Postischemic Contractile Dysfunction Research Articles

GLP-1(7-36) and GLP-1(9-36) Protect Murine Hearts from Post-Ischemic Contractile Dysfunction

Background: Increasing evidence suggests that glycemic control is necessary but insufficient to alter cardiovascular complications in Type 2 DM. Glucagon like peptide-1(GLP-1) is a novel anti-glycemic agent that possesses both insulinotropic and insulinomimetic actions and has been shown to be cardio-protective in both animals and humans. However, it is uncertain as to whether the protective effects are mediated by the native peptide, GLP-1(7-36) or its active metabolite, GLP-1(9-36). The goal of the present study was to determine the effects of GLP-1 (7-36) versus GLP-1(9-36) on post-ischemic contractile dysfunction.

Overexpression of heat shock protein 27 protects against ischaemia/reperfusion-induced cardiac dysfunction via stabilization of troponin I and T

Heat shock protein 27 (Hsp27) renders cardioprotection from ischaemia/reperfusion (I/R) injury, but little is known about its role in myofilaments. We proposed that increased expression of Hsp27 may improve post-ischaemic contractile dysfunction by preventing I/R-induced cardiac troponin I (cTnI) and troponin T (cTnT) degradation. Adenovirus-mediated Hsp27 overexpression improved contractile function in perfused rat hearts subjected to global no-flow I/R (30-min/30-min). Such improvement was further confirmed in Hsp27-overexpressing cardiomyocytes subjected to simulated I/R (20-min/30-min). Moreover, these cells showed restored myofilament response to Ca(2+) but not intracellular Ca(2+) transients. The protection correlated with attenuation of I/R-induced cTnI and cTnT degradation. Confocal microscopy revealed co-localization of Hsp27 with these proteins. Co-immunoprecipitation and pull-down assays further confirmed that Hsp27 interacted with the COOH-terminus of cTnI and the NH(2)-terminus of cTnT and that Hsp27 overexpression decreased the interaction between mu-calpain (a protease mediating proteolysis of cTnI and cTnT) and cTnI or cTnT under I/R. The findings reveal a novel role of Hsp27 in the protection of cTnI and cTnT from I/R-induced degradation by preventing their proteolytic cleavage via interacting with these proteins. Such protection may result in restored post-ischaemic myofilament response to Ca(2+) and improved post-ischaemic contractile function.

Effect of acute and chronic simvastatin treatment on post-ischemic contractile dysfunction in isolated rat heart

Statins are powerful lipid-lowering drugs, widely used in patients with hyperlipidemia and coronary artery disease. It was found, however, that statins appear to have a pleiotropic effect beyond their lipid-lowering ability. They exert anti-inflammatory, antithrombotic and antioxidant effects, increase nitric oxide production and improve endothelial dysfunction. The aim of our study was to examine the effect of chronic and acute treatment with simvastatin on the contractile function of the isolated perfused rat heart after ischemia/reperfusion injury. Contractile function was measured on isolated rat hearts, perfused according to Langendorff under constant pressure. The hearts were subjected to 20 min of global ischemia, followed by 40 min of reperfusion. To investigate the acute effect, simvastatin at a concentration of 10 micromol/l was added to the perfusion solution during reperfusion. In chronic experiments the rats were fed simvastatin at a concentration of 10 mg/kg for two weeks before the measurement of the contractile function. Acute simvastatin administration significantly increased reparation of the peak of pressure development [(+dP/dt)(max)] (52.9+/-8.2 %) after global ischemia, as compared with the control group (28.8+/-5.2 %). Similar differences were also observed in the time course of the recovery of [(+dP/dt)(max)]. Chronic simvastatin was without any protective effect. Our results reveal that the acute administration of simvastatin during reperfusion, unlike the chronic treatment, significantly reduced contractile dysfunction induced by ischemia/reperfusion injury. This supports the idea of possible cardioprotective effect of statin administration in the first-line therapy of the acute coronary syndrome.

Lack of the effect of superoxide dismutase and catalase on Na+,K+-ATPase activity in stunned rabbit hearts

Reactive oxygen species (ROS) have been implicated in the mechanism of postischemic contractile dysfunction, known as myocardial stunning. In this study, we examined protective effects of antioxidant enzymes, superoxide dismutase (SOD) and catalase, against ischemia/reperfusion-induced cardiac dysfunction and inhibition of Na+,K+-ATPase activity. Isolated Langendorff-perfused rabbit hearts were subjected to 15 min of global normothermic ischemia followed by 10 min reperfusion. The hearts treated with SOD plus catalase did not show significant recovery of left ventricular (LV) end-diastolic pressure compared with untreated ischemic reperfused hearts. Treatment with antioxidants had no protective effects on developed LV pressure or its maximal positive and negative first derivatives (+/-LVdP/dt). Myocardial stunning was accompanied by significant loss in sarcolemmal Na+,K+-ATPase activity and thiol group content. Inhibition of enzyme activity and oxidation of SH groups were not prevented by antioxidant enzymes. These results suggest that administration of SOD and catalase in perfusate do not protect significantly against cardiac dysfunction in stunned rabbit myocardium.

Differential effects of the sodium calcium exchange inhibitor, KB-R7943, on ischemia and reperfusion injury in isolated guinea pig ventricular myocytes

Effects of the Na +–Ca 2+ exchange (NCX) inhibitor KB-R7943 on electrical and contractile function were examined in guinea pig ventricular myocytes exposed to ischemia and reperfusion. Action potentials and transmembrane currents were recorded with microelectrodes; contractions were measured with an edge detector. Cells were exposed to simulated ischemia (hypoxia, hypercapnia, hyperkalemia, acidosis, lactate accumulation, no glucose) for 20 min and reperfused with Tyrode's solution. Experiments were conducted at 37 °C in the absence or presence of KB-R7943. Low concentrations of KB-R7943 (0.1 μM) had little impact on changes in contractions, membrane potential, or Ca 2+ current induced by ischemia and reperfusion. However, higher concentrations of KB-R7943 (0.5 and 1.0 μM) reduced the magnitude of Ca 2+ current and promoted action potential abbreviation in both ischemia and reperfusion. High concentrations of KB-R7943 also promoted post-ischemic contractile dysfunction (stunning) in reperfusion. In the absence of KB-R7943, the arrhythmogenic transient inward current ( I TI) plus aftercontractions occurred upon reperfusion, and some cells exhibited irreversible cell injury (hypercontracture). Higher concentrations of KB-R7943 (0.5 and 1.0 μM) did not affect the occurrence or magnitude of I TI and aftercontractions and did not affect the occurrence of hypercontracture. In contrast, 0.1 μM KB-R7943 virtually abolished I TI, aftercontractions and hypercontracture. Thus, low concentrations of KB-R7943 protected against ischemia and reperfusion injury, but higher concentrations of drug actually exacerbated detrimental effects of ischemia and reperfusion. These results suggest that inhibition of I TI may contribute to the antiarrhythmic effects of KB-R7943 on reperfusion-induced arrhythmias.

Abstract 552: Mitochondrial KCa and Anion Channels Influence Mitochondrial Volume and Respiratory Rate: Relevance to Cardioprotection

Ion channels in the mitochondrial inner membrane influence the response of the heart to ischemia and reperfusion - the opening of mitochondrial K + channels (mitoK Ca or mitoK ATP ) induce cardioprotection while the excessive activation of an inner membrane anion channel (IMAC) contributes to post-ischemic contractile and electrical dysfunction. Here we examined the effects of K Ca channel openers and inhibitors on mitochondrial volume and respiration in KCl-based medium in energized isolated guinea-pig heart mitochondria. Inhibition of IMAC with a mitochondrial benzodiazepine ligand was also assessed. Mitochondrial swelling and contraction were evoked by the sequential addition of 5mM glutamate (Glu) and 5mM malate (Mal), respectively. Both the swelling and contraction responses involved energy-dependent fluxes of K + and Cl − . The Glu-induced swelling response was suppressed by preincubation with either the K Ca channel toxin paxilline, or by the IMAC blocker 4′-chlorodiazepam (4-Cl-DZP). The K Ca opener NS1619, at concentrations <10μM, initiated mitochondrial swelling responses either in the absence or presence of substrates. This effect was blocked by paxilline or charybdotoxin. Massive and irreversible swelling was induced by [NS1619] >10μM. Under similar conditions, mitochondrial respiratory control (State3/State4 VO 2 ratio) was maintained at >15 for [NS1619] <10μM, but was strongly suppressed above 20μM due to an increase in state4- and a decrease in state3-respiration. 4-Cl-DZP did not affect respiration and only slightly increased the K 0.5 for ADP. The findings indicate that mitochondrial matrix volume is modulated by mitoK Ca channel activity under normal conditions and in the presence of K Ca channel openers, while IMAC channels underlie compensatory anion movements in response to K + flux. Physiological regulation of mitochondrial volume by ion transport is in turn linked to altered respiratory flux, with important implications for cardioprotection and recovery of mitochondrial function in the post-ischemic heart.

Optimal Time Duration for Low-Pressure Controlled Reperfusion to Efficiently Protect Ischemic Rat Heart

Previous studies have shown the capacity of low-pressure (LP) reperfusion to protect the ischemic heart. The present study sought to determine the optimal time for the application of LP reperfusion. Isolated rat hearts (n = 30) were exposed to 40 minutes of global warm ischemia followed by 70 minutes of reperfusion. Reperfusion was performed under LP (LP = 70 cm H 2O) for 0 (control group), 5 (group LP-5), 10 (group LP-10), 30 (group LP-30), or 60 (group LP-60) minutes. Following the LP period the hearts were reperfused with normal pressure (100 cm H 2O) until the end of reperfusion. Cardiac function was assessed during reperfusion using the Langendorff model. Myocardial necrosis was assessed by measuring LDH leakage in the coronary effluents. Functional recovery was reduced among the control and LP-5 groups with rate-pressure products (RPP) averaging 3788 ± 499 and 5333 ± 892 mm Hg/min, respectively. RPP was significantly improved in other groups with RPP averaging 7363 ± 1159, 7441 ± 863, and 7269 ± 692 mm Hg/min in LP-10, LP-30, and LP-60 ( P < .01). Similarly, necrosis measured by LDH leakage was significantly reduced in LP-10, LP-30, and LP-60 hearts ( P < .01). This study demonstrated that LP reperfusion improves postischemic contractile dysfunction and attenuates necrosis when applied for at least 10 minutes.

Remote Preconditioning- Endocrine Factors in Organ Protection Against Ischemic Injury

Cardiovascular disease is the leading cause of death in the United States and developing world. Experimental and clinical studies have demonstrated that a number of interventions including brief periods of ischemia or hypoxia and certain endogenous factors such as opioids, bradykinin, growth factors or pharmacological agents are capable of protecting the heart against post-ischemic contractile dysfunction, arrhythmias and myocardial infarction. This conventional cardioprotection occurs via an autocrine or paracrine action in which these protective factors are released from the heart to act upon itself. Over the last ten years, a growing body of evidence indicates that a brief ischemic insult on one organ releases endogenous factors that protect other organs against a prolonged ischemic insult. This phenomenon, termed remote preconditioning or preconditioning at a distance, implicates an endocrine action, and may involve humoral or neural-endocrine signaling. This review will summarize the endocrine factors identified and implicated in this inter-organ cytoprotection.

Role of S-nitrosothiol transport in the cardioprotective effects of S-nitrosocysteine in rat hearts

The objective of this study was to determine if prior exposure of rat hearts to S-nitrosocysteine (CysNO) was able to provide protection against reperfusion injury. We probed NO release using the extracellular NO scavenger oxyhemoglobin (oxyHb), and we examined the involvement of the amino acid transport system L (L-AT), a known transporter of CysNO, using the L-AT competitor, l-leucine (L-Leu). Isolated (9- to 12-week-old Wistar male) rat hearts (six to eight per group) were perfused with CysNO (10 μM) for 30 min with or without the L-AT competitor L-Leu (1 mM) before 30 min of ischemia. Cardiac function was assessed before, during, and after treatment and during 120 min of reperfusion after ischemia. Functional recovery (rate–pressure product) was significantly improved in the CysNO group compared to hearts in the CysNO + L-Leu group and the control group ( p < 0.05). Necrosis, measured by triphenyltetrazolium chloride staining, was significantly reduced in CysNO hearts ( p < 0.05) and this improvement was reversed by L-Leu. The NO scavenger oxyHb (20 μM) was perfused either concomitant with CysNO or just before ischemia. In neither case did oxyHb affect the cardioprotection afforded by CysNO. OxyHb alone, given in either time window, did not alter the course of ischemia–reperfusion injury. When nitrite was used in place of CysNO, no protective effects were observed. Perfusion with CysNO increased tissue S-nitrosothiol (RSNO) levels from an unmeasurable background to a value of about 15.7 ± 4.1 pmol RSNO/mg protein, as measured by triiodide-based chemiluminescence in the presence and absence of mercury(II) chloride. In the presence of l-Leu, this value dropped to 0.4 ± 0.3 pmol RSNO/mg protein. This study demonstrates that exposure to CysNO before ischemia increases tissue S-nitrosothiol levels, improves postischemic contractile dysfunction, and attenuates necrosis. The mechanism of cardioprotection requires the uptake of CysNO via the L-AT and does not seem to involve NO release either during CysNO exposure or during ischemia. This suggests that the protective effects of CysNO are mediated through the posttranslational modification of cellular proteins through an NO-independent transnitrosation mechanism.

Caloric restriction restores ischemic tolerance in aged hearts: Effects on pro-survival kinases

It has been reported that calorie restriction (CR) improves ischemic tolerance in aged myocardium, though mechanisms by which this occurs are not well understood. We sought to identify potential involvement of pro-survival kinases in this cardioprotective response. To this end, hearts from young and aged mice (standard diet or CR) were isolated and perfused. All hearts were subjected to 25 min global ischemia and 45 min reperfusion. Hearts were then frozen for Western analysis. Young hearts showed a moderate degree of ischemic tolerance with > 50% recovery of contractile function (EDP, 21 ᠲ mm Hg; LVDP, 57 ᠴ%). Standard diet aged hearts displayed a significantly greater degree of post-ischemic contractile dysfunction (EDP, 43 ᠲ mm Hg; LVDP, 25 ᠳ%). In stark contrast, CR aged hearts recovered similarly to young hearts (EDP, 7 ᠱ mm Hg; LVDP, 60 ᠳ%). Moreover, both LDH and cardiac troponin release was significantly elevated in the standard diet aged mouse, but equivalent to the young hearts in the aged CR group. Examination of pro-survival kinases (Akt, p70S6K, GSK3 ERK1/2 and p38 MAPK) primarily reveals an increase in both phosphorylated and total Akt and GSK3ߠwith CR in aged hearts. In addition, ratios of phosphorylated:total ERK1/2 and p38 MAPK were identical between young and aged CR. While significantly different from young hearts, there was no distinction in p70S6K observed in the two aged diets. In summary, we show that caloric restriction effectively restores ischemic tolerance in aged hearts, possibly via alterations in Akt, GSK3 ERK1/2 and p38 MAPK.

Mitochondrial contribution to the molecular mechanism of heart acclimatization to chronic hypoxia: role of nitric oxide

A remarkable number of adaptive responses; including changes in the cardiovascular, respiratory and hematologic systems; takes place during acclimatization to natural or simulated high altitude. This adaptation to chronic hypoxia confers the heart an improved tolerance to all major deleterious consequences of acute O2 deprivation, not only reducing infarct size but also alleviating post-ischemic contractile dysfunction and ventricular arrhythmias. There is growing evidence about the involvement of mitochondria and NO in the establishment of cardioprotection. This review focuses on evidence about the putative role of different effectors of heart acclimatization to chronic hypoxia. Along with classical parameters, we consider NO, specially that generated by mtNOS, mitochondrial respiratory chain, mitoK(ATP) channels, reactive oxygen species and control of gene expression by HIF-1.

Inhibition of MMP-2 activation and release as a novel mechanism for HDL-induced cardioprotection

High density lipoproteins (HDL) protect the heart against ischemia/reperfusion (I/R) injury, and matrix metalloproteinase-2 (MMP-2) directly contributes to cardiac contractile dysfunction after I/R. To investigate the possible involvement of MMP-2 inhibition in HDL-mediated cardioprotection, isolated rat hearts underwent 20 min of low-flow ischemia and 30 min of reperfusion. Plasma-derived and synthetic HDL attenuated the I/R-induced cardiac MMP-2 activation and release in a dose-dependent way. The attenuation of I/R-induced MMP-2 activation by HDL correlated with the reduction of post-ischemic contractile dysfunction and cardiomyocyte necrosis. These results indicate prevention of MMP-2 activation as a novel mechanism for HDL-mediated cardioprotection.

Increased superoxide anion production is associated with early atherosclerosis and cardiovascular dysfunctions in a rabbit model

Hypercholesterolemia (HC) has been associated with impairment of vascular and myocardial functions. As HC could generate an alteration in the oxidative status, we studied the effects of a 1-month cholesterol diet on cardiovascular oxidative stress. New Zealand rabbits received cholesterol (1%) or normal chow for 1 month. At 30 days, superoxide anion levels, assessed by ESR spectroscopy, NAD(P)H oxidase (NOX) activity, and dihydroethidium (DHE) staining of aortas were higher in the cholesterol-fed (CF) group compared with control (respectively, 4.0 +/- 0.6 Arbitrary Units/mg (AU/mg) vs. 2.6 +/- 0.3, p < 0.05; 4231 +/- 433 vs. 2931 +/- 373 AU/mg, p<0.05; 21.4 +/- 1.2 vs. 12.9 +/- 1.7% fluorescence/mm2, p < 0.001). NOX gp91 phox and p67 phox expression in the aortas were higher in the CF group vs. control (1.5 +/- 0.2 vs. 0.5 +/- 0.2, p < 0.001; 0.9 +/- 0.2 vs. 0.3 +/- 0.2, p<0.05). The endothelium-dependent relaxation evaluated on the iliac arteries was higher in control than in the CF group (64.8 +/- 10.1 vs. 13.1 +/- 3.70%, p<0.001). The cardiac diastolic pressure estimated on isolated hearts was higher in the CF group than in control (21.1 +/- 4.1 vs. 10.3 +/- 1.4 mmHg, p<0.05) after 60 min of ischemia. Hypercholesterolemia induced increased levels of superoxide in the aortas and a higher expression of NOX subunits, associated with altered vasorelaxation. The increased diastolic pressure observed in hearts, consistent with a post-ischemic contractile dysfunction might be mediated by the production of superoxide.

Controlled reperfusion after hypothermic heart preservation inhibits mitochondrial permeability transition-pore opening and enhances functional recovery

We investigated whether low-pressure reperfusion may attenuate postischemic contractile dysfunction, limits necrosis and apoptosis after a prolonged hypothermic ischemia, and inhibits mitochondrial permeability transition-pore (MPTP) opening. Isolated rats hearts (n = 72) were exposed to 8 h of cold ischemia and assigned to the following groups: 1) reperfusion with low pressure (LP = 70 cmH(2)O) and 2) reperfusion with normal pressure (NP = 100 cmH(2)O). Cardiac function was assessed during reperfusion using the Langendorff model. Mitochondria were isolated, and the Ca(2+) resistance capacity (CRC) of the MPTP was determined. Malondialdehyde (MDA) production, caspase-3 activity, and cytochrome c were also assessed. We found that functional recovery was significantly improved in LP hearts with rate-pressure product averaging 30,380 +/- 1,757 vs. 18,000 +/- 1,599 mmHg/min in NP hearts (P < 0.01). Necrosis, measured by triphenyltetrazolium chloride staining and creatine kinase leakage, was significantly reduced in LP hearts (P < 0.01). The CRC was increased in LP heart mitochondria (P < 0.01). Caspase-3 activity, cytochrome c release, and MDA production were reduced in LP hearts (P < 0.001 and P < 0.01). This study demonstrated that low-pressure reperfusion after hypothermic heart ischemia improves postischemic contractile dysfunction and attenuates necrosis and apoptosis. This protection could be related to an inhibition of mitochondrial permeability transition.

5-HT 2 receptor blocker sarpogrelate prevents downregulation of antiapoptotic protein Bcl-2 and protects the heart against ischemia–reperfusion injury

Even though reperfusion is the treatment of choice in patients admitted with acute myocardial infarction, reperfusion itself has been demonstrated to activate various pathological factors especially following procedures of cardiac revascularization. 5-hydroxytryptamine (5HT) is one such factor activated during reperfusion and is known to trigger the post ischemic contractile dysfunction and pathological apoptosis. Here we demonstrate the potential effects of the 5-HT 2A antagonist sarpogrelate in protecting the myocardium against reperfusion injury of heart. Male Wistar rats weighing between 220 and 240 g were subjected to 30 min left coronary artery (LCA) occlusion and 120 min reperfusion. Sarpogrelate (4 mg/kg) was infused intravenously for 30 min either before LCA occlusion or at reperfusion. Following reperfusion the samples were collected for infarction area, immunohistochemistry, western blotting and myocardial metabolite analysis. Sarpogrelate infusion before ischemia resulted in (a) significant recovery of post ischemic cardiac functions (LVDP, EDP), (b) significant reduction in the infarct size among the risk area after triphenyl tetrazolium chloride staining ( p < 0.001), (c) decreased tissue water content ( p < 0.05), (d) well preserved myocardial ATP ( p < 0.05), (e) reduction in Bcl-2 downregulation and caspase 3 activation and (g) less prevalence of apoptotic cells (3.1 ± 0.4% to 15.2 ± 0.6%, drug versus control). Treating the rats with sarpogrelate during reperfusion also showed similar results. This study thus demonstrates the protective effects of sarpogrelate and supports the role for 5-HT2A inhibition in preventing the reperfusion injury of the heart.

β2-Adrenergic stimulation is involved in the contractile dysfunction of the stunned heart

Endogenous catecholamines released during myocardial ischemia have been considered both to aggravate cell injury and exacerbate arrhythmias and to exert a protective action on the post-ischemic contractile function. The present work was addressed to look for evidence to explain this controversy. The effects of cardiac catecholamine depletion and of alpha- and beta-adrenoceptor (AR) blockade on the post-ischemic contractile dysfunction, as well as its possible relationship with cardiac oxidative stress, were studied in isolated and perfused rat hearts submitted to 20 min of ischemia and 30 min of reperfusion (stunning). Catecholamine depletion improves the contractile recovery in the stunned heart. This mechanical effect was associated with decreased levels of lipid peroxidation. A similar enhancement of the contractile function during reperfusion was detected after the simultaneous blockade of alpha 1- and beta-ARs with prazosin plus propranolol. To ascertain which specific AR pathway was involved in the effects of catecholamines on the stunned heart, selective AR blockers, prazosin (alpha 1-blocker), atenolol (beta 1-blocker), ICI 118,551 (beta 2-blocker) and selective inhibitors of Gi-PI3K pathway (pertussis toxin and wortmannin) were alternatively combined. The results indicate that catecholamines released during ischemia exert a dual action on the contractile behavior of the stunned heart: a deleterious effect, related to the activation of the beta 2-AR-Gi-PI3K-pathway, which was counteracted by a beneficial effect, triggered by the stimulation of alpha 1-AR. Neither the depression nor the enhancement of the post-ischemic contractile recovery were related with the increase in ROS formation induced by endogenous catecholamines.

Cardioprotection induced by cardiac-specific overexpression of fibroblast growth factor-2 is mediated by the MAPK cascade

Our laboratory showed previously that cardiac-specific overexpression of FGF-2 [FGF-2 transgenic (Tg)] results in increased recovery of contractile function and decreased infarct size after ischemia-reperfusion injury. MAPK signaling is downstream of FGF-2 and has been implicated in other models of cardioprotection. Treatment of FGF-2 Tg and wild-type hearts with U-0126, a MEK-ERK pathway inhibitor, significantly reduced recovery of contractile function after global low-flow ischemia-reperfusion injury in FGF-2 Tg (86 +/- 2% vehicle vs. 66 +/- 4% U-0126; P < 0.05) but not wild-type (61 +/- 7% vehicle vs. 67 +/- 7% U-0126) hearts. Similarly, MEK-ERK inhibition significantly increased myocardial infarct size in FGF-2 Tg (12 +/- 3% vehicle vs. 31 +/- 2% U-0126; P < 0.05) but not wild-type (30 +/- 4% vehicle vs. 36 +/- 7% U-0126) hearts. In contrast, treatment of FGF-2 Tg and wild-type hearts with SB-203580, a p38 inhibitor, did not abrogate FGF-2-induced cardioprotection from postischemic contractile dysfunction. Instead, inhibition of p38 resulted in decreased infarct size in wild-type hearts (30 +/- 4% vehicle vs. 11 +/- 2% SB-203580; P < 0.05) but did not alter infarct size in FGF-2 Tg hearts (12 +/- 3% vehicle vs. 14 +/- 1% SB-203580). Western blot analysis of ERK and p38 activation revealed signaling alterations in FGF-2 Tg and wild-type hearts during early ischemia or reperfusion injury. In addition, MEK-independent ERK inhibition by p38 was observed during early ischemic injury. Together these data suggest that activation of ERK and inhibition of p38 by FGF-2 is cardioprotective during ischemia-reperfusion injury.

Low-pressure reperfusion alters mitochondrial permeability transition

We hypothesized that low-pressure reperfusion may limit myocardial necrosis and attenuate postischemic contractile dysfunction by inhibiting mitochondrial permeability transition pore (mPTP) opening. Male Wistar rat hearts (n = 36) were perfused according to the Langendorff technique, exposed to 40 min of ischemia, and assigned to one of the following groups: 1) reperfusion with normal pressure (NP = 100 cmH(2)O) or 2) reperfusion with low pressure (LP = 70 cmH(2)O). Creatine kinase release and tetraphenyltetrazolium chloride staining were used to evaluate infarct size. Modifications of cardiac function were assessed by changes in coronary flow, heart rate (HR), left ventricular developed pressure (LVDP), the first derivate of the pressure curve (dP/dt), and the rate-pressure product (RPP = LVDP x HR). Mitochondria were isolated from the reperfused myocardium, and the Ca(2+)-induced mPTP opening was measured using a potentiometric approach. Lipid peroxidation was assessed by measuring malondialdehyde production. Infarct size was significantly reduced in the LP group, averaging 17 +/- 3 vs. 33 +/- 3% of the left ventricular weight in NP hearts. At the end of reperfusion, functional recovery was significantly improved in LP hearts, with RPP averaging 10,392 +/- 876 vs. 3,969 +/- 534 mmHg/min in NP hearts (P < 0.001). The Ca(2+) load required to induce mPTP opening averaged 232 +/- 10 and 128 +/- 16 microM in LP and NP hearts, respectively (P < 0.001). Myocardial malondialdehyde was significantly lower in LP than in NP hearts (P < 0.05). These results suggest that the protection afforded by low-pressure reperfusion involves an inhibition of the opening of the mPTP, possibly via reduction of reactive oxygen species production.

Effect of ischemia and reperfusion on protein oxidation in isolated rabbit hearts.

Reactive oxygen species and other oxidants are involved in the mechanism of postischemic contractile dysfunction, known as myocardial stunning. The present study investigated the oxidative modification of cardiac proteins in isolated Langendorff-perfused rabbit hearts subjected to 15 min normothermic ischemia followed by 10 min reperfusion. Reperfusion under these conditions resulted in only 61.8+/-2.7 % recovery of developed pressure relative to preischemic values and this mechanical dysfunction was accompanied by oxidative damage to cardiac proteins. The total sulfhydryl group content was significantly reduced in both ventricle homogenates and mitochondria isolated from stunned hearts. Fluorescence measurements revealed enhanced formation of bityrosines and conjugates of lipid peroxidation-end products with proteins in cardiac homogenates, whereas these parameters were unchanged in the mitochondrial fraction. Reperfusion did not alter protein surface hydrophobicity, as detected by a fluorescent probe 1-anilino-8-naphthalenesulfonate. Our results indicate that oxidation of proteins in mitochondria and possibly in other intracellular structures occurs during cardiac reperfusion and might contribute to ischemia-reperfusion injury.

Effect of free radical scavengers on myocardial function and Na+, K+-ATPase activity in stunned rabbit myocardium

Objectives. The role of reactive oxygen species (ROS) in the mechanism of myocardial stunning was investigated. Material and methods. Isolated Langendorff-perfused rabbit hearts were subjected to 15 min normothermic ischemia followed by 10 min reperfusion with Krebs-Henseleit solution±mannitol or histidine. Results. In hearts reperfused without free radical scavenger the left ventricular developed pressure as well as its maximal positive and negative first derivatives (+dP/dt, −dP/dt) was significantly depressed, whereas end diastolic pressure (LVEDP) increased when compared to preischemic values. Treatment with mannitol had little protective effects, whereas singlet oxygen scavenger histidine significantly improved the recovery of LVEDP and −dP/dt. Sarcolemmal Na+, K+-ATPase activity (control, 400±41 nmol Pi.min−1.mg−1) was depressed in untreated stunned hearts (260±27 nmol Pi.min−1.mg−1), but was almost completely recovered in hearts pretreated with histidine (364±27 nmol Pi.min−1.mg−1). The inhibition of Na+, K+-ATPase was only slightly prevented by mannitol (302±29 nmol Pi.min−1.mg−1l). Conclusions. The results suggest that ROS-induced inhibition of Na+, K+-ATPase activity is involved in the mechanism of postischemic contractile dysfunction and support the view that singlet oxygen may be one of the major causes of oxidative injury during ischemia and reperfusion.