Size-limiting Effect Of Ischemic Preconditioning Research Articles

Modulation of Inula racemosa Hook Extract on Cardioprotection by Ischemic Preconditioning in Hyperlipidaemic Rats.

Hyperlipidemia (HL) is a major cause of ischemic heart diseases. The size-limiting effect of ischemic preconditioning (IPC), a cardioprotective phenomenon, is reduced in HL, possibly because of the opening of the mitochondrial permeability transition pore (MPTP). The objective of this study is to see what effect pretreatment with Inula racemosa Hook root extract (IrA) had on IPC-mediated cardioprotection on HL Wistar rat hearts. An isolated rat heart was mounted on the Langendorff heart array, and then ischemia reperfusion (I/R) and IPC cycles were performed. Atractyloside (Atr) is an MPTP opener. The animals were divided into ten groups, each consisting of six rats (n = 6), to investigate the modulation of I. racemosa Hook extract on cardioprotection by IPC in HL hearts Sham control, I/R Control, IPC control, I/R + HL, I/R + IrA + HL, IPC + HL, IPC + NS + HL, IPC + IrA+ HL, IPC + Atr + oxidative stress, mitochondrial function, integrity, and hemodynamic parameters are evaluated for each group. The present experimental data show that pretreatment with IrA reduced the LDH, CK-MB, size of myocardial infarction, content of cardiac collagen, and ventricular fibrillation in all groups of HL rat hearts. This pretreatment also reduced the oxidative stress and mitochondrial dysfunction. Inhibition of MPTP opening by Atr diminished the effect of IrA on IPC-mediated cardioprotection in HL rats. The study findings indicate that pretreatment with IrA e restores IPC-mediated cardioprotection in HL rats by inhibiting the MPTP opening.

Ischemic preconditioning protects the heart against ischemia-reperfusion injury in chronic kidney disease in both males and females

BackgroundUremic cardiomyopathy is a common cardiovascular complication of chronic kidney disease (CKD) characterized by left ventricular hypertrophy (LVH) and fibrosis enhancing the susceptibility of the heart to acute myocardial infarction. In the early stages of CKD, approximately 60% of patients are women. We aimed to investigate the influence of sex on the severity of uremic cardiomyopathy and the infarct size-limiting effect of ischemic preconditioning (IPRE) in experimental CKD.MethodsCKD was induced by 5/6 nephrectomy in 9-week-old male and female Wistar rats. Two months later, serum and urine laboratory parameters were measured to verify the development of CKD. Transthoracic echocardiography was performed to assess cardiac function and morphology. Cardiomyocyte hypertrophy and fibrosis were measured by histology. Left ventricular expression of A- and B-type natriuretic peptides (ANP and BNP) were measured by qRT-PCR and circulating BNP level was measured by ELISA. In a subgroup of animals, hearts were perfused according to Langendorff and were subjected to 35 min global ischemia and 120 min reperfusion with or without IPRE (3 × 5 min I/R cycles applied before index ischemia). Then infarct size or phosphorylated and total forms of proteins related to the cardioprotective RISK (AKT, ERK1,2) and SAFE (STAT3) pathways were measured by Western blot.ResultsThe severity of CKD was similar in males and females. However, CKD males developed more severe LVH compared to females as assessed by echocardiography. Histology revealed cardiac fibrosis only in males in CKD. LV ANP expression was significantly increased due to CKD in both sexes, however, LV BNP and circulating BNP levels failed to significantly increase in CKD. In both sexes, IPRE significantly decreased the infarct size in both the sham-operated and CKD groups. IPRE significantly increased the phospho-STAT3/STAT3 ratio in sham-operated but not in CKD animals in both sexes. There were no significant differences in phospho-AKT/AKT and phospho-ERK1,2/ERK1,2 ratios between the groups.ConclusionThe infarct size-limiting effect of IPRE was preserved in both sexes in CKD despite the more severe uremic cardiomyopathy in male CKD rats. Further research is needed to identify crucial molecular mechanisms in the cardioprotective effect of IPRE in CKD.

Lovastatin interferes with the infarct size-limiting effect of ischemic preconditioning and postconditioning in rat hearts

Statins have been shown to be cardioprotective; however, their interaction with endogenous cardioprotection by ischemic preconditioning and postconditioning is not known. In the present study, we examined if acute and chronic administration of the 3-hydroxy-3-methylglutaryl CoA reductase inhibitor lovastatin affected the infarct size-limiting effect of ischemic preconditioning and postconditioning in rat hearts. Wistar rats were randomly assigned to the following three groups: 1) vehicle (1% methylcellulose per os for 12 days), 2) chronic lovastatin (15 mg.kg(-1).day(-1) per os for 12 days), and 3) acute lovastatin (1% methylcellulose per os for 12 days and 50 micromol/l lovastatin in the perfusate). Hearts isolated from the three groups were either subjected to a nonconditioning (aerobic perfusion followed by 30-min coronary occlusion and 120-min reperfusion, i.e., test ischemia-reperfusion), preconditioning (three intermittent periods of 5-min ischemia-reperfusion cycles before test ischemia-reperfusion), or postconditioning (six cycles of 10-s ischemia-reperfusion after test ischemia) perfusion protocol. Preconditioning and postconditioning significantly decreased infarct size in vehicle-treated hearts. However, preconditioning failed to decrease infarct size in acute lovastatin-treated hearts, but the effect of postconditioning remained unchanged. Chronic lovastatin treatment abolished postconditioning but not preconditioning; however, it decreased infarct size in the nonconditioned group. Myocardial levels of coenzyme Q9 were decreased in both acute and chronic lovastatin-treated rats. Western blot analysis revealed that both acute and chronic lovastatin treatment attenuated the phoshorylation of Akt; however, acute but not chronic lovastatin treatment increased the phosphorylation of p42 MAPK/ERK. We conclude that, although lovastatin may lead to cardioprotection, it interferes with the mechanisms of cardiac adaptation to ischemic stress.

Lovastatin interferes with the infarct size-limiting effect of ischemic preconditioning and postconditioning

Lovastatin interferes with the infarct size-limiting effect of ischemic preconditioning and postconditioning

A new paradigm: cross talk of protein kinases during reperfusion saves life!

timely reperfusion after myocardial ischemia is the prerequisite to reduce irreversible tissue injury ([16][1]). However, reperfusion itself after a prolonged ischemic period may contribute to tissue damage, including reversible contractile dysfunction of viable cardiomyocytes ([4][2], [14][3]) as

Dilazep dihydrochloride prolongs the infarct size-limiting effect of ischemic preconditioning in rabbits.

Recent studies have indicated the key role of adenosine receptor activation as a trigger for ischemic preconditioning (PC). Hence, the augmentation of endogenous adenosine may potentiate the cardioprotective effects of PC. In this study. we aimed to test the hypothesis that dilazep dihydrochloride, an adenosine transport inhibitor, potentiates the PC effect. Protocol 1: Infarcts were produced in open-chest anesthetized rabbits by 30-min occlusion of a coronary artery and 2 days' reperfusion. PC was elicited by a preceding 5-min occlusion and either 5, 40, or 120 min of reperfusion. PC with the 5-min reperfusion markedly limited the infarct size after the 30-min ischemia (infarct size to area at risk (IS): 10%+/-3% vs 41%+/-3%, P < 0.05). PC was not protective when the reperfusion periods were 40 or 120 min (IS: 47%+/-5% and 44%+/-3%. P = not significant (NS) vs control, respectively). However, concomitant treatment with dilazep (0.2mg/kg) preserved the PC effect in the 40-min reperfusion group (18%+/-5%, P < 0.05 vs control) but not in the 120-min reperfusion group (43%+/-4%, P = NS vs control). Protocol 2: Infarct was produced in a similar rabbit model by either a 45- or 50-min occlusion of a coronary artery and 2 days of reperfusion. PC was elicited by a preceding 5-min occlusion and a 5-min reperfusion. PC was protective in the 45-min occlusion group (30%+/-7% vs 67%+/-3%, P < 0.05) but not in the 50-min occlusion group (74%+/-4% vs 79%+/-5%, P = NS). Treatment with dilazep (0.2mg/kg) failed to retrieve protection in this preconditioned group (77%+/-6%, P = NS vs 50-min occlusion group without PC). Thus, dilazep prolonged the infarct size-limiting effect of PC, but failed to retrieve protection in the group with a longer sustained ischemia.

Protein tyrosine kinase is not involved in the infarct size-limiting effect of ischemic preconditioning in canine hearts.

Protein kinase C (PKC) plays an important role in ischemic preconditioning (IP). Because (1) tyrosine kinase is located at the downstream of PKC for IP in the rabbit hearts and (2) we have reported that ecto-5'-nucleotidase is the substrate for PKC and plays a crucial role for the infarct size-limiting effect, we tested whether tyrosine kinase activation contributes to either activation of ecto-5'-nucleotidase or the infarct size-limiting effect of the early phase of IP in the canine heart. In dogs, the IP procedure (4 cycles of 5-minute occlusion of coronary artery) and exposure to 12, 13-phorbol myristate acetate (PMA) each activated myocardial ecto-5'-nucleotidase and Lck tyrosine kinase. Genistein (10, 30, and 100 microg. kg(-)(1). min(-)(1) IC), an inhibitor of tyrosine kinase, attenuated the activation of Lck tyrosine kinase but did not attenuate the activation of ecto-5'-nucleotidase due to either IP or PMA. In the other canine hearts, IP attenuated infarct size (49+/-5 versus 11+/-3 or 16+/-3%, P<0.01) due to 90 minutes of coronary occlusion followed by 6 hours of reperfusion, which was not blunted by 3 or 2 (30 and 100 microg. kg(-)(1). min(-)(1)) doses of genistein (infarct sizes, 15+/-4, 13+/-4, and 13+/-3%, respectively, and 17+/-3 and 15+/-4%, respectively) or lavendustin A. Tyrosine kinase does not activate ecto-5'-nucleotidase or trigger the infarct size-limiting effect of the early phase of IP in canine hearts.

Adenosine and cardioprotection in the diseased heart.

Biological and mechanical stressors such as ischemia, hypoxia, cellular ATP depletion, Ca2+ overload, free radicals, pressure and volume overload, catecholamines, cytokines, and renin-angiotensin may independently cause reversible and/or irreversible cardiac dysfunction. As a defense against these forms of stress, several endogenous self-protective mechanisms are exerted to avoid cellular injury. Adenosine, a degradative substance of ATP, may act as an endogenous cardioprotective substance in pathophysiological conditions of the heart, such as myocardial ischemia and chronic heart failure. For example, when brief periods of myocardial ischemia precede sustained ischemia, infarct size is markedly limited, a phenomenon known as ischemic preconditioning. We found that ischemic preconditioning activates the enzyme responsible for adenosine release, ie, ecto-5'-nucleotidase. Furthermore, the inhibitor of ecto-5'-nucleotidase reduced the infarct size-limiting effect of ischemic preconditioning, which establishes the cause-effect relationship between activation of ecto-5'-nucleotidase and the infarct size-limiting effect. We also found that protein kinase C is responsible for the activation of ecto-5'-nucleotidase. Protein kinase C phosphorylated the serine and threonine residues of ecto-5'-nucleotidase. Therefore, we suggest that adenosine produced via ecto-5'-nucleotidase gives cardioprotection against ischemia and reperfusion injury. Also, we found that plasma adenosine levels are increased in patients with chronic heart failure. Ecto-5'-nucleotidase activity increased in the blood and the myocardium in patients with chronic heart failure, which may explain the increases in adenosine levels in the plasma and the myocardium. In addition, we found that further elevation of plasma adenosine levels due to either dipyridamole or dilazep reduces the severity of chronic heart failure. Thus, we suggest that endogenous adenosine is also beneficial in chronic heart failure. We propose potential mechanisms for cardioprotection attributable to adenosine in pathophysiological states in heart diseases. The establishment of adenosine therapy may be useful for the treatment of either ischemic heart diseases or chronic heart failure.

Chronic treatment with an ACE inhibitor, temocapril, lowers the threshold for the infarct size-limiting effect of ischemic preconditioning.

This study examined whether chronic inhibition of the angiotensin-converting enzyme (ACE) lowers the threshold of preconditioning (PC) and potentiates cardio-protection of subthreshold PC. Rabbits were orally administered either 0.5 mg/kg/day temocapril or placebo for 14 days and were randomly subjected to subthreshold PC (i.e., PC with 2 minutes ischemia/5 minutes reperfusion) or no PC before a 30-minute coronary artery occlusion and a 3-hour reperfusion. The size of the infarct was determined by tetrazolium staining and was expressed as the percent of the area at risk (%IS/AR). At the end of the experiment, the lungs were sampled for a tissue ACE activity assay. There was no significant difference in %IS/AR among the rabbits given placebo alone, placebo plus 2 minutes PC, and temocapril alone (%IS/AR = 59.5 +/- 5.8%, 43.6 +/- 3.7%, and 56.7 +/- 7.1%, respectively). However, %IS/AR was significantly reduced in the group that received temocapril and 2 minutes PC before infarction (%IS/AR = 35.4 +/- 4.8%, P < 0.05 vs. placebo control and temocapril control). The lung tissue ACE activity did not differ in the placebo-treated rabbits with and without PC (12.6 +/- 2.8 vs. 13.7 +/- 2.0 nmol/g protein/min) but was suppressed in temocapril-treated rabbits despite 2 minutes PC (0.9 +/- 0.1 vs. 1.5 +/- 0.2 nmol/g protein/min, both P < 0.05 vs. placebo-treated groups). The present results suggest that chronic inhibition of ACE is beneficial for potentiating the anti-infarct effect of subthreshold PC.

It is time to ask what adenosine can do for cardioprotection.

Prevention and attenuation of ischemia and reperfusion injury in patients with acute coronary syndrome are critically important for cardiologists. To save these patients from deleterious ischemic insults, there are three different strategies. The first strategy is to increase ischemic tolerance before the onset of myocardial ischemia; the second is to attenuate the ischemia and reperfusion injury when an irreversible process of myocardial cellular injury occurs; the third is to treat the ischemic chronic heart failure that is caused by acute myocardial infarction. Adenosine, which is known to be cardioprotective against ischemia and reperfusion injury, may merit being used for these three cardioprotection strategies. First of all, adenosine induces collateral circulation via induction of growth factors, and triggers ischemic preconditioning, both of which induce ischemic tolerance in advance. Secondly, endogenous adenosine may mediate the infarct size-limiting effect of ischemic preconditioning, and exogenous adenosine is known to attenuate ischemia and reperfusion injury. Thirdly, we also revealed that adenosine metabolism is changed in patients with chronic heart failure, and increases in adenosine levels may attenuate the severity of ischemic heart failure. Therefore, adenosine therapy may improve the pathophysiology of ischemic chronic heart failure. Taking these factors together, we hereby propose potential tools for cardioprotection attributable to adenosine in ischemic hearts, and we postulate the use of adenosine therapy before, during, and after the onset of acute myocardial infarction.

Role of Protein Kinase C-α in Activation of Ecto-5′-nucleotidase in the Preconditioned Canine Myocardium

We have reported that activation of protein kinase C (PKC) increases ecto-5′-nucleotidase activity, which may contribute to the infarct size-limiting effect of ischemic preconditioning. Since we have reported that Ca2+- and phospholipid-sensitive PKC is activated due to ischemic preconditioning, we further tested 1) whether PKC-α or -β is translocated to the cellular membrane of the preconditioned canine myocardium, and 2) whether activation of PKC contributes to the increase in ecto-5′-nucleotidase activity via phosphorylation-dependent mechanisms. Four times of 5 minutes coronary occlusion separated by 5 minutes of reperfusion (ischemic preconditioning) translocated PKC-α to the cellular membrane in the canine hearts, although PKC-β, -δ, -ϵ, and -ζ were not translocated. The activity of Ca2+- and phospholipid-sensitive PKC increased, which was attenuated by the removal of either Ca2+or phosphatidylserine. Ecto-5′-nucleotidase was also activated in the preconditioned myocardium compared with control. Inhibition of PKC due to GF109203X blunted the activation of myocardial ecto-5′-nucleotidase. Okadaic acid (an inhibitor of phosphatase) enhanced the increases in ecto-5′-nucleotidase activity due to preconditioning, and this enhancement was blunted by GF109203X. We conclude that ischemic preconditioning activates PKC-α, and thus ecto-5′-nucleotidase.

Selective Blockade of AT1Angiotensin II Receptors Abolishes Ischemic Preconditioning in Isolated Rabbit Hearts

The aim of the present study was to assess the participation of angiotensin II receptors in the triggering mechanism of ischemic preconditioning. Isolated buffer-perfused rabbit hearts were subjected to 40 min of regional ischemia (37°C) followed by 60 min of reperfusion. Ischemic preconditioning was induced with three cycles of 5-min ischemia and 10-min reperfusion given prior to the 40-min ischemic period. Infarct size and ventricular function were assessed. Ischemic preconditioning reduced infarct size to 5.2±1.2% of the area at risk (mean±s.e.m.,P<0.001) when compared to controls (26.4±3.0%), but did not protect against ventricular dysfunction. Activation of angiotensin II receptors with angiotensin II (100 nm) also limited infarct size (9.6±2.2%,P<0.01vcontrol group). Inhibition of angiotensin II receptors with [Sar1, Val5, Ala8]-angiotensin II (saralasin, 1μm) blocked the protection of ischemic preconditioning against necrosis (29.7±3.2%) while it did not increase infarct size in saralasin-treated control hearts (31.5±3.9%). Furthermore, inhibition of the AT1subtype of the angiotensin II receptor with losartan (20μm), but not inhibition of the AT2subtype with PD-123,319 ditrifluoroacetate (10μm), abolished the infarct size-limiting effect of ischemic preconditioning. We conclude that the AT1angiotensin II receptor participates in ischemic preconditioning. Thus, in the isolated rabbit heart, activation of AT1receptors must occur before prolonged ischemia for ischemic preconditioning to limit infarction.

Role of activation of ectosolic 5'-nucleotidase in the cardioprotection mediated by opening of K+c channels.

We tested the hypothesis that the opening of ATP-sensitive K+ channels contributes to activation of ectosolic 5'-nucleotidase and the infarct size-limiting effect of ischemic preconditioning. In open-chest dogs, the left anterior descending coronary artery was occluded four times for 5 min each, separated by a 5-min period of reperfusion (ischemic preconditioning, n = 8). After this procedure, the coronary artery was occluded for 90 min, followed by 6 h of reperfusion. Infarct size was smaller in this group than in the group (control, n = 8) with a 45 min interval instead of the ischemic preconditioning procedure (40.1 +/- 3.9 vs. 6.4 +/- 1.9%). Glibenclamide blunted the infarct size-limiting effect of ischemic preconditioning (infarct size, 37.3 +/- 5.8%; n = 7), and transient exposures to cromakalim and nicorandil mimicked it [infarct size, 10.1 +/- 3.1 (n = 7) and 11.1 +/- 2.7% (n = 8), respectively]. Ectosolic and cytosolic 5'-nucleotidase activity increased in the ischemic preconditioning group compared with that in the control group; this preconditioning-induced increase in 5'-nucleotidase activity was blunted by glibenclamide (n = 5) and mimicked by cromakalim (n = 5) and nicorandil (n = 5). The infarct size-limiting effect due to cromakalim and nicorandil was blunted by alpha,beta-methyleneadenosine 5'-diphosphate, an inhibitor of ectosolic 5'-nucleotidase [infarct size, 37.7 +/- 5.6 (n = 9) and 36.8 +/- 4.8% (n = 7), respectively] and 8-sulfophenyltheophylline (infarct size with cromakalim, 44.7 +/- 4.6%; n = 7). We conclude that activation of ectosolic 5'-nucleotidase due to the openers of ATP-sensitive K+ channels contributes to the infarct size- limiting effect of ischemic preconditioning.

Activation of ecto-5'-nucleotidase and cardioprotection by ischemic preconditioning.

Since Jennings and his colleagues found that preceding brief periods of myocardial ischemia limit infarct size following sustained ischemia and reperfusion (11), termed as “ischemic preconditioning”, many basic and clinical researchers have focused on the cellular mechanisms of this potent cardioprotection. Downey and his colleagues (10) observed that 8-sulfophenyltheophylline blunts the infarct size-limiting effect of ischemic preconditioning in the rabbit heart. This evidence may point to two hypotheses: 1) adenosine makes the myocardium resistant against the lethal ischemic injury (adenosine as a trigger substance), and 2) ischemic preconditioning makes adenosine act more effectively during the lethal ischemic insult (adenosine as a mediator substance). Since adenosine is known to have many different beneficial cardiovascular effects against ischemia and reperfusion injury (1, 3), we hypothesized the latter mechanism as a candidate of the cellular mechanism of ischemic preconditioning.

Effects of ischemic and adenosine preconditioning on interstitial fluid adenosine and myocardial infarct size.

The accumulation of adenosine during a brief coronary occlusion has been proposed to mediate the infarct size-limiting effect of ischemic preconditioning. The purpose of this study was to compare the effects of ischemic preconditioning and a transient adenosine infusion on myocardial interstitial fluid (ISF) adenosine levels and infarct size. Microdialysis fibers (10.0 mm length) were placed in the left ventricular myocardium of pentobarbital sodium-anesthetized rabbits to estimate ISF adenosine. Ischemic preconditioning was induced by 5 min of coronary artery occlusion and 10 min of reperfusion before 45 min of occlusion. Adenosine preconditioning was induced with 5 min of intravenous adenosine infusion (140 micrograms.kg-1.min-1) followed by a 10-min washout before the prolonged occlusion. Myocardial infarct size was determined by triphenyltetrazolium chloride staining after 3 h of reperfusion. Five minutes of ischemia and 5 min of adenosine infusion produced comparable increases in dialysate adenosine levels (from 0.19 +/- 0.02 to 0.69 +/0- 0.11 and 0.28 +/- 0.10 to 0.71 +/- 0.18 microM, respectively) that decreased to baseline before the prolonged ischemia; however, ischemic-preconditioned hearts exhibited elevated dialysate adenosine levels for the first 5 min of reperfusion. Ischemic-preconditioned hearts exhibited significantly reduced dialysate adenosine concentrations for the first 20 min of the prolonged occlusion (P < 0.05 vs. control), and infarct size was reduced from 41 +/- 6 to 10 +/- 4% of risk area. Adenosine preconditioning had no effect on dialysate adenosine levels during prolonged ischemia but did reduce infarct size to 25 +/- 5% of risk area. These results indicate that a transient increase in ISF adenosine can reduce myocardial infarct size, but adenosine alone does not fully replicate the protective effects of ischemic preconditioning.

Nucleoside transport inhibitors enhance the infarct size-limiting effect of ischemic preconditioning.

We recently observed that dipyridamole pretreatment significantly enhanced the infarct size (IS)-limiting effect of preconditioning (PC), which was attenuated by adenosine receptor antagonist. This potentiation of PC was interpreted to result from inhibition of nucleoside transport by dipyridamole, but contribution of other pharmacologic actions of dipyridamole could not be excluded. To confirm that inhibition of nucleoside transport leads to PC enhancement, we assessed alteration of mild PC by two different nucleoside transport inhibitors, dilazep and R75231, which, unlike dipyridamole, lack action on phosphodiesterase (PDE) and prostacyclin. Myocardial infarction was induced in rabbits by 30-min coronary occlusion and 72-h reperfusion. IS and area at risk (AAR) were determined by histology and fluorescent particles, respectively. Rabbits either were untreated or received dilazep (0.34 mg/kg intravenously, i.v.) or R75231 (0.05 mg/kg i.v.) before coronary occlusion. In other groups of rabbits, PC was conducted with 2-min ischemia and 5-min reperfusion with or without injection of the nucleoside transport inhibitor (0.34 or 0.10 mg/kg dilazep or 0.05 mg/kg of R75231) before PC. IS expressed as percentage of AAR (%IS/AAR) was 43.9 +/- 2.3% (SE) in untreated controls; dilazep (0.34 mg/kg) and R75231 alone did not modify IS (%IS/AAR = 50.6 +/- 4.7 and 42.7 +/- 11.9%, respectively). PC tended to reduce IS (%IS/AAR = 33.3 +/- 3.5%), but the combination of dilazep or R75231 with PC significantly limited %IS/AAR (%IS/AAR = 22.5 +/- 5.0% after low-dose dilazep plus PC, 27.6 +/- 4.9% after high-dose dilazep plus PC, and 19.9 +/- 3.6%, after R75231 plus PC).(ABSTRACT TRUNCATED AT 250 WORDS)

Infarct size-limiting effect of ischemic preconditioning is blunted by inhibition of 5'-nucleotidase activity and attenuation of adenosine release.

We have previously reported that ischemic preconditioning increases 5'-nucleotidase activity and adenosine release during ischemia and reperfusion. However, its direct cause-and-effect relation has not been proven. To test the idea that the infarct size-limiting effect of ischemic preconditioning is blunted by inhibition of ectosolic 5'-nucleotidase activity, we assessed 5'-nucleotidase activity, adenosine release, and infarct size caused by sustained ischemia with and without an exposure to alpha,beta,-methylene adenosine 5'-diphosphate (AOPCP) in the ischemia-preconditioned myocardium. In 67 open-chest dogs, the left anterior descending coronary artery was cannulated and perfused with an extracorporeal bypass tube from the carotid artery. After hemodynamic stabilization, the coronary artery was occluded four times for 5 minutes separated by 5 minutes of reperfusion (ischemic preconditioning, n = 10). After this procedure, the coronary artery was occluded for 90 minutes followed by 6 hours of reperfusion. Infarct size normalized by the risk area was smaller than the control group (n = 8, 41.0 +/- 2.6% versus 6.8 +/- 1.9%), although there were no significant differences in the endomyocardial collateral flow measured at 80 minutes of ischemia (8.5 +/- 1.1 versus 9.4 +/- 1.0 mL/100 g per minute). Ectosolic and cytosolic 5'-nucleotidase activity and adenosine release were increased during reperfusion in the ischemic preconditioning group compared with the control group, and the activity of ectosolic 5'-nucleotidase was markedly reduced by AOPCP (n = 10). AOPCP affected neither adenosine-induced coronary vasodilation nor increases in myocardial oxygen consumption during an intracoronary infusion of isoproterenol (n = 10). To test whether the increase in 5'-nucleotidase activity decreases infarct size, we infused AOPCP 10 minutes before the ischemic preconditioning procedure and continued for 60 minutes after the onset of reperfusion (n = 8). AOPCP blunted the infarct size-limiting effect (infarct size, 38.8 +/- 4.9%). AOPCP without ischemic preconditioning did not increase infarct size (n = 9). Furthermore, when AOPCP was infused during the ischemic preconditioning procedure (n = 6) or during 60 minutes of reperfusion (n = 6), the infarct size-limiting effect was partially blunted (infarct size, 21.3 +/- 2.5% and 19.5 +/- 2.4%, respectively). Increases in ectosolic 5'-nucleotidase activity and adenosine release are primarily responsible for the infarct size-limiting effect of ischemic preconditioning. Exposures to adenosine during the ischemic preconditioning procedure and enhanced release of adenosine during reperfusion synergistically contribute to the infarct size-limiting effects.

Myocardial infarct size-limiting effect of ischemic preconditioning was not attenuated by oxygen free-radical scavengers in the rabbit.

The limiting effect of ischemic preconditioning on infarct size has been reported in canine hearts, which contain considerable amounts of xanthine oxidase, a free radical-producing enzyme. Furthermore, a recent study suggested that free radicals generated during preconditioning may contribute to the cardioprotective effect of preconditioning. The present study examined 1) whether preconditioning limits infarct size in rabbits, which, like humans, lack myocardial xanthine oxidase and 2) whether the cardioprotective effect of PC is mediated by free radicals. A branch of the circumflex coronary artery in rabbits was occluded for 30 minutes and then reperfused for 72 hours. Myocardial infarct size and area at risk were determined by histology and fluorescent particles, respectively. Five groups were studied: an untreated control group, a preconditioned group (PC group), a high-dose superoxide dismutase (SOD)-treated preconditioned group (high-dose SOD-PC group), a low-dose SOD-treated preconditioned group (low-dose SOD-PC group), and a SOD-plus-catalase-treated preconditioned group (SOD/CAT-PC group). Preconditioning was performed with four episodes of 5 minutes of ischemia and 5 minutes of reperfusion. The free radical scavengers (30,000 units/kg SOD for high-dose SOD-PC group, 15,000 units/kg SOD for low-dose SOD-PC group, and 30,000 units/kg SOD plus 55,000 units/kg catalase for SOD/CAT-PC group) were infused intravenously over 60 minutes starting 20 minutes before preconditioning. Infarct size as the percentage of area at risk was 45.1 +/- 3.5% (mean +/- SEM) in the control group (n = 11), 13.3 +/- 3.0% in the PC group (n = 12), 9.7 +/- 1.8% in the high-dose SOD-PC group (n = 8), 11.9 +/- 2.2% in the low-dose SOD-PC group (n = 6), and 9.6 +/- 2.3% in the SOD/CAT-PC group (n = 6) (p less than 0.05 versus control for the last four values). The differences in infarct size as the percent of area at risk among the PC, high-dose SOD-PC, low-dose SOD-PC, and SOD/CAT-PC groups were not significant. Ischemic preconditioning delays ischemic myocardial necrosis regardless of myocardial xanthine oxidase content. Free radicals are unlikely to have a major role in the mechanism of the preconditioning in rabbits.