Glucose,insulin and potassium (GIK) during reperfusion mediates improved myocardial bioenergetics

Previous studies suggest glucose, insulin and potassium (GIK) infusion during ischemia reduces infarct size and improves post-ischemic myocardial function in acute myocardial infarction and following surgical revascularization of the heart. The potential use of GIK when given only during reperfusion after a period of global ischemia, as might occur during cardiac arrest, is unclear. To test the hypothesis that GIK reperfusion improves post-ischemic myocardial bioenergetics and function, we utilized a perfused heart model. Hearts from Sprague-Dawley rats (350-450 g) were perfused at 85 mmHg with oxygenated Krebs-Henseleit bicarbonate containing 5.5 mM glucose and 0.2 mM octanoic acid. Following 20 min of global ischemia, hearts were reperfused for 30 min with original solution (control) or GIK in two different doses (10 or 20 mM glucose each with insulin 10 U/l and K(+) 7 meq/l). Hearts perfused with GIK solutions had significantly higher ATP, creatine phosphate, energy charge, and NADP(+) and lower AMP and inosine levels compared with control after 30 min of reperfusion. Hearts reperfused with GIK had significantly higher developed pressure and higher dP/dt than control reperfused hearts. Reperfusion with GIK improved post-ischemic recovery of both contractile function and the myocardial bioenergetic state. GIK may be a viable adjunctive reperfusion therapy following the global ischemia of cardiac arrest to improve post-resuscitation cardiac dysfunction.     

A comparison between ranolazine and CVT-4325, a novel inhibitor of fatty acid oxidation, on cardiac metabolism and left ventricular function in rat isolated perfused heart during ischemia and reperfusion

Inhibition of fatty acid oxidation has been reported to be cardioprotective against myocardial ischemic injury; however, recent studies have questioned whether the cardioprotection associated with putative fatty acid oxidation inhibitors, such as ranolazine and trimetazidine, are due to changes in substrate oxidation. Therefore, the goals of this study were to compare the effects of ranolazine with a new fatty acid oxidation inhibitor, CVT-4325 [(R)-1-(2-methylbenzo[d]thiazol-5-yloxy)-3-(4-((5-(4-(trifluoromethyl)phenyl)-1,2,4-oxadiazol-3-yl)methyl)-piperazin-1-yl)propan-2-ol], on carbohydrate and fatty acid oxidation and on left ventricular (LV) function in the response to ischemia/reperfusion in rat isolated perfused hearts. Metabolic fluxes were determined in hearts perfused in an isovolumic Langendorff mode using 13C nuclear magnetic resonance isotopomer analysis or in isolated working hearts using [14C]glucose and [3H]palmitate, with and without 10 microM ranolazine or 3 microM CVT-4325. Isovolumic perfused hearts were also subjected to 30 min of low-flow ischemia (0.3 ml/min) and 60 min of reperfusion, and working hearts were subjected to 15 min of zero-flow ischemia and 60 min of reperfusion. Regardless of the experimental protocol, ranolazine had no effect on carbohydrate or fatty acid oxidation, whereas CVT-4325 significantly reduced fatty acid oxidation up to approximately 7-fold with a concomitant increase in carbohydrate oxidation. At these same concentrations, although ranolazine significantly improved LV functional recovery following ischemia/reperfusion, CVT-4325 had no significant protective effect. These results demonstrate that at pharmacologically relevant concentrations, ischemic protection by ranolazine was not mediated by inhibition of fatty acid oxidation and conversely that inhibition of fatty acid oxidation with CVT-4325 was not associated with improved LV functional recovery.     

Role of fatty acids in the recovery of cardiac function during resuscitation from hemorrhagic shock

This study tested the hypothesis that removal of fatty acids as a fuel source would improve cardiac efficiency at the expense of reduced cardiac contractile function in the isolated working heart after hemorrhage-retransfusion. Non-heparinized male Sprague-Dawley rats were anesthetized with ketamine-xylazine and were hemorrhaged to a mean arterial blood pressure of 40 mmHg for 1 h. Two-thirds volume of shed blood was reinfused together with 0.9% NaCl in a volume equal to 2.3 times the shed blood volume, followed by continuous infusion of 0.9% NaCl at 10 mL/kg per h for 3 h. Hearts were removed and perfused in closed, recirculating working mode for 60 min to measure hydraulic work and cardiac efficiency. Rates of glycolysis and glucose oxidation were assessed with [5-3H/U-14C] glucose (11 mM) in the absence or presence of 0.4 mM palmitate. Compared to baseline measurements, hemorrhage-retransfusion significantly reduced arterial blood glucose (228+/-7 versus 118+/-12 mg/dL) and non-esterified fatty acid concentrations (0.36+/-0.01 versus 0.30+/-0.02 mM), while elevating blood lactate (0.8+/-0.1 versus 2.5+/-0.4 mM). Perfusion of sham hearts with glucose-only did not alter cardiac work compared to shams perfused with glucose plus palmitate. However, shocked hearts perfused with glucose-only demonstrated a significant reduction in cardiac work compared to shocked hearts perfused with glucose plus palmitate and compared to sham hearts perfused with glucose only (P < 0.05, repeated measures ANOVA). Shocked hearts perfused with glucose plus palmitate showed no reduction in cardiac work compared to shams. Shocked hearts perfused with glucose-only had increased glucose oxidation rates compared to shams perfused with glucose plus palmitate. In sham hearts perfused with glucose-only, myocardial glycogen and triacylglycerol contents were significantly reduced compared to hearts freeze-clamped in situ. These endogenous fuels were not decreased in shocked hearts. These data indicate that hemorrhagic shock renders the heart unable to mobilize endogenous fuels, and suggest that withdrawal of fatty acid oxidation will impair myocardial energy metabolism during resuscitation.     

The effects of ryanodine on calcium uptake by the sarcoplasmic reticulum of ischemic and reperfused rat myocardium

Summary— The effects of ischemia and reperfusion on sarcoplasmic reticulum (SR) calcium uptake were measured in crude heart homogenates of rats and were compared to published results for rabbit hearts. Isolated rat hearts ( n = 5 in each group) were Langendorff-perfused at 37 °C and were either kept normally perfused (control group), or submitted to 15 min normothermic ischemia (ischemic group), or reperfused for 10 min after 15 min ischemia (reperfused group). Mechanical function recovered to 50–60% of control after 10 min reperfusion following ischemia. Ca uptake (control V_max; 23.0 ± 2.20 nmol·min⁻¹·mg of protein⁻¹) decreased during ischemia (V_max: 15.7 ± 1.60 nmol·min⁻¹·mg⁻¹) but recovered to control level on reperfusion (V_max: 20.8 ± 2.02 nmol·min⁻¹·mg⁻¹). An increased Ca uptake was obtained when the measurements were carried out in the presence of ryanodine (430 μM) to block Ca leakage through SR Ca-release channels. The relative magnitude of ryanodine effect in the ischemic myocardium (increase: 77.2 ± 18.20%) was more marked than in control (32.0 ± 8.22%) or reperfused myocardium (39.0 ± 10.66%). This result is different from that of rabbit myocardium where similar ryanodine effect is present in all groups (56.7 ± 13.76%, 50.0 ± 13.56% and 54.2 ± 6.88% in control, ischemic and reperfused hearts, respectively) and suggests that a component of cytosolic Ca overload via SR Ca-release channels is present during ischemia in rat, but not in rabbit myocardium.     

Lactate improves cardiac efficiency after hemorrhagic shock

This study was undertaken to examine the role of lactate on cardiac function and metabolism after severe acute hemorrhagic shock. Anesthetized, nonheparinized rats were bled to a mean arterial pressure of 25-30 mm Hg for 1 h; controls were not bled. Their hearts were removed, and cardiac work and efficiency (work/oxygen consumption) were measured in the isolated working heart mode for 60 min. The hearts were perfused with one of five substrate combinations: 1) glucose (11 mM), 2) glucose + 0.4 mM palmitate, 3) glucose + 0.4 mM palmitate + 8.0 mM lactate, 4) glucose + 1.2 mM palmitate, or 5) glucose + 1.2 mM palmitate + 8.0 mM lactate. After perfusion, hearts were freeze-clamped, and tissue contents of free coenzyme-A (CoA), acetyl CoA, and succinyl CoA were measured, as was myocardial pyruvate dehydrogenase (PDH) activity. The addition of 8.0 mM lactate significantly improved cardiac work in shocked hearts perfused with 0.4 mM palmitate and increased cardiac efficiency in the presence of either 0.4 mM or 1.2 mM palmitate. Compared to control hearts, shocked hearts exhibited a 20-30% decrease in PDH activity. Shocked hearts perfused with lactate demonstrated no increase in acetyl CoA content but did have a significant increase in tissue succinyl CoA compared to control hearts perfused with lactate or shocked hearts perfused without lactate. In the heart recovering from severe hemorrhagic shock, lactate improves cardiac efficiency in the presence of free fatty acids, possibly by a anaplerosis of the tricarboxylic acid cycle.     

Reversible blockade of electron transport during ischemia protects mitochondria and decreases myocardial injury following reperfusion

Cardiac mitochondria sustain damage during ischemia and reperfusion, contributing to cell death. The reversible blockade of electron transport during ischemia with amobarbital, an inhibitor at the rotenone site of complex I, protects mitochondria against ischemic damage. Amobarbital treatment immediately before ischemia was used to test the hypothesis that damage to mitochondrial respiration occurs mainly during ischemia and that protection of mitochondria during ischemia leads to decreased cardiac injury with reperfusion. Langendorff-perfused Fischer-344 rat hearts were treated with amobarbital (2.5 mM) or vehicle for 1 min immediately before 25 min of global ischemia. Both groups were reperfused for 30 min without additional treatment. Subsarcolemmal (SSM) and interfibrillar (IFM) populations of mitochondria were isolated after reperfusion. Ischemia and reperfusion decreased state 3 and increased state 4 respiration rate in both SSM and IFM. Amobarbital treatment protected oxidative phosphorylation measured following reperfusion and improved the coupling of respiration. Cytochrome c content measured in SSM and IFM following reperfusion decreased in untreated, but not in amobarbital-treated, hearts. H(2)O(2) release from SSM and IFM isolated from amobarbital-treated hearts during reperfusion was markedly decreased. Amobarbital treatment before ischemia improved recovery of contractile function (percentage of preischemic developed pressure: untreated 51 +/- 4%, n = 12; amobarbital 70 +/- 4%, n = 11, p < 0.01) and substantially reduced infarct size (untreated 32 +/- 2%, n = 7; amobarbital 13 +/- 2%, n = 7, p < 0.01). Thus, mitochondrial damage occurs mainly during ischemia rather than during reperfusion. Reperfusion in the setting of preserved mitochondrial respiratory function attenuates the mitochondrial release of reactive oxygen species, enhances contractile recovery, and decreases myocardial infarct size.     

AMP-activated protein kinase mediates ischemic glucose uptake and prevents postischemic cardiac dysfunction, apoptosis, and injury

AMP-activated protein kinase (AMPK) is an important regulator of diverse cellular pathways in the setting of energetic stress. Whether AMPK plays a critical role in the metabolic and functional responses to myocardial ischemia and reperfusion remains uncertain. We examined the cardiac consequences of long-term inhibition of AMPK activity in transgenic mice expressing a kinase dead (KD) form of the enzyme. The KD mice had normal fractional shortening and no heart failure, cardiac hypertrophy, or fibrosis, although the in vivo left ventricular (LV) dP/dt was lower than that in WT hearts. During low-flow ischemia and postischemic reperfusion in vitro, KD hearts failed to augment glucose uptake and glycolysis, although glucose transporter content and insulin-stimulated glucose uptake were normal. KD hearts also failed to increase fatty acid oxidation during reperfusion. Furthermore, KD hearts demonstrated significantly impaired recovery of LV contractile function during postischemic reperfusion that was associated with a lower ATP content and increased injury compared with WT hearts. Caspase-3 activity and TUNEL-staining were increased in KD hearts after ischemia and reperfusion. Thus, AMPK is responsible for activation of glucose uptake and glycolysis during low-flow ischemia and plays an important protective role in limiting damage and apoptotic activity associated with ischemia and reperfusion in the heart.     

Reperfusion injury induces apoptosis in rabbit cardiomyocytes.

The most effective way to limit myocardial ischemic necrosis is reperfusion, but reperfusion itself may result in tissue injury, which has been difficult to separate from ischemic injury. This report identifies elements of apoptosis (programmed cell death) in myocytes as a response to reperfusion but not ischemia. The hallmark of apoptosis, nucleosomal ladders of DNA fragments (approximately 200 base pairs), was detected in ischemic/reperfused rabbit myocardial tissue but not in normal or ischemic-only rabbit hearts. Granulocytopenia did not prevent nucleosomal DNA cleavage. In situ nick end labeling demonstrated DNA fragmentation predominantly in myocytes. The pattern of nuclear chromatin condensation was distinctly different in reperfused than in persistently ischemic tissue by transmission electron microscopy. Apoptosis may be a specific feature of reperfusion injury in cardiac myocytes, leading to late cell death.     

Levosimendan: effects of a calcium sensitizer on function and arrhythmias and cyclic nucleotide levels during ischemia/reperfusion in the Langendorff-perfused guinea pig heart.

The majority of clinically used inotropes act by increasing cytosolic calcium levels, which may hypothetically worsen reperfusion stunning and provoke arrhythmias. We tested the hypothesis that the calcium sensitizer levosimendan (levo) given during ischemia alone or ischemia and reperfusion would improve reperfusion function without promoting arrhythmias. The Langendorff-perfused guinea pig heart, subjected to 40-min low-flow ischemia (0.4 ml/min) with or without levo (10-300 nM) given during ischemia or ischemia/reperfusion was used. Left ventricular developed pressure (LVDP) was used as an index of mechanical function. The effect of levo (300 nM) or dobutamine (0.1 microM) on the incidence of ischemia/reperfusion arrhythmias was also investigated. Control hearts (vehicle-perfused) had LVDPs of 69.4 +/- 2.1 mm Hg whereas hearts treated with levo during ischemia and reperfusion (300 nM) had LVDPs of 104.5 +/- 2.7 mm Hg (p <.05). Hearts treated with levo during ischemia alone (10 nM) had reperfusion LVDPs of 95.8 +/- 4.2 mm Hg (p <.05) after 30-min reperfusion. Hearts treated with both levo and 10 microM glibenclamide (K(ATP) channel blocker) during ischemia had reperfusion LVDPs of 73.4 +/- 4.3 mm Hg after 30-min reperfusion. Of control hearts, 25% developed reperfusion ventricular tachycardia but not ventricular fibrillation. Levo-treated hearts had no ischemia/reperfusion arrhythmias whereas 83% (p <.05 versus control) of dobutamine-treated hearts developed ventricular tachycardia and 33% (p <.05 versus levo) developed reperfusion ventricular fibrillation. Levo improved reperfusion function without promoting arrhythmias in this model. This was possibly achieved by opening the K(ATP) channels during ischemia and sensitizing myocardial contractile apparatus instead of elevating cytosolic calcium levels in reperfused hearts.     

Blockade of electron transport before cardiac ischemia with the reversible inhibitor amobarbital protects rat heart mitochondria

Cardiac ischemia damages the mitochondrial electron transport chain. Irreversible blockade of electron transport at complex I by rotenone decreases ischemic damage to cardiac mitochondria by decreasing the loss of cytochrome c and preserving respiration through cytochrome oxidase. Therapeutic intervention to protect myocardium during ischemia and reperfusion requires the use of a reversible inhibitor that allows resumption of oxidative metabolism during reperfusion. Amobarbital is a reversible inhibitor at the rotenone site of complex I. We asked whether amobarbital administered immediately before ischemia protected respiratory function. Isolated rat hearts were perfused for 15 min followed by 25-min global ischemia at 37 degrees C. Amobarbital-treated hearts received drug for 1 min before ischemia. Subsarcolemmal (SSM) and interfibrillar (IFM) populations of mitochondria were isolated after ischemia, and oxidative phosphorylation was measured. Amobarbital protected oxidative phosphorylation, including through cytochrome oxidase, in both SSM and IFM in a dose-dependent manner, with an optimal dose of 2 to 2.5 mM. Amobarbital also preserved cytochrome c content in both SSM and IFM. Thus, reversible blockade of the electron transport chain during ischemia protects mitochondrial respiration.     

Effect of nifedipine in hypothermic cardioplegia: a phosphorus-31 nuclear magnetic resonance study

The ability of nifedipine to enhance myocardial protection was assessed on isolated perfused rat hearts subjected to 180 min of hypothermic (20 degrees C), global ischemia, followed by 45 min of normothermic reperfusion. Intracellular pH, ATP, Pi and phosphocreatine content were serially measured at 4 min intervals by phosphorus-31 nuclear magnetic resonance spectroscopy and correlated with simultaneously recorded hemodynamic parameters. Addition of nifedipine (0.075 mumol/l and 0.5 mumol/l) to Saint Thomas' cardioplegic solution reduced Pi accumulation during ischemic arrest and increased phosphocreatine levels during reperfusion. Post-ischemic functional recovery was not improved at a drug concentration of 0.075 mumol/l and was depressed at 0.5 mumol/l. These results clearly show that the presence of nifedipine in Saint Thomas' cardioplegic solution does not provide significant additional myocardial protection under hypothermic conditions.     

The effect of intracoronary diltiazem on regional myocardial function and development of infarcts in porcine hearts

The effect of intracoronary (i.c.) pretreatment with diltiazem on regional myocardial function and the development of infarcts was investigated in regionally ischemic, reperfused porcine hearts. The left anterior descending coronary artery (LAD) was distally ligated in 16 pigs for 20-90 min followed by 24 h of reperfusion. Eight pigs were treated with increasing doses of i.c. diltiazem (0.375 mg/min, 0.75 mg/min, 1 mg/min) prior to ischemia. Eight pigs served as controls. Regional myocardial function was assessed by implanted ultrasonic crystals. Infarct size was determined as ratio of infarcted (tetrazolium stain) to ischemic myocardium (dye technique). I.c. diltiazem mainly depressed early systolic shortening (isovolumetric contraction) and lengthening during the first half of diastole. Pretreatment with this calcium antagonist significantly delayed the development of infarcts. In control experiments, a mean infarct size of 74% was found after 45-min ischemia. At that time no infarction was observed in the treated group, where infarcts started to evolve after 60-min ischemia. It is concluded that the favorable action of i.c. diltiazem can mainly be ascribed to a reduced myocardial oxygen consumption at the onset of ischemia.     

Oxygen radicals generated at reflow induce peroxidation of membrane lipids in reperfused hearts.

To test whether generation of oxygen radicals during postischemic reperfusion might promote peroxidation of cardiac membrane lipids, four groups of Langendorff-perfused rabbit hearts were processed at the end of (a) control perfusion, (b) 30 min of total global ischemia at 37 degrees C without reperfusion, (c) 30 min of ischemia followed by reperfusion with standard perfusate, (d) 30 min of ischemia followed by reperfusion with the oxygen radical scavenger human recombinant superoxide dismutase (h-SOD). The left ventricle was homogenized and tissue content of malonyldialdehyde (MDA), an end product of lipid peroxidation, was measured on the whole homogenate as well as on various subcellular fractions. Reperfusion was accompanied by a significant increase in MDA content of the whole homogenate and of the fraction enriched in mitochondria and lysosomes. This phenomenon was not observed in hearts subjected to ischemia but not reperfused, and was similarly absent in those hearts which received h-SOD at reflow. Reperfused hearts also had significantly greater levels of conjugated dienes (another marker of lipid peroxidation) in the mitochondrial-lysosomal fraction. Again, this phenomenon did not occur in ischemic hearts or in reperfused hearts treated with h-SOD. Unlike the effect on tissue MDA and conjugated dienes, reperfusion did not significantly stimulate release of MDA in the cardiac effluent. Treatment with h-SOD was also associated with significant improvement in the recovery of cardiac function. In conclusion, these data directly demonstrate that postischemic reperfusion results in enhanced lipid peroxidation of cardiac membranes, which can be blocked by h-SOD, and therefore is most likely secondary to oxygen radical generation at reflow.     

蛋白激酶C在缺血预处理限制心肌梗死范围效应中作用的观察

目的：探讨蛋白激酶Ｃ（ＰＫＣ）在缺血预处理限制心肌梗死范围中的作用。方法：采用非循环式Ｌａｎｇｅｎｄｏｒｆ离体兔心恒压灌流缺血再灌注模型,观察特异性ＰＫＣ对缺血预处理心肌保护作用的影响。结果：单纯缺血３０分钟及再灌注６０分钟可引起３３．９６％±１２．０７％的心肌坏死;在缺血前如给予５分钟全心缺血及１０分钟再灌注的预处理可缩小心肌梗死范围至１５．２６％±５．１４％,２组比较Ｐ＜０．０５;Ｃｈｅｌｅｒｙｔｈｒｉｎｅ（ＰＫＣ阻滞剂）预灌流心脏不能影响心肌梗死范围（３６．５１％±６．９１％）,但如在预处理的再灌注期用Ｃｈｅｌｅｒｙｔｈｒｉｎｅ灌流心脏,则可取消缺血预处理的心肌保护作用,其心肌梗死范围为３２．１９％±８．３１％,与对照组比较差异无统计学意义。结论：蛋白激酶Ｃ介导了缺血预处理限制心肌梗死范围的作用。     

Effect of amiloride on age-dependent cardiac dysfunction after ischemia/reperfusion in the isolated, perfused rat heart

This study was intended to compare the cardiac consequences of ischemia/reperfusion and amiloride treatment in immature (2-3 wk), juvenile (4-6 wk), and adult (3-5 mo) rats using an isolated, perfused heart model. Male immature, juvenile, and adult rats were anticoagulated and anesthetized. Hearts were harvested and coronary arteries were perfused on a Langendorff apparatus via retrograde perfusion of the aorta at a constant coronary flow (initially determined by perfusing the heart at 50 mm Hg perfusion pressure) with oxygenated Krebs-Henseleit-Bicarbonate (KHB) solution. Left ventricular peak systolic (LVPSP) and end diastolic (LVEDP) pressures were measured via a balloon-tipped catheter placed in the left ventricle through the mitral valve. Following a 20-30 min stabilization period, hearts underwent 30 min of normothermic ischemia and were then reperfused with Krebs-Henseleit-Bicarbonate alone for 30 min, or Krebs-Henseleit-Bicarbonate containing 500 microM amiloride for 5 min followed by Krebs-Henseleit-Bicarbonate alone for 25 min (n = 6/age group). Left ventricular generated pressure was calculated (left ventricular peak systolic-left ventricular end diastolic) and used as a measure of ventricular function. All hearts demonstrated a decrease in generated pressure, respectively, from preischemic levels at 15 and 30 min of reperfusion, although this decrease was significantly less for the immature hearts. Ischemia/reperfusion injury was attenuated by amiloride in adult and juvenile hearts, whereas ischemia/reperfusion injury was worsened by amiloride in immature hearts. Although immature hearts were relatively resistant to ischemia/reperfusion injury compared with adult and juvenile hearts, the presence of amiloride during reperfusion resulted in more severe ventricular dysfunction in immature hearts. These data suggest a differential age-dependent mechanism of sarcolemmal ion exchange in response to ischemia/reperfusion.     

Concentration-dependent effects of prostacyclin on the response of the isolated guinea pig heart to ischemia and reperfusion: possible involvement of the slow inward current

Isolated guinea pig hearts were subjected to 40 min of low flow global ischemia followed by 30 min of reperfusion. The effects of prostacyclin (PGI2) (10 pg/ml-10 ng/ml) on the response of hearts to ischemia and reperfusion were studied. Ischemia caused a 55% decline in contractile force and the development of contracture. Sinus bradycardia and varying degrees of atrioventricular conduction block were observed. Reperfusion was associated with rapid recovery of contractile force and a gradual decline in resting tension. PGI2 (1 ng/ml) enhanced ischemic contracture significantly at 10 and 20 min (P less than .05). Hearts made ischemic in the presence of PGI2 developed higher degrees of atrioventricular conduction block when compared to controls. Reperfusion in the presence of 1 or 10 ng/ml of PGI2 caused a significant decline in recovery of force (P less than .05) and enhanced reperfusion-associated contracture. We examined the influence of verapamil (100 ng/ml) and lowering external calcium to 1.25 mM on hearts subjected to ischemia and reperfusion in the absence of presence of PGI2 (1 ng/ml). Neither verapamil nor 1.25 mM calcium exerted significant effects on the decline of contractile force during ischemia or on recovery of contractility upon reperfusion. However, verapamil did reverse the reperfusion-associated cardiodepressant effects of PGI2. Verapamil abolished contracture in control hearts after 5 and 10 min of reperfusion and prevented the enhancement of contracture in hearts reperfused in the presence of PGI2.(ABSTRACT TRUNCATED AT 250 WORDS)     

The rapid and reversible activation of a calcium-independent plasmalogen-selective phospholipase A2 during myocardial ischemia.

Recent studies have demonstrated the existence of two members of a novel family of calcium-independent plasmalogen-selective phospholipases A2 in mammalian myocardium (Wolf, R. A., and R. W. Gross. 1985. J. Biol. Chem. 260:7295-7303; and Hazen, S. L., D. A. Ford, and R. W. Gross. 1991. J. Biol. Chem. 266:5629-5633). To examine the potential role of these calcium-independent phospholipases A2 in mediating membrane dysfunction during early myocardial ischemia, the temporal course of alterations in phospholipase A2 activity during global ischemia in Langendorf perfused rabbit hearts was quantified and compared with traditionally accepted markers of myocytic ischemic injury and anaerobic metabolism. We now report that membrane-associated calcium-independent plasmalogen-selective phospholipase A2 activity increased over 400% during 2 min of global ischemia (P less than 0.01), was near maximally activated (greater than 10-fold) after only 5 min of ischemia, and remained activated throughout the entire ischemic interval examined (2-60 min). Activation of membrane-associated plasmalogen-selective phospholipase A2 after 5 min of myocardial ischemia was rapidly reversible during reperfusion of ischemic tissue. Both the activation of phospholipase A2 and its reversibility during reperfusion were temporally correlated to alterations in myocytic anaerobic metabolism. Furthermore, activation of membrane-associated phospholipase A2 was essentially complete before electron microscopic evidence of cellular damage. Collectively, these results identify dynamic alterations in calcium-independent plasmalogen-selective phospholipase A2 activity during myocardial ischemia which precede irreversible cellular injury and demonstrate that activation of plasmalogen-selective phospholipase A2 is amongst the earliest biochemical alterations in ischemic myocardium.     

Inhibition of peroxynitrite precursors, NO and O2, at the onset of reperfusion improves myocardial recovery

AIM OF STUDY: Previous reports note an increase in both reactive oxygen species (ROS) and nitric oxide (*NO) at the onset of myocardial reperfusion. We tested the hypothesis that inhibition of *NO or ROS production at the time of reperfusion improves recovery of post-ischemic myocardial function. METHODS AND MATERIALS: Isolated rat hearts were perfused with temperature controlled (37.4 degrees C) modified Krebs Henseleit buffer solution at 85 mm Hg. Following 20 min of global ischemia, hearts were reperfused for the first 10 min with: (1) standard buffer (control), (2) buffer with a NOS inhibitor, N-nitro-L-arginine methyl ester (L-NAME), (3) buffer with superoxide dismutase (SOD) or (4) buffer with N-morpholinosydnonimine hydrochloride (SIN-1), a peroxynitrite generator. Tissue O(2) and *NO were continuously measured with thin electrochemical probes embedded in the wall of the LV. ROS was measured with the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) (40 mM). LV contractile function was continuously monitored. RESULTS: Recovery of LV contractile function was significantly improved in hearts initially reperfused with L-NAME and SOD and significantly depressed in hearts reperfused with SIN-1 compared with control (p<0.01, n=5-8 per group). DMPO-adduct during reperfusion (measure of ROS) was significantly decreased with SOD (p<0.001 versus L-NAME and Control, n=4 per group) and unchanged with L-NAME and SIN-1 compared with Control. With L-NAME, tissue *NO and PO(2) were significantly decreased, independent of coronary flow, during reperfusion compared with control and SIN-1. CONCLUSIONS: Inhibition of O(2)*(-) or *NO at the time of reperfusion improves early reperfusion LV function and alters tissue oxygen tension. In contrast to pre-ischemic treatments, intervention to reduce peroxynitrite generation at the onset of reperfusion can effectively improve post-ischemic myocardial recovery.     

Temporal differences in actions of calcium channel blockers on K+ accumulation, cardiac function, and high-energy phosphate levels in ischemic guinea pig hearts

We investigated temporal differences in the protective action of three types of Ca2+ channel blockers in myocardial ischemia, focusing particularly on the blocking ability under depolarizing conditions. The effects of diltiazem, verapamil, and nifedipine on extracellular potassium concentration ([K+]e), acidosis, and level of metabolic markers were examined during 30-min global ischemia and postischemic left ventricular (LV) function in isolated guinea pig hearts. Diltiazem and verapamil, but not nifedipine, inhibited the late phase (15-30 min) of [K+]e elevation, whereas all three blockers delayed the onset of the early phase (0-8 min) of [K+]e elevation. Diltiazem and verapamil inhibited ischemic contracture and improved postischemic LV function to a greater extent. These differences appeared to be linked to preservation of ATP and creatine phosphate and delay of cessation of anaerobic glycolytic activity. Maneuvers to preserve energy sources during ischemia (decrease in external Ca2+ concentration or pacing at a lower frequency) attenuated the late phase of [K+]e elevation. Inhibition of LV pressure was potentiated 12- and 8.2-fold by diltiazem and verapamil, respectively, at 8.9 mM K+ as compared with 2.9 mM K+, whereas that by nifedipine was unchanged. These results indicate that the differential cardioprotection of Ca2+ channel blockers in the late period of ischemia correlates with preservation of high-energy phosphates as a result of different Ca2+ channel blocking abilities under high [K+]e conditions.     

无创伤双后肢缺血后处理对移植胰腺缺血再灌注损伤的远隔保护

背景：缺血预处理及缺血后处理是近年来提出减轻缺血再灌注损伤有效方法。目的：探讨无创伤双后肢缺血后处理对移植胰腺缺血再灌注损伤的影响及机制。方法：18只糖尿病SD大鼠数字表法随机分为3组,对照组仅行开腹术;缺血再灌注组仅行胰腺移植;缺血后处理组,移植前行非创伤性双后肢缺血后处理。结果与结论：缺血再灌注组血糖和胰腺组织中丙二醛水平均高于缺血后处理组（P〈0.01）、而超氧化物歧化酶活性低于缺血后处理组（P〈0.01）;与缺血后处理组比较,缺血再灌注组胰腺组织凋亡指数明显增高（P〈0.01）。结果提示,无创伤双后肢缺血后处理对大鼠移植胰的缺血再灌注损伤具有保护作用,机制可能与可通过减少超氧化物歧化酶失活,从而清除氧自由基以及减少胰腺细胞凋亡等有关。