ｍｇ·ｋｇ－１）缺血后舒张压下降明显与对照组相比差异显著，随着再灌注时间延长舒张压逐步回升。结论：心肌顿抑发生后伴有血流动力学变化及血小板聚集率增高，缺血前静脉应用呋喃丙吡啶治疗对心肌损伤具有保护作用。     

Effects of U74006F, a novel inhibitor of lipid peroxidation, in stunned reperfused canine myocardium.

U74006F, a novel 21-amino steroid is a potent inhibitor of iron-mediated lipid peroxidation and has been shown to be of therapeutic benefit in central nervous system ischemia. As oxygen radicals have been implicated in the development of postischemic myocardial dysfunction, we examined the efficacy of U74006F to enhance the recovery of function in a canine model of stunned, reperfused myocardium. Twenty-six dogs were randomized to either a vehicle (n = 11), U74006F (n = 10), or U74006F-paced group (n = 5). U74006F (6 mg/kg i.v.) was administered 15 min prior to coronary artery occlusion. Myocardial blood flows were measured by the microsphere technique, and function data were obtained by sonomicrometry. Both U74006F-treated groups demonstrated a significant increase in posterior wall thickening as compared to the vehicle treatment (U74006F-paced, 27.0 +/- 12.8%; U74006F, 22.4 +/- 11%; vehicle, -13.5 +/- 9.9%, p less than 0.001 following 3 h of reperfusion). Enhanced function recovery was accompanied by lower heart rates in the U74006F-treated group following reperfusion (treated versus vehicle, 109 +/- 6.7 versus 131 +/- 8.8 beats/min, p = 0.004). The U74006F-paced group was maintained at the same rate as the vehicle group, with no diminution in function recovery compared to the unpaced group. No effects in systemic hemodynamics or nutrient blood flow were evident as a function of drug treatment. We conclude that pretreatment with U74006F enhances the recovery of function in stunned canine myocardium via the inhibition of oxygen radicals and lipid peroxidation products. This activity suggests that this compound represents a new therapeutic adjunct in reperfusion and recanalization therapies.     

Xamoterol recruits an inotropic reserve in the acutely failing,reperfused canine myocardium without detrimental effects on its subsequent recovery

Summary The present study tested (1) whether xamoterol recruits an inotropic reserve in reperfused myocardium and (2) whether acute inotropic stimulation by xamoterol has deleterious consequences on the long-term recovery of the reperfused myocardium. Sixteen anaesthetized, open-chest dogs were bilaterally vagotomized and heart rate kept constant by left atrial pacing. The distal left circumflex coronary artery was occluded for 15 min and then reperfused for 8 h. The coronary occlusion resulted in regional myocardial dyskinesia and myocardial function remained severely depressed after release of the occlusion. At 10 min reperfusion, 8 dogs received xamoterol (100 g/kg i. v.), whereas the remaining 8 dogs served as controls and received saline. Xamoterol increased mean systolic wall thickening velocity (from 1.47 ± 2.34 (SD) mm/s at 10 min reperfusion to 7.13 ± 3.55 mm/s at 30 min reperfusion and 7.64 ± 2.48 mm/s at 1 h reperfusion, respectively, both P < 0.05). In the control group, only a slow recovery of mean systolic wall thickening velocity was observed (from 3.14 ± 3.30 mm/s to 2.96 ± 3.74 mm/s and 4.03 ± 3.00 mm/s at 10 min, 30 min, and 1 h reperfusion, respectively). At 8 h reperfusion, mean systolic wall thickening velocity was similar in both groups (7.97 ± 4.23 mm/s in the xamoterol-group and 6.87 ± 4.00 mm/s in the placebo-group). Histological examination revealed no difference in the extent of necrosis between the two groups after 8 h reperfusion. We conclude that (1) xamoterol recruits an inotropic reserve in reperfused myocardium, and (2) this recruitment of an inotropic reserve does not compromise functional recovery and structural integrity of the reperfused myocardium.Send offprint requests to G. Heusch at the above address