Preconditioning by 17beta-estradiol in isolated rat heart depends on PI3-K/PKB pathway, PKC, and ROS

To study the cell signaling events leading to 17beta-estradiol (E(2))-induced acute cardioprotection, we subjected isolated rat hearts to three 5-min cycles of 10 microM E(2) before 30 min of regional ischemia, followed by 2 h of reperfusion. Protection was judged by changes in infarct size in percentage of risk zone volume. To test the importance of phosphoinositide 3-kinase (PI3-K), protein kinase C (PKC), or reactive oxygen species (ROS) in E(2)-induced protection, we combined wortmannin (1 microM), chelerythrine (2 microM), and 2-mercaptopropionylglycine (300 microM), respectively, with E(2) exposure. Changes in phosphorylation of protein kinase B (PKB) and selected PKC isoforms were tested by immunoblotting of total lysates and subcellular fractions, along with assessment of PKC translocation from soluble to membrane fraction of heart tissue homogenates. Intracellular ROS levels induced by E(2) preconditioning were investigated. E(2) preconditioning led to significant reduction in infarct size from 31.8 +/- 5.3 to 20.2 +/- 2.6% in male hearts and from 42.7 +/- 4.7 to 17.1 +/- 3.4% in female hearts (P < 0.05). Protection was abolished by wortmannin (30.0 +/- 3.2%), chelerythrine (45.1 +/- 4.4%), and 2-mercaptopropionylglycine (36.8 +/- 4.7%). E(2) preconditioning induced phosphorylation of PKB, PKCalpha, and PKCepsilon and membrane translocation of PKCepsilon and PKCdelta. Intracellular ROS levels were found elevated after transient treatment with hormone. Therefore, our data demonstrate the ability of E(2) to induce preconditioning-like cardioprotection via cell signaling events shared by classic ischemic preconditioning.     

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 相似文献   

Cardioprotective effect of resveratrol via HO-1 expression involves p38 map kinase and PI-3-kinase signaling, but does not involve NFkappaB

Recent studies have demonstrated that resveratrol (trans-3,4',5-trihydroxy stilbene), a phytoalexin found in the skin and seeds of grapes, can pharmacologically precondition (PC) the heart through a nitric oxide (NO)-dependent and adenosine receptors-mediated mechanism. Since NO can induce the expression of heme oxygenase-1 (HO-1), we examined if HO-1 induction has a direct role in resveratrol-preconditioning of the heart. Eight groups of rats were studied during 7 days: (i) control rats; (ii) rats receiving resveratrol (gavage, 2.5 mg/kg); (iii) rats injected tin protoporphyrin (SnPP), a HO-1 inhibitor, i.p. on days 1, 3 and 6; (iv) rats injected 202190 (SB), a p38MAPK inhibitor, i.p. for 7 days; (v) rats injected 294002 (LY), a Akt inhibitor, i.p. for 7days; (vi) rats receiving resveratrol and SnPP; (vii) rats receiving resveratrol and SB; and (viii) rats receiving resveratrol and LY. After the treatments, the rats were sacrificed, and the hearts isolated and subjected to 30 min global ischemia followed by 2 h of reperfusion. The results shown a significant cardioprotection with resveratrol as evidenced by superior post-ischemic ventricular recovery, reduced myocardial infarct size, and decreased number of apoptotic cardiomyocytes. SnPP treatment abolished the cardioprotective effect of resveratrol. Resveratrol induced the activation of nuclear factor kappa-beta(NFkappaB), the phosphorylation of p38MAP kinase beta and Akt, as well as the inhibition of p38 MAP kinase alpha; all these effects but the activation of NFkappaB, were completely reversed by treatment with SnPP. These results indicate that resveratrol generates cardioprotection by preconditioning the heart by HO-1-mediated mechanisms, which are regulated by p38MAP kinase and Akt survival signaling, but non-dependent on NFkappaB activation.     

HDAC Inhibition Elicits Myocardial Protective Effect through Modulation of MKK3/Akt-1

We and others have demonstrated that HDAC inhibition protects the heart against myocardial injury. It is known that Akt-1 and MAP kinase play an essential role in modulation of myocardial protection and cardiac preconditioning. Our recent observations have shown that Akt-1 was activated in post-myocardial infarction following HDAC inhibition. However, it remains unknown whether MKK3 and Akt-1 are involved in HDAC inhibition-induced myocardial protection in acute myocardial ischemia and reperfusion injury. We sought to investigate whether the genetic disruption of Akt-1 and MKK3 eliminate cardioprotection elicited by HDAC inhibition and whether Akt-1 is associated with MKK3 to ultimately achieve protective effects. Adult wild type and MKK3^−/−, Akt-1^−/− mice received intraperitoneal injections of trichostatin A (0.1mg/kg), a potent inhibitor of HDACs. The hearts were subjected to 30 min myocardial ischemia/30 min reperfusion in the Langendorff perfused heart after twenty four hours to elicit pharmacologic preconditioning. Left ventricular function was measured, and infarct size was determined. Acetylation and phosphorylation of MKK3 were detected and disruption of Akt-1 abolished both acetylation and phosphorylation of MKK3. HDAC inhibition produces an improvement in left ventricular functional recovery, but these effects were abrogated by disruption of either Akt-1 or MKK3. Disruption of Akt-1 or MKK3 abolished the effects of HDAC inhibition-induced reduction of infarct size. Trichostatin A treatment resulted in an increase in MKK3 phosphorylation or acetylation in myocardium. Taken together, these results indicate that stimulation of the MKK3 and Akt-1 pathway is a novel approach to HDAC inhibition -induced cardioprotection.     

Autophagy in myocardium of murine hearts subjected to ischemia followed by reperfusion

Autophagy in myocardium has been thought to be cardioprotective, but its extent after transient or prolonged myocardial ischemia remains unclear. Accordingly, we characterized its magnitude in myocardium of murine hearts subjected to ischemia with or without reperfusion. Ten-week-old transgenic GFP-LC3 mice and C57Bl6 mice were subjected to coronary ligation for 1 or 4 h followed by 24 h of reperfusion (1HTL, 4HTL) or to 24 h of persistent ligation (24HPL). Their hearts were analyzed by fluorescence microscopy, electron microscopy, and by Western blotting. Fluorescent GFP-LC3 dots indicative of autophagy were absent in infarct zones and reduced markedly in the peri-infarct zones compared with dots in sham controls (p ≤ 0.05). The LC3-II/LC3-I ratio indicative of autophagy did not increase in LV homogenates from hearts following ischemia. Phosphorylation of ribosomal protein S6 increased in LV homogenates in hearts from mice subjected to 4HTL and 24HPL (p ≤ 0.05). Virtually no autophagic cells recognizable by electron microscopy were evident in infarct or peri-infarct zones. Autophagy is virtually absent within 24 h in the center of zones of infarction and is decreased significantly in the peri-infarct zones compared with that in normal hearts.     

Role of 12-lipoxygenase in volatile anesthetic-induced delayed preconditioning in mice

Delayed cardiac protection mediated by 12-lipoxygenase (12-LO) expression and activity has been linked to opioid receptor stimulation. The role of 12-LO in volatile anesthetic-induced delayed cardiac protection has not been determined. We tested the hypothesis that expression and activity of 12-LO mediate delayed cardiac protection induced by isoflurane in the mouse heart in vivo. Mice were pretreated with 1.4% isoflurane for 30 min and allowed to recover for 1, 12, or 24 h. Immunoblot analysis showed isoflurane significantly enhanced 12-LO protein expression at 12 and 24 h after isoflurane exposure, and this induction of 12-LO was confirmed by immunohistochemistry of whole heart sections at 24 h. The induced protein expression appeared to be localized to intercalated disc regions adjoining adjacent cardiac myocytes. Additionally, mice +/- isoflurane (24 h previously) were subjected to 30 min coronary artery occlusion followed by 2 h of reperfusion in the presence and absence of a 12-LO inhibitor. Isoflurane reduced infarct size (27.1 +/- 2.2% of the area at risk; n = 8) compared with the control group (44.6 +/- 3.6%, n = 8). Baicalein (3 mg/kg), a selective 12-LO inhibitor, blocked the delayed protective effects of isoflurane pretreatment on infarct size (40.6 +/- 3.6%, n = 8). These data suggest that 12-LO is an important mediator of isoflurane-induced delayed preconditioning.     

CRYAB and HSPB2 deficiency alters cardiac metabolism and paradoxically confers protection against myocardial ischemia in aging mice

The abundantly expressed small molecular weight proteins, CRYAB and HSPB2, have been implicated in cardioprotection ex vivo. However, the biological roles of CRYAB/HSPB2 coexpression for either ischemic preconditioning and/or protection in situ remain poorly defined. Wild-type (WT) and age-matched ( approximately 5-9 mo) CRYAB/HSPB2 double knockout (DKO) mice were subjected either to 30 min of coronary occlusion and 24 h of reperfusion in situ or preconditioned with a 4-min coronary occlusion/4-min reperfusion x 6, before similar ischemic challenge (ischemic preconditioning). Additionally, WT and DKO mice were subjected to 30 min of global ischemia in isolated hearts ex vivo. All experimental groups were assessed for area at risk and infarct size. Mitochondrial respiration was analyzed in isolated permeabilized cardiac skinned fibers. As a result, DKO mice modestly altered heat shock protein expression. Surprisingly, infarct size in situ was reduced by 35% in hearts of DKO compared with WT mice (38.8 +/- 17.9 vs. 59.8 +/- 10.6% area at risk, P < 0.05). In DKO mice, ischemic preconditioning was additive to its infarct-sparing phenotype. Similarly, infarct size after ischemia and reperfusion ex vivo was decreased and the production of superoxide and creatine kinase release was decreased in DKO compared with WT mice (P < 0.05). In permeabilized fibers, ADP-stimulated respiration rates were modestly reduced and calcium-dependent ATP synthesis was abrogated in DKO compared with WT mice. In conclusion, contrary to expectation, our findings demonstrate that CRYAB and HSPB2 deficiency induces profound adaptations that are related to 1) a reduction in calcium-dependent metabolism/respiration, including ATP production, and 2) decreased superoxide production during reperfusion. We discuss the implications of these disparate results in the context of phenotypic responses reported for CRYAB/HSPB2-deficient mice to different ischemic challenges.     

Exercise enhances myocardial ischemic tolerance via an opioid receptor-dependent mechanism

Exercise increases serum opioid levels and improves cardiovascular health. Here we tested the hypothesis that opioids contribute to the acute cardioprotective effects of exercise using a rat model of exercise-induced cardioprotection. For the standard protocol, rats were randomized to 4 days of treadmill training and 1 day of vigorous exercise (day 5), or to a sham exercise control group. On day 6, animals were killed, and global myocardial ischemic tolerance was assessed on a modified Langendorff apparatus. Twenty minutes of ischemia followed by 3 h of reperfusion resulted in a mean infarct size of 42 +/- 4% in hearts from sham exercise controls and 21 +/- 3% (P < 0.001) in the exercised group. The cardioprotective effects of exercise were gone by 5 days after the final exercise period. To determine the role of opioid receptors in exercise-induced cardioprotection, rats were exercised according to the standard protocol; however, just before exercise on days 4 and 5, rats were injected subcutaneously with 10 mg/kg of the opioid receptor antagonist naltrexone. Similar injections were performed in the sham exercise control group. Naltrexone had no significant effect on baseline myocardial ischemic tolerance in controls (infarct size 43 +/- 4%). In contrast, naltrexone treatment completely blocked the cardioprotective effect of exercise (infarct size 40 +/- 5%). Exercise was also associated with an early increase in myocardial mRNA levels for several opioid system genes and with sustained changes in a number of genes that regulate inflammation and apoptosis. These findings demonstrate that the acute cardioprotective effects of exercise are mediated, at least in part, through opioid receptor-dependent mechanisms that may include changes in gene expression.     

一氧化氮和线粒体ATP敏感性钾通道介导血管紧张素转换酶抑制剂加强阈下预处理的作用

实验采用离体大鼠心脏Langendorff灌流模型,观察含巯基（卡托普利）和不含巯基（培哚普利拉）的两种血管紧张素转换酶抑制剂（angiotensin-converting enzyme inhibitors,ACEI）对抗心肌缺血的作用,并探讨一氧化氮（nitric oxide,NO）和线粒体ATP敏感性钾通道（mimchondrial ATP-sensitive potassium channel,mitoKATP channel）是否参与ACEI的心肌保护作用。结果表明：（1）给予大鼠心脏2min全心停灌和10min复灌作为闽下缺血预处理（subthreshold preconditioning,sPC）、卡托普利或培哚普利拉单独使用,均不能改善长时间缺血复灌（缺血30min＋复灌120min）引起的心肌损伤。（2）当两种ACEI分别和sPC联合使用时,与sPC组相比,缺血心脏在长时间缺血后的复灌期问左室舒张末压（left ventricular end-diastolic pressure,LVEDP）明显降低,左宦发展压（left ventricular developed pressure,LVDP）和冠脉流量明显增高,乳酸脱氢酶（lactate dehydrogenase,LDH）的释放量和心肌梗死面积明显低于sPC组。（3）利用NOS抑制剂L-NAME和mitoKATP通道的抑制剂5-HD灌流10min后,可明显抑制卡托普利／培哚普利拉和sPC联合使用引起的LVEDP降低,并使LVDP和冠脉流量降低,LDH的释放量和心肌梗死面积明显增高（P〈0．05）。（4）sPC、卡托普利或培哚普利拉单独使用,心脏NO的产生增加。ACEI和sPC联合使用,与三者单独使用相比NO的浓度亦明显增高（P〈0．05）。结果提示：含与不含巯基的ACEI与闽下缺血预处理联合使用均可使大鼠心脏功能明显改善,其心肌保护作用的机制可能通过NO途径,并和mitoKATP通道的激活有关。     

HSP70.1 and -70.3 are required for late-phase protection induced by ischemic preconditioning of mouse hearts

We investigated the role of inducible heat shock proteins 70.1 and 70.3 (HSP70.1 and HSP70.3, respectively) in myocardial ischemic preconditioning (IP) in mice. Wild-type (WT) mice and HSP70.1- and HSP70.3-null [HSP70.1/3(-/-)] mice were subjected to IP and examined 24 h later during the late phase of protection. IP significantly increased steady-state levels of HSP70.1 and HSP70.3 mRNA and expression of inducible HSP70 protein in WT myocardium. To assess protection against tissue injury, mice were subjected to 30 min of regional ischemia and 3 h of reperfusion. In WT mice, IP reduced infarct size by 43% compared with sham IP-treated mice. In contrast, IP did not reduce infarct size in HSP70.1/3(-/-) mice. Absence of inducible HSP70.1 and HSP70.3 had no effect, however, on classical or early-phase protection produced by IP, which significantly reduced infarct size in HSP70.1/3(-/-) mice. We conclude that inducible HSP70.1 and HSP70.3 are required for late-phase protection against infarction following IP in mice.     

PPAR-alpha activation as a preconditioning-like intervention in rats in vivo confers myocardial protection against acute ischaemia-reperfusion injury: involvement of PI3K-Akt

Peroxisome proliferator-activated receptors (PPAR) regulate the expression of genes involved in lipid metabolism, energy production, and inflammation. Their role in ischaemia-reperfusion (I/R) is less clear, although research indicates involvement of PPARs in some forms of preconditioning. This study aimed to explore the effects of PPAR-α activation on the I/R injury and potential cardioprotective downstream mechanisms involved. Langendorff-perfused hearts of rats pretreated with the selective PPAR-α agonist WY-14643 (WY, pirinixic acid; 3 mg·(kg body mass)·day(-1); 5 days) were subjected to 30 min ischaemia - 2 h reperfusion with or without the phosphatidylinositol 3-kinase (PI3K)-Akt inhibitor wortmannin for the evaluation of functional (left ventricular developed pressure, LVDP) recovery, infarct size (IS), and reperfusion-induced arrhythmias. A 2-fold increase in baseline PPAR-α mRNA levels (qPCR) in the WY-treated group and higher post-I/R PPAR-α levels compared with those in untreated controls were accompanied by similar changes in the expression of PPAR-α target genes PDK4 and mCPT-1, regulating glucose and fatty acid metabolism, and by enhanced Akt phosphorylation. Post-ischaemic LVDP restoration in WY-treated hearts reached 60% ± 9% of the pre-ischaemic values compared with 24% ± 3% in the control hearts (P < 0.05), coupled with reduced IS and incidence of ventricular fibrillation that was blunted by wortmannin. Results indicate that PPAR-α up-regulation may confer preconditioning-like protection via metabolic effects. Downstream mechanisms of PPAR-α-mediated cardioprotection may involve PI3K-Akt activation.     

Angiotensin preconditioning of the heart

Angiotensin II (Ang II) has been found to exert preconditioning-like effect on mammalian hearts. Diverse mechanisms are known to exist to explain the cardioprotective abilities of Ang II preconditioning. The present study hypothesized, based on the recent report that Ang II generates reactive oxygen species (ROS) through NADPH oxidase, that Ang II preconditioning occurs through redox cycling. To test this hypothesis, a group of rat hearts was treated with Ang II in the absence or presence of an NADPH oxidase inhibitor, apocynin; or a cell-permeable ROS scavenger, N-acetyl cysteine (NAC). Ang II pretreatment improved postischemic ventricular recovery; reduced myocardial infarction; and decreased the number of cardiomyocyte apoptosis, indicating its ability to precondition the heart against ischemic injury. Both apocynin and NAC almost abolished the preconditioning ability of Ang II. Ang II resulted in increase in ROS activity in the heart, which was reduced by either NAC or apocynin. Ang II also increased both the NADPH oxidase subunits gp91 phox and p22phox mRNA expression, which was abolished with apocynin and NAC. Our results thus demonstrate that the Ang II preconditioning was associated with enhanced ROS activities and increased NADPH oxidase subunits p22phox and gp91phox expression. Both NAC and apocynin reduced ROS activities simultaneously abolishing preconditioning ability of Ang II, suggesting that Ang II preconditioning occurs through redox cycling. That both NAC and apocynin reduced ROS activities and abolished Ang II-mediated increase in p22phox and gp91phox activity further suggest that such redox cycling occurs via both NADPH oxidase-dependent and -independent pathways.     

A Randomized Pilot Trial of Remote Ischemic Preconditioning in Heart Failure with Reduced Ejection Fraction

Background

Remote ischemic preconditioning (RIPC) induced by transient limb ischemia confers multi-organ protection and improves exercise performance in the setting of tissue hypoxia. We aimed to evaluate the effect of RIPC on exercise capacity in heart failure patients.

Methods

We performed a randomized crossover trial of RIPC (4×5-minutes limb ischemia) compared to sham control in heart failure patients undergoing exercise testing. Patients were randomly allocated to either RIPC or sham prior to exercise, then crossed over and completed the alternate intervention with repeat testing. The primary outcome was peak VO2, RIPC versus sham. A mechanistic substudy was performed using dialysate from study patient blood samples obtained after sham and RIPC. This dialysate was used to test for a protective effect of RIPC in a mouse heart Langendorff model of infarction. Mouse heart infarct size with RIPC or sham dialysate exposure was also compared with historical control data.

Results

Twenty patients completed the study. RIPC was not associated with improvements in peak VO2 (15.6+/−4.2 vs 15.3+/−4.6 mL/kg/min; p = 0.53, sham and RIPC, respectively). In our Langendorff sub-study, infarct size was similar between RIPC and sham dialysate groups from our study patients, but was smaller than expected compared to healthy controls (29.0%, 27.9% [sham, RIPC] vs 51.2% [controls]. We observed less preconditioning among the subgroup of patients with increased exercise performance following RIPC (p<0.04).

Conclusion

In this pilot study of RIPC in heart failure patients, RIPC was not associated with improvements in exercise capacity overall. However, the degree of effect of RIPC may be inversely related to the degree of baseline preconditioning. These data provide the basis for a larger randomized trial to test the potential benefits of RIPC in patients with heart failure.

Trial Registration

ClinicalTrials.gov +++++{"type":"clinical-trial","attrs":{"text":"NCT01128790","term_id":"NCT01128790"}}NCT01128790     

Exogenous NO triggers preconditioning via a cGMP- and mitoKATP-dependent mechanism

Exogenous nitric oxide (NO) triggers a preconditioning-like effect in heart via a pathway that is dependent on reactive oxygen species. This study examined the signaling pathway by which the NO donor S-nitroso-N-acetylpenicillamine (SNAP, 2 microM) triggers its anti-infarct effect. Isolated rabbit hearts experienced 30 min of regional ischemia and 120 min of subsequent reperfusion. Infarct size was determined by triphenyltetrazolium chloride staining. Infarct size was reduced from 30.5 +/- 3.0% of the risk zone in control hearts to 10.2 +/- 2.0% in SNAP-treated hearts. Bracketing the SNAP infusion with either the guanylyl cyclase blocker 1H-[1,2,4]oxadiazole[4,3-a]quinoxalin-1-one (2 microM) or the mitochondrial ATP-sensitive K(+) (mitoK(ATP)) channel blocker 5-hydroxydecanoate (200 microM) completely blocked the infarct-sparing effect of SNAP (34.3 +/- 3.8 and 32.2 +/- 1.6% infarction, respectively). Pretreatment of hearts with 8-(4-chlorophenylthio)-guanosine 3',5'-cyclic monophosphate (10 microM), which is a cell-permeable cGMP analog that activates protein kinase G, mimicked the preconditioning effect of SNAP by reducing infarct size to 7.5 +/- 1.1% of the risk zone. This salutary effect was abolished by either the free radical scavenger N-(2-mercaptopropionyl)glycine (1 mM) or 5-hydroxydecanoate (100 microM; 28.9 +/- 2.7 and 33.6 +/- 5.0% infarction of the risk zone, respectively). To confirm these functional data and the effect of SNAP on the guanylyl cyclase-protein kinase G signaling pathway, cGMP levels were measured. SNAP increased the level from 0.18 +/- 0.04 to 0.61 +/- 0.14 pmol/mg of protein (P < 0.05). These data suggest that exogenous NO triggers the preconditioning effect by initiating a cascade of events including stimulation of guanylyl cyclase to make cGMP, activation of protein kinase G, opening of mitoK(ATP) channels, and, finally, production of reactive oxygen species.     

Opioid receptor contributes to ischemic preconditioning through protein kinase C activation in rabbits

Recent studies have reported that protection from ischemic preconditioning (PC) is blocked by the opioid receptor antagonist naloxone (NAL). We tested whether an opioid agonist could mimic PC in the rabbit heart, whether that protection involved protein kinase C (PKC) activation, and whether opioid receptors act in concert with other PKC-coupled receptors. Rabbit hearts were subjected to 30min coronary occlusions and were reperfused for either 3 (in situ) or 2 (in vitro) h. Infarct size was determined by staining with triphenyltetrazolium chloride. In untreated in situ hearts 38.5 ± 1.6% of the risk zone infarcted. PC with 5 min ischemia/10 min reperfusion significantly limited infarction to 12.7 ± 2.9% (p < 0.01). NAL infusion did not modify infarction (39.6 ± 1.6%) in non-PC hearts, but blocked the effect of one cycle of PC (34.4 ± 3.6% infarction). NAL, however, could not block cardioprotection when PC was amplified with 3 cycles of ischemia/reperfusion (9.9 ± 1.4% infarction, p < 0.01 vs. control). Morphine could also mimic ischemic preconditioning, but only at a dose much higher than would be used clinically (3 mg/kg). In isolated hearts pretreatment with morphine (0.3 M) significantly limited infarction to 9.3 ± 1.2% (p < 0.01 vs. 32.0 ± 3.1% in controls). This cardioprotective effect of morphine could be blocked by either the PKC inhibitor chelerythrine (30.4 ± 2.6% infarction) or NAL (34.0 ± 2.6% infarction). Neither chelerythrine nor NAL by itself modified infarction in non-PC hearts. NAL could not block protection from one cycle of PC in isolated hearts indicating that an intact innervation may be required for endogenous opioid production. Thus, opioid receptors, like other PKC-coupled receptors, participate in the triggering PC in the rabbit heart.     

Effect of aging on the ability of preconditioning to protect rat hearts from ischemia-reperfusion injury

Ischemic preconditioning (IP) reduces infarct size in young animals; however, its impact on aging is underinvestigated. The effect of variations in IP stimuli was studied in young, middle-aged, and aged rat hearts. Isolated hearts underwent 35 min of regional ischemia and 120 min of reperfusion. Hearts with IP were subjected to either one ischemia-reperfusion cycle (5 min of ischemia and 5 min of reperfusion per cycle) or three successive cycles before 35 min of regional ischemia. Additional studies investigated the effects of pharmacological preconditioning in aged hearts using the adenosine A(1) receptor agonist 2-chloro-N(6)-cyclopentyladenosine, the protein kinase C analog 1,2-dioctanoyl-sn-glycerol, and the mitochondrial ATP-sensitive potassium (K(ATP))-channel opener diazoxide. Infarct sizes indicated that the aged rat heart could not be preconditioned via ischemic or pharmacological means. The middle-aged rat heart had a blunted IP response compared with the young adult (only an increased IP stimulus caused a significant reduction in infarct size). These results suggest that there are defects within the IP signaling cascade of the aged heart. Clinical relevance is important if we are to use any IP-like mimetics to the benefit of an aging population.     

Hyperosmotic pretreatment reduces infarct size in the rat heart

Preconditioning of the heart can be achieved by an ischemia/reperfusion stimulus, but also by stretching of the heart by an acute volume overload. Since manipulations of the extracellular osmolality affects cell size, we hypothesized that hyperosmotic pretreatment of the isolated perfused rat heart could reduce infarct size following regional ischemia (RI). Langendorff perfused rat hearts were subjected to 30 min RI by ligature of the main branch of the left coronary artery followed by 120 min reperfusion (control group). Ischemic preconditioning (IP-5') was achieved by 5 min total global ischemia and 5 min reperfusion prior to RI. Hyperosmotic pretreatment was accomplished by perfusion with a hyperosmotic buffer (600 mOsm/kg H2O by adding mannitol) for 1 min, 2 min or 5 min. At the end of the experiments, the hearts were cut into 2 mm slices, incubated with triphenyltetrazoliumchloride before scanning and computerized for estimation of infarct size. The average infarct size (as percentage of area at risk) in the control group was 42% and was significantly reduced to 16% by ischemic preconditioning and to 17% by 2 min hyperosmotic pretreatment. Neither 1 min nor 5 min hyperosmotic pretreatment reduced infarct size as compared to the controls. The infarct reducing effect of 2 min hyperosmotic pretreatment was not blunted by inhibition of protein kinase C (chelerytrine chloride), the Na+/H+-exchanger (HOE 694) or stretch-activated anion channels (gadolinium chloride). The results indicate that short-lasting hyperosmotic perturbations of the extracellular environment may precondition the heart to a subsequent ischemic insult.