Significant Reduction In Infarct Size Research Articles

Inhaled Sevoflurane Produces Better Delayed Myocardial Protection at 48 Versus 24 Hours After Exposure

Ischemia preconditioning produces a delayed window of cardioprotection against subsequent ischemia and reperfusion injury. Contradictory results have been reported regarding the ability of inhaled anesthetics to produce similar effects. Our investigation was designed to test whether inhaled sevoflurane is capable of producing a delayed window of anesthetic preconditioning and to compare the differences at 24 and 48 h after exposure. Male Fischer-344 rats, 2-4 mo old, were exposed to sevoflurane (2.5% for 60 min). Twenty-four or 48 h after exposure, the hearts were isolated and perfused for 30 min (equilibration) followed by 25 min of ischemia and then 60 min of reperfusion. Control hearts received no treatment before ischemia. Left ventricular (LV) function, creatine kinase (CK), and infarct size (IS) were measured. Nuclear magnetic resonance was used to measure Na+(i), [Ca2+]i, and pH(i). There was improved LV function and significant reduction in IS and CK and in both the 24- and 48-h delayed groups compared with the controls. There was also a significant recovery of LV function and reduction in IS and CK in the 48-h group when compared with the 24-h group. There was significant adenosine triphosphate preservation in both the 24- and 48-h groups, as well as a significant reduction in acidosis, [Ca2+]I, and Na+(i) in response to ischemia in both the groups versus the control. Sevoflurane is capable of producing a delayed window of preconditioning, and it takes more than 24 h to produce maximal protective effects.

Treatment With the Gap Junction Modifier Rotigaptide (ZP123) Reduces Infarct Size in Rats With Chronic Myocardial Infarction

Treatment with non-selective drugs (eg, long-chain alcohols, halothane) that reduce gap junction intercellular communication (GJIC) is associated with reduced infarct size after myocardial infarction (MI). Therefore, it has been suggested that gap junction intercellular communication stimulating compounds may increase infarct size. The antiarrhythmic peptide analogue rotigaptide (ZP123) increases cardiac gap junction intercellular communication and the purpose of the present study was to examine the effects of rotigaptide treatment on infarct size. Myocardial infarction was induced in male rats by ligation of the left anterior descending artery (LAD). Rats (n = 156) were treated with rotigaptide at three dose levels or vehicle from the onset of ischemia and for 3 weeks following LAD occlusion. Infarct size was determined using histomorphometry after 3 weeks treatment. Rotigaptide treatment producing steady state plasma levels of 0.8 +/- 0.1, 5.5 +/- 0.5, and 86 +/- 8 nmol/L had no effect on mortality, but reduced infarct size to 90 +/- 10% (P = 0.41), 67 +/- 7% (P = 0.005), and 82 +/- 7% (P = 0.13), respectively relative to vehicle-treated myocardial infarction rats (100 +/- 12%). In contrast to what was predicted, our data demonstrates that rotigaptide treatment was associated with a significant infarct size reduction. We conclude that whereas treatment with non-selective inhibitors of gap junction intercellular communication cause a reduction in infarct size, this information cannot be extrapolated to the effects of compounds that selectively increase gap junction intercellular communication.

Similarities between Ischemic Preconditioning and 17??-Estradiol Mediated Cardiomyocyte KATP Channel Activation Leading to Cardioprotective and Antiarrhythmic Effects during Ischemia/Reperfusion in the Intact Rabbit Heart

The aims of our present work were to assess whether treatment with either ischemic preconditioning (IPC) or 17beta-estradiol or both combined produce proarrhythmic or antiarrhythmic effects, and whether opening of the sarcolemmal or mitochondrial KATP channels is relatable to this effect; to assess biochemically the effects of IPC and/or 17beta-estradiol on oxidant stress and antioxidant defenses in the myocardium; to examine the effects of nitric oxide (NO) synthase inhibitor, Nomega-nitro-L-arginine methyl ester (L-NAME) pretreatment in rabbits treated with either IPC or 17beta-estradiol (because 17beta-estradiol evoked NO release has been implicated in KATP activation and IPC); and examine the effects of ischemic preconditioning and 17beta-estradiol on myocardial energy metabolism during ischemia and reperfusion in a well-standardized model of reperfusion arrhythmias in anesthetized adult male New Zealand White rabbits (n = 124) subjected to 30 minutes occlusion of the left coronary artery followed by 120 minutes of reperfusion. Pretreatment with either 17beta-estradiol (10 microg/kg, i.v.) or one cycle of ischemic preconditioning prior to the period of coronary occlusion offers significant infarct size reduction (18.6 +/- 2.2% and 19.4 +/- 1.9%, respectively versus 40.1 +/- 3.9% in saline control and 39.2 +/- 3.2% in vehicle control groups; P < 0.01) and antiarrhythmic effects. Both 17beta-estradiol and ischemic preconditioning treatment significantly attenuated the incidence of life-threatening arrhythmias like sustained VT (13% and 13%, respectively versus 100% in saline control and 100% in vehicle control groups; P < 0.001) and other arrhythmias (25% and 25%, respectively versus 100% in saline control and 100% in vehicle control groups; P < 0.001), and were quite effective in increasing the number of animals that survived without developing any arrhythmia during ischemia and reperfusion. 5-hydroxydecanoate(5-HD; 5 mg/kg, i.v.) alone offered no cardioprotective and antiarrhythmic activities. Pretreatment with 5-HD but not HMR 1883 (3 mg/kg, i.v.) abolished the beneficial effects of 17beta-estradiol and ischemia preconditioning on reperfusion-induced arrhythmias and cardioprotection suggesting that such effects have been achieved via the selective activation of cardiomyocyte mitochondrial KATP channels rather than sarcolemmal KATP channels. The reduced reperfusion arrhythmic incidence and durations induced by estrogen was not significantly altered by ICI 182 720 (2.5 mg/kg, i.v.). The lack of effect of ICI 182 720 on antiarrhythmic and infarct-limiting effects of 17beta-estradiol and ischemic preconditioning suggest that these favorable effects are rapid, direct, and non-genomic effects. This study demonstrates similarities between 17beta-estradiol and ischemic preconditioning of the rabbit myocardium in terms of cardioprotection, antiarrhythmic, and metabolic activities. Ischemic preconditioning and 17beta-estradiol appear to share a final common effector; the mitochondrial KATP channel.

Neuroprotective effects of KCL-440, a new poly(ADP-ribose) polymerase inhibitor, in the rat middle cerebral artery occlusion model

It is reported that ischemic brain injury is mediated by the activation of poly(ADP-ribose) polymerase (PARP). In this study, we examined the pharmacological profile of KCL-440, a new PARP inhibitor, and its neuroprotective effects in the rat acute cerebral infarction model induced by photothrombotic middle cerebral artery (MCA) occlusion. In an in vitro study, KCL-440 exhibited potency with regard to inhibition of PARP activity, with an IC 50 value of 68 nM. An in vivo pharmacokinetic study showed that the brain concentration of KCL-440 was sufficient to inhibit PARP activity during the intravenous infusion of KCL-440 at the rate of 1 mg/kg/h. KCL-440 at various doses or saline was administered for 24 h immediately after the MCA occlusion. Administration of KCL-440 led to a dose-dependent reduction in the infarct size at 24 h after MCA occlusion. Infarct sizes were 44.8% ± 3.0% ( n = 8), 40.5% ± 1.1% ( n = 8), 38.2% ± 1.4% ( n = 8), 35.1% ± 2.1% ( n = 8), 34.2% ± 2.3% ( n = 7), 32.6% ± 1.9% ( n = 8), and 31.0% ± 2.1% ( n = 5) at doses of 0, 0.01, 0.03, 0.1, 0.3, 1.0, and 3.0 mg/kg/h. When compared to the control group, a statistically significant difference was observed in the doses that were higher than 0.03 mg/kg/h. When the infusion of KCL-440 (1 mg/kg/h, n = 8) was started at 1 h after the MCA occlusion, a significant reduction in infarct size was observed; this was not observed when KCL-440 infusion was started 2 or 3 h after the MCA occlusion. Furthermore, increased poly(ADP-ribose) immunostaining was confirmed at the ischemic border zone 2 h after the MCA occlusion, and it was reduced by KCL-440 treatment. These results suggest that KCL-440 is a possible neuroprotective agent with high blood–brain barrier permeability and high PARP inhibitory activity.

Short‐ and long‐term effects of melatonin on myocardial post‐ischemic recovery

Melatonin, the chief secretory product of the pineal gland, has been shown to protect the heart against ischemia-reperfusion injury. This was attributed to its free radical scavenging and broad-spectrum antioxidant properties. The possibility that melatonin may act via its receptor and intracellular signaling, has not yet been addressed in this regard. In all previous studies, only the acute effects of melatonin on the heart, were evaluated. The aims of the present study were to: (i) compare the acute and long-term effects of melatonin on infarct size and functional recovery of the ischemic heart, and (ii) evaluate the role of the melatonin receptor in cardioprotection. For evaluation of the short-term effects of melatonin on contractile recovery and infarct size, the isolated perfused working rat heart was subjected to 20 min global ischemia or 35 min regional ischemia respectively, and melatonin (25-50 microm) administered either before and during reperfusion, or before ischemia or during reperfusion after ischemia. The melatonin receptor was manipulated using luzindole and N-acetyltryptamine. The long-term effects of melatonin were evaluated 24 hr after melatonin administration (2.5 or 5.0 mg/kg, i.p.) or after oral administration for 7 days (20 or 40 microg/mL). Infarct size and mechanical recovery during reperfusion of the working heart were used as endpoints. Melatonin (50 microm), when administered either before and during reperfusion after ischemia or during reperfusion only, significantly improved cardiac output and work performance and reduced infarct size compared with untreated controls. Luzindole (5 microm), a melatonin receptor antagonist, abolished these cardioprotective effects. Long-term administration of melatonin (i.p. or orally for 7 days) caused a significant reduction in infarct size of hearts subjected to 35 min regional ischemia. The cardioprotection persisted for 2-4 days after discontinuation of treatment. In summary, the results obtained suggest that melatonin induces short- as well as long-term protection and that the melatonin receptor is also involved in its cardioprotective actions.

Central role of neuronal IkappaBalpha kinase (IKK) in cerebral ischemia

The transcription factor NF-kappaB is activated in cerebral ischemia and promotes ischemic neurodegeneration with inflammatory and apoptotic effects. Activation of NF-kappaB is triggered by IkappaB-kinase (IKK) but may also occur independent of IKK. To find out if IKK is critical for damage in cerebral ischemia, we used both genetic and pharmacological aproaches to reduce IKK activity. Mice with floxed IKK-2 allels were crossed with a nestin-Cre line resulting in selective deficiency of IKK-2 in neural cells or with a CaMKII-Cre line resulting in selective deficiency in neurons. Western-blot-analysis of brain tissue was performed to prove the deficiency of IKK-2 protein. Furthermore, we expressed a dominant-negative mutant of IKK-2 in neurons, which inhibits both subunits of IKK. As a pharmacological approach, we injected BMS-345541, a selective inhibitor of IKK, or NaCl 0.9% as control icv at different time points. All mice underwent an occlusion of the distal branches of the middle cerebral artery (MCAO) by coagulation as a model of permanent cerebral ischemia. IKK was rapidly activated after MCAO followed by decrease of total IkappaB protein. These processes were reduced in mice with genetic inhibition of IKK. We found a significant reduction of infarct size 48 hours after MCAO in all genetically altered mouse lines. Equally, BMS-345541 in different doses reduced the infarct size up to 60% when injected as late as 4.5 hours after MCAO. The neuroprotective effect of BMS-345541 was confirmed when infarct size was measured 2 weeks after MCAO. TUNEL staining evaluated by laser scanning cytometry showed a reduction of apoptotic signs after treatment with BMS-345541. In summary these data clarify the central role of IKK in ischemic brain damage as shown consistently by different genetic and pharmacological approaches. Above all our experiments suggest that IKK is a worthwile drug target in stroke therapy with a promising therapeutic time frame.

Assessment of lithium treatment using fMRI during stroke recovery in rats

An important function of lithium as a neuroprotective agent has been documented previously. Significant reduction in infarct size and DNA damages with the corresponding improvement of neurological scores were reported as results of post stroke lithium administration. In the current study, therapeutic efficacy of chronic lithium treatment was assessed by various MRI-derived parameters (i.e., apparent diffusion coefficient (ADC), fractional anisotropy (FA), and vessel size index (VSI)) and quantified with fMRI activity using electrical stimulation of rat forelimbs. The fMRI responses were examined by measuring the local hemodynamic MRI signal intensity based on blood oxygenation level dependence (BOLD) and cerebral blood volume weighted (CBVw) responses. The ipsilesional fMRI activations in the somatosensory (SS) cortex of lithium-treated (n=6) and saline-treated rats (n=6) two weeks after middle cerebral artery occlusion (MCAO) were compared. ADC and FA values of local brain tissues were significantly correlated with the magnitude of fMRI responses for the lithium-treated rats while no such correlations were seen in the saline controls. The ipsilesional/contralesional BOLD signal intensity response ratios of lithium-treated rats were larger than those of saline-treated control rats. In contrast, the CBVw response ratios were similar between two groups. These results demonstrated that the lithium-treatment of post-stroke animal models positively enhanced BOLD fMRI response in the lesion hemisphere. Overall activation volume and magnitude were larger for saline-treated rats than the lithium-treated group in both contra and ipsilesional somatosensory cortices. However, for both BOLD and CBVw responses, the mean activated volume ratio (ipsilesional/contralesional) was higher for lithium-treated rats than controls. The CBVw activation signal magnitude ratios (Figure 1 upper panel) were significantly higher than BOLD ratios for the control animals; however, similar for the lithium-treated rats. As shown in Figure 1 (bottom panel), functional status index (FSI) which represents the mean ratio between BOLD and CBVw ipsi/contralesional activation signal intensity ratio for the individual somatosensory cortex was significantly higher for lithium-treated rats than the control group. From the acquired ADC maps, ex vacuo dilation of the ipsilesional lateral ventricles was identified for most of rats while other structural damages (e.g., FA) were also evident at this late stage (2 weeks) of recovery. Among the acquired structural parameters, ADC and FA were significantly correlated with CBVw fMRI activation magnitude (i.e., area under the curve) for the lithium-treated rats while no such correlations were found for the saline controls. When compared to the contralesional values, the ipsilesional mean VSI ratio (ipsi/contra) was larger for the lithium-treated rats; however, the micro vascular volume (i.e., Delta-R2) ratio (ipsi/contra) was larger for control rats. These results imply that possible vascular transformation affects fMRI characteristics.

Pretreatment with an adenosine A1 receptor agonist and lidocaine: A possible alternative to myocardial ischemic preconditioning

The heart possesses an extraordinary ability to remember short episodes of sublethal ischemia and reperfusion (angina), which protects the myocardium and coronary vasculature from a subsequent lethal insult, a phenomenon known as ischemic preconditioning. A therapeutic goal for more than 2 decades has been to develop a pharmacologic mimetic comparable with ischemic preconditioning. Our aim was to investigate the preconditioning effect of a new combinatorial therapy targeting adenosine A1 receptors and voltage-dependent sodium fast channels in the in vivo rat model of regional ischemia. Ischemia-reperfusion was achieved by placing a reversible tie around the left coronary artery in anesthetized and ventilated Sprague-Dawley rats (n = 37). Rats were randomly assigned to 1 of 5 groups: (1) saline control (n = 13); (2) ischemic preconditioning (n = 6); (3) lidocaine only (608 microg . kg -1 . min -1 , n = 5); (4) adenosine A1 receptor agonist 2-chloro-N6-cyclopentyladenosine (CCPA; 5 microg/kg, n = 7); and (5) CCPA plus lidocaine (n = 6). Ischemic preconditioning was achieved by using 3 cycles of ischemia and reperfusion lasting 3 minutes each. Lidocaine was infused continuously 5 minutes before and throughout 30 minutes of ischemia and ceased at reperfusion. A bolus of CCPA was infused 5 minutes before ligation along with a constant infusion of lidocaine (as above). All animals were reperfused for 120 minutes for infarct size measurement. Fifty-four percent of saline control rats, 17% of ischemic preconditioning-treated rats, and 29% of CCPA-treated rats died during ischemia from ventricular fibrillation. Infarct size of saline control animals was 61% +/- 5%. Pretreating with CCPA and lidocaine infusion resulted in no deaths, no severe arrhythmias, and significant infarct size reduction compared with that seen in saline control animals (P < .05). Remarkably, infarct size reduction in CCPA plus lidocaine-treated rats (12% +/- 4%) was equivalent to that achieved with ischemic preconditioning (11% +/- 3%), whereas infarct size in rats undergoing CCPA-only and lidocaine-only treatments was 42% +/- 7% and 60% +/- 6%, respectively. Although CCPA plus lidocaine treatment reduced heart rate, mean arterial pressure, and systolic pressure during ischemia, no correlation was found between these variables and infarct size reduction. We conclude that activating adenosine A1 receptor subtype with CCPA and concomitantly modulating sodium fast channels with lidocaine was comparable with ischemic preconditioning and might offer a new therapeutic window to minimize myocardial damage during surgical ischemia and reperfusion.

Letter Regarding Article by Shyu et al, “Functional Recovery of Stroke Rats Induced by Granulocyte Colony-Stimulating Factor–Stimulated Stem Cells”

To the Editor: Shuy and coworkers investigated the effect of granulocyte colony-stimulating factor (G-CSF) in a right cerebral artery ligation model in the rat.1 They reported that G-CSF treatment led to a significant reduction in infarct size, improved neural plasticity, and enhanced vascularization. They concluded that the latter effects are largely the result of mobilization of autologous hemopoietic stem cells into circulation, enhancing their translocation into ischemic brain, and thus significantly improving lesion repair. The authors omitted one important physiological mechanism in the brain—the impact of the collateral circulation (arteriogenesis) on the restoration of tissue perfusion on hypoperfusion. Our recent studies into brain arteriogenesis2–4 demonstrated for the first time that similar to the cardiac and peripheral circulation, cerebrovascular occlusions are partly compensated by the growth of preexisting collateral arteries (ie, by the active remodeling of the arterial vessel wall that is characteristic of …

Susceptibility of the heart to ischaemia–reperfusion injury and exercise‐induced cardioprotection are sex‐dependent in the rat

The cardioprotective effects of short-term exercise against myocardial ischaemia-reperfusion injury in male and female rats were examined. We subjected male and female rats to 0 (Sed; n = 8 males and 8 females), 1 (1 day; n = 10 males and 8 females), or 5 (5 day; n = 6 males and 6 females) days of treadmill running. Langendorff-perfused hearts underwent 1 h of regional ischaemia and 2 h of reperfusion, and infarct size (expressed as a percentage of the zone at risk; ZAR), left ventricular pressure development, and coronary flow were measured for each heart. Preischaemic pressure development and coronary flow did not differ between the sexes nor were they influenced by exercise. Sed females had significantly smaller infarct sizes (25 +/- 3%) than Sed male hearts (37 +/- 3%; P < 0.001). Short-term running significantly reduced infarct size following 1 day (27 +/- 3%; P < 0.05) and 5 days (30 +/- 4%; P < 0.10) of exercise in males. One day of running did not reduce infarct size in females (19 +/- 3%; P = NS), but 5 day females did show a significant reduction in infarct size (13 +/- 2%; P < 0.05). There was no relationship between postischaemic coronary vascular hyperaemia and infarct size across sexes or exercise training groups. Hearts from Sed females exhibited significantly higher manganese superoxide dismutase (MnSOD) protein expression than hearts from Sed males, but short-term exercise (neither 1 nor 5 days) did not alter MnSOD protein in either sex. Increased sarcolemmal ATP-sensitive K(+) (K(ATP)) channel subunit protein expression (SUR2A and/or K(ir)6.2) correlated closely with sex-dependent and exercise-acquired protection against myocardial infarction. These data indicate that: (1) sex-dependent and exercise-induced differences in the susceptibility of the heart to ischaemia-reperfusion injury are not associated with improved coronary flow or postischaemic hyperaemia; (2) increased MnSOD protein expression is not necessary for exercise-induced protection from infarction; and (3) one possible mechanism for sex-dependent and exercise-mediated reductions in infarct size involves an increased protein expression of cardiac sarcolemmal K(ATP) channels.

A Peroxisome Proliferator-Activated Receptor-γ Agonist Reduces Infarct Size in Transient but Not in Permanent Ischemia

Activators of peroxisome proliferator-activated receptor-gamma (PPARgamma), a member of the PPAR family, increase levels of CuZn-superoxide dismutase (SOD) in cultured endothelium, suggesting a mechanism by which it may exert its protective effect within the brain. These properties raise the question of whether a PPARgamma agonist may be neuroprotective in models of ischemia without reperfusion, in which oxidative injury is less prevalent. In 2 groups of rats, 90 minutes of middle cerebral artery (MCA) occlusion was followed by 1 day of reperfusion, with 1 group receiving pioglitazone (a PPARgamma agonist) starting 72 hours before MCA occlusion (MCAO) and continuing through the day of occlusion, whereas the other group received vehicle only. In 2 comparable groups, the MCA was occluded permanently. One day after occlusion, the animals were tested neurologically and infarct volumes were calculated. In a separate group, rats were treated with pioglitazone or vehicle for 4 days. Tissue was obtained from the cortex and the striatum 2 hours into reperfusion after 90 minutes of MCAO, and the tissue was examined for CuZn-SOD by Western blot. Results show a significant reduction in infarct size in the treated rats, with transient MCAO but not permanent MCAO. There was also an improvement in neurological score in the treated animals after transient MCAO. The level of CuZn-SOD was increased in the cortex in treated animals. These data, which show that a PPARgamma agonist reduces infarct size in transient but not permanent MCAO, suggest that the role of PPARgamma is specific to events occurring during reperfusion. Our data point to CuZn-SOD as the mediator of this neuroprotection.

Protective Effects of Evodiamine on Myocardial Ischemia-Reperfusion Injury in Rats

Previous investigations have indicated that the pharmacological activities of evodiamine, a major alkaloidal component of the dried, unripe fruit of Evodia rutaecarpa Bentham (Rutaceae), are associated with the stimulation of calcitonin gene-related peptide (CGRP) release and that CGRP protects the myocardium against ischemia-reperfusion injury. In the present study, we have examined whether evodiamine protects against myocardial ischemia-reperfusion injury in rats and whether the protective effects of evodiamine are related to the activation of capsaicin-sensitive sensory nerves. Rats were pretreated with evodiamine 10 min before the experiment, and then the left main coronary artery of rat hearts was subjected to 60 min occlusion followed by 180 min reperfusion. Infarct size, the activity of serum creatine kinase, serum concentrations of TNF-alpha and plasma concentrations of CGRP were measured. Pretreatment with evodiamine (30 or 60 microg/kg, i.v.) markedly increased the content of CGRP in plasma concomitantly with a significant reduction in infarct size, the activity of serum creatine kinase, and TNF-alpha level, and the effects of evodiamine were completely abolished by capsazepine (5.0 mg/kg, s.c.), a competitive vanilloid receptor antagonist. These results suggest that evodiamine exerts a protection against myocardial ischemia-reperfusion injury in rats and that the protective effects of evodiamine are related to stimulation of CGRP release via activation of vanilloid receptors.

Sarcolemmal KATP channel triggers delayed ischemic preconditioning in rats

Previous work from our laboratory has shown that the sarcolemmal K(ATP) channel (sK(ATP)) is required as a trigger for delayed cardioprotection upon exogenous opioid administration. We also established that the mitochondrial K(ATP) (mK(ATP)) channel is not required for triggering delayed delta-opioid-induced infarct size reduction. Because mechanistic differences have been found among delta-opioids and that due to ischemic preconditioning (IPC), we determined whether the triggering mechanism of delayed IPC-induced infarct size reduction involves either the sK(ATP) or mK(ATP). Male Sprague-Dawley rats received either sham surgery or IPC (3- to 5-min cycles of ischemia and reperfusion) 24 h before being subjected to 30 min of ischemia and 2 h of reperfusion. Infarct size was determined and expressed as a percentage of the area at risk, with significance compared with sham reported at P </= 0.001. A subset of both sham and IPC-treated rats received either the selective sK(ATP) channel antagonist, HMR-1098 (6 mg/kg), or the selective mK(ATP) channel antagonist, 5-hydroxydeconoic acid (5-HD; 10 mg/kg), given 5 min before IPC. Rats subjected to IPC demonstrated a significant reduction in infarct size compared with sham (29.2 +/- 4.7 vs. 59.3 +/- 2.5%, respectively; P </= 0.001). Prior administration of HMR-1098, but not 5-HD, abolished IPC-induced infarct size reduction (48.8 +/- 2.9 and 28.8 +/- 4.0%, respectively; P </= 0.001). Furthermore, administration of HMR 24 h after IPC, before index ischemia, did not abrogate IPC-induced infarct size reduction (33.0 +/- 5.0 vs. 29.2 +/- 4.7%, respectively; P </= 0.001). These data suggest that the sK(ATP) channel is required as a trigger but not a mediator for delayed IPC-induced infarct size reduction in rat hearts.

Acute adenosine preconditioning is mediated by p38 MAPK activation in discrete subcellular compartments

Although acute adenosine preconditioning (PC) is well established, the signaling pathways mediating this cardioprotection remain unclear. Because adenosine receptor agonists activate p38 MAPK and this kinase has been implicated in ischemic and pharmacological PC, the purpose of this study was to determine the role of p38 MAPK in acute adenosine receptor PC. The role of p38 MAPK activation in discrete subcellular compartments during ischemia-reperfusion was also determined. The following groups were used in an in vivo rat ischemia-reperfusion model: 1) control (10% DMSO i.v.), 2) the A(1)/A(2a) adenosine receptor AMP-579 (50 microg/kg i.v.), 3) AMP-579 + the A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, 100 microg/kg i.v.), 4) AMP-579 + the p38 MAPK inhibitor SB-203580 (1 mg/kg i.v.), and 5) SB-203580 alone. p38 MAPK activation was measured by Western blot analysis in cytosolic, mitochondrial, membrane, and nuclear/myofilament fractions obtained from hearts at preischemic, ischemic, and reperfusion time points. A significant reduction in infarct size was observed with AMP-579 PC, an effect blocked by DPCPX or SB-203580 pretreatment. AMP-579 treatment was associated with a significant increase in p38 MAPK activation in the nuclear/myofilament fraction before ischemia, whereas no activation of this kinase occurred during ischemia or reperfusion. In contrast, p38 MAPK was activated in the mitochondrial fraction by ischemia and in the cytosolic, mitochondrial, and membrane fractions by reperfusion in the control group. SB-203580 blocked the AMP-579-induced increase in phosphorylation of the downstream p38 substrate activating transcription factor-2. These results suggest a role for p38 MAPK activation in discrete subcellular compartments in acute adenosine A(1) receptor PC.

Sildenafil Citrate (Viagra) Induces Cardioprotective Effects after Ischemia/Reperfusion Injury in Infant Rabbits

Infants undergoing surgery for congenital heart disease are at risk for myocardial ischemia during cardiopulmonary bypass, circulatory arrest, or low-flow states. The purpose of this study was to demonstrate the effects of sildenafil, a selective phosphodiesterase-5 (PDE-5) inhibitor on myocardial functional improvement and infarct size reduction during ischemia/reperfusion injury in infant rabbits. Infant rabbits (aged 8 wk) were treated with sildenafil citrate (0.7 mg/kg i.v.) or normal saline 30 min before sustained ischemia for 30 min and reperfusion for 3 h. Transesophageal echocardiography (TEE) was used to assess left ventricular cardiac output (LVCO) and aortic velocity time integral (VTI). After ischemia/reperfusion, risk area was demarcated by Evan's blue dye and infarct size determined by computer morphometry of triphenyltetrazolium chloride-stained sections. The sildenafil-treated group had preservation and elevation in LVCO (143% of baseline, p < 0.05) and an elevated aortic VTI (145% of baseline, p < 0.05) after 30 min of ischemia compared with the control group LVCO (72% of baseline, p < 0.05) and aortic VTI (73% of baseline, p < 0.05). This is a statistically significant increase in LVCO and aortic VTI in the sildenafil group compared with controls (n = 6/group, p < 0.05). The sildenafil-treated group had significant reduction in infarct size (15.5 +/- 1.2 versus 33 +/- 2.3 in the saline group, % risk area, mean +/- SEM, n = 10-15/group, p < 0.05). For the first time, we have shown that sildenafil citrate promotes myocardial protection in infant rabbits as evidenced by postischemic preservation and elevation in LVCO and aortic VTI and reduction in infarct size.

Failure to demonstrate myocardial protective effects of the ultra short-acting calcium antagonist clevidipine in a closed-chest reperfusion porcine model.

Restoration of myocardial perfusion is essential in acute myocardial infarction for the salvaging of myocardial tissue. However, reperfusion per se can provoke myocardial necrosis within the jeopardized tissue. Yet, no intervention has been successfully applied to the clinical situation in this matter. Clevidipine, an ultra-short acting calcium antagonist, has, in open-chest animal models, shown to reduce the extent of reperfusion injury. In the present study we intended to reproduce those findings in a closed-chest porcine model with a clinically applicable set up. Pigs were subjected to balloon occlusion of the left anterior descending coronary artery (LAD) for 45 minutes. During 25 minutes, starting 1 minute prior to reperfusion, clevidipine, Intralipid, or saline was infused antegradely into the endangered myocardium. As no significant effects on infarct size were achieved, the model was modified. In a second phase, different anesthesias were evaluated addressing the same issue. Nonetheless no significant effects on infarct size were observed. Different techniques of occluding LAD, in an open-chest model, were investigated in a third phase, and revealed no significant differences between the techniques. However, when comparing all the closed- versus open-chest models, significant reduction in infarct size by the use of clevidipine was only obtained in the open-chest models. We could not demonstrate any significant myocardial protective effect with clevidipine in our porcine, closed-chest, acute infarct, and reperfusion model. However, in a modified open-chest model we obtained significant reduction in infarct size. Further studies are required to explain the discrepancies.

ERYTHROPOIETIN ATTENUATES THE TISSUE INJURY ASSOCIATED WITH HEMORRHAGIC SHOCK AND MYOCARDIAL ISCHEMIA

Here we investigate the effects of erythropoietin (EPO) on the tissue/organ injury caused by hemorrhagic shock (HS), endotoxic shock, and regional myocardial ischemia and reperfusion in anesthetized rats. Male Wistar rats were anesthetized with thiopental sodium (85 mg/kg i.p.) and subjected to hemorrhagic shock (HS; i.e., mean arterial blood pressure reduced to 45 mmHg for 90 min, followed by resuscitation with shed blood for 4 h), endotoxemia (for 6 h), or left anterior descending coronary artery occlusion (25 min) and reperfusion (2 h). HS and endotoxemia resulted in renal dysfunction and liver injury. Administration of EPO (300 IU/kg i.v., n = 10) before resuscitation abolished the renal dysfunction and liver injury in hemorrhagic, but not endotoxic, shock. HS also resulted in significant increases in the kidney of the activities of caspases 3, 8, and 9. This increase in caspase activity was not seen in HS rats treated with EPO. In cultured human proximal tubule cells, EPO concentration-dependently reduced the cell death and increase in caspase-3 activity caused by either ATP depletion (simulated ischemia) or hydrogen peroxide (oxidative stress). In the heart, administration of EPO (300 IU/kg i.v., n = 10) before reperfusion also caused a significant reduction in infarct size. In cultured rat cardiac myoblasts (H9C2 cells), EPO also reduced the increase in DNA fragmentation caused by either serum deprivation (simulated ischemia) or hydrogen peroxide (oxidative stress). We propose that the acute administration of EPO on reperfusion and/or resuscitation will reduce the tissue injury caused by ischemia-reperfusion of the heart (and other organs) and hemorrhagic shock.

The salvage potential of coronary sinus interventions: meta-analysis and pathophysiologic consequences

Objectives Intermittent coronary sinus occlusion has been described to be effective in salvaging ischemic myocardium. This meta-analysis aims to review the efficacy of intermittent coronary sinus occlusion and intermittent coronary sinus occlusion in combination with retroperfusion of arterial blood as methods of myocardial salvage. Methods A Medline search was performed to review the published literature on intermittent coronary sinus occlusion. The study inclusion criterion was a randomized, placebo-controlled trial with area of infarction (expressed as a percentage of the area at risk) as the primary end point. Results Seven experimental trials comprising 125 test animals were found that analyzed the effects of intermittent coronary sinus occlusion on ischemic damage during coronary occlusion. A further 5 studies comprising 88 animals were designed to evaluate the effect of intermittent coronary sinus occlusion in combination with retroperfusion of arterial blood on the infarct size. A meta-analysis of the 7 studies analyzing the effect of intermittent coronary sinus occlusion revealed a significant reduction in infarct size of 29.3% in the treatment group compared with that in the placebo group ( P < .001; 95% confidence interval, −40.9 to −17.7). A meta-analysis of the 5 trials analyzing the effect of intermittent coronary sinus occlusion in combination with retroperfusion revealed a reduction in infarct size of 39.4% in the treatment group compared with that in the placebo group ( P < .001; 95% confidence interval, −48.9 to −29.9). Comparison between intermittent coronary sinus occlusion and intermittent coronary sinus occlusion in combination with retroperfusion of arterial blood showed no statistical difference ( P = .19). An inverse relationship between achieved coronary sinus pressure increase per minute and infarct size could be found in the intermittent coronary sinus occlusion group ( r = −0.92; P < .007), whereas in combination with retroperfusion, there was a negative correlation both between achieved coronary sinus pressure and the amount of the retroperfusate and myocardial salvage ( r = −0.97; P < .004). Conclusions The use of intermittent coronary sinus occlusion and intermittent coronary sinus occlusion in combination with retroperfusion of arterial blood significantly decreases ischemic damage during coronary occlusions. Intermittent coronary sinus occlusion in combination with retroperfusion exhibits no significant profit in salvaging the ischemic myocardium in comparison with that provided by intermittent coronary sinus occlusion alone.

Adenosine and a nitric oxide donor enhances cardioprotection by preconditioning with isoflurane through mitochondrial adenosine triphosphate-sensitive K+ channel-dependent and -independent mechanisms.

Preconditioning with isoflurane has been shown to confer cardioprotection via activation of mitochondrial adenosine triphosphate-sensitive K+ (mito K(ATP)) channels. However, the relative contribution of mito K(ATP) channel and non-mito K(ATP) channel mechanisms to isoflurane-mediated cardioprotection has not been investigated. Isolated and buffer-perfused rat hearts were used. Flavoprotein fluorescence was monitored as an index for mito K(ATP) channel activity. Isovolumic left ventricular function and infarct size were measured as indices for cardioprotection. Flavoprotein fluorescence, which was monitored as an index for mito K(ATP) channel activity, was increased by isoflurane and a known mito K(ATP) channel opener, diazoxide, in a 5-hydroxydecanoate-sensitive manner. Although flavoprotein oxidation induced by diazoxide was dissipated soon after its removal from the buffer, flavoprotein oxidation induced by isoflurane was sustained after cessation of the treatment. The sustained increase in flavoprotein oxidation was associated with a significant reduction in infarct size after 30 min of ischemia followed by 120 min of reperfusion. Although adenosine and S-nitroso-N-acetyl-penicillamine each alone did not increase flavoprotein fluorescence, nor did they confer significant cardioprotection, coadministration of adenosine and S-nitroso-N-acetyl-penicillamine with isoflurane conferred a highly significant reduction of infarct size and improvement of left ventricular function without increasing flavoprotein oxidation over isoflurane alone. The early treatment with 5-hydroxydecanoate before and during preconditioning completely reversed flavoprotein oxidation and inhibited the infarct-sparing effect of isoflurane and combined preconditioning with isoflurane, adenosine, and S-nitroso-N-acetyl-penicillamine. The late treatment with 5-hydroxydecanoate after preconditioning abolished flavoprotein oxidation and the infarct-sparing effect of isoflurane but only partially inhibited cardioprotection conferred by the combined preconditioning, despite complete abrogation of flavoprotein oxidation. Mito K(ATP) channel activation is the essential trigger of both preconditioning with isoflurane and combined preconditioning with isoflurane, adenosine, and S-nitroso-N-acetyl-penicillamine. Mito K(ATP) channel activation is also a crucial mediator of cardioprotection afforded by preconditioning with isoflurane. However, enhanced cardioprotection conferred by combined preconditioning is mediated through both mito K(ATP) channel-dependent and -independent mechanisms.

The Low Molecular Weight Heparin Enoxaparin Reduces Infarct Size in a Rat Model of Temporary Focal Ischemia

Low molecular weight heparins (LMWHs) have significantly reduced infarct size in animal studies but they have not been effective in clinical trials, probably because they were administered after ischemic injury had become irreversible. The present study was designed to explore the temporal characteristics of the LMWH enoxaparin with the objective of determining the duration of the treatment window in a rat model of temporary focal ischemia. Focal cerebral ischemia was induced by the intraluminal suture, middle cerebral artery occlusion (MCAO) method. Enoxaparin (10 mg/kg) was administered subcutaneously twice; the first dose was administered to different groups of animals either 1 h before or 1.5, 3, or 5 h after MCAO, and the second 4, 6, 9, or 25 h after the first dose. At 48 h animals were tested for motor coordination and brains were removed for determination of infarct size. Results showed that infarct size and degree of motor impairment were a function of time of the first and the second treatments. If the first treatment was given 1 h prior to MCAO, significant reductions in infarct size were obtained when the second treatment was given up to 6 h after the first (5 h after MCAO), but not when the second dose was given at longer intervals. If the first treatment was given 1.5 h after stroke onset, reduced infarct size was observed only in the group treated for the second time 4 h after the first injection. If the first treatment was given 3 h after MCAO, infarct size was not reduced in any group. However, if enoxaparin was administered intravenously rather than subcutaneously, significant reductions in infarct size were obtained when the first dose was given as long as 5 h after MCAO. These findings indicate that enoxaparin can reduce infarct size in rats when administered prior to stroke onset and suggest that the drug might be considered for prophylactic treatment in a phase 3 clinical trial.