Adenosine-enhanced ischemic preconditioning: adenosine receptor involvement during ischemia and reperfusion

Adenosine-enhanced ischemic preconditioning (APC) extends the cardioprotection of ischemic preconditioning (IPC) by both significantly decreasing myocardial infarct size and significantly enhancing postischemic functional recovery. In this study, the role of adenosine receptors during ischemia-reperfusion was determined. Rabbit hearts (n = 92) were used for Langendorff perfusion. Control hearts were perfused for 180 min, global ischemia hearts received 30-min ischemia and 120-min reperfusion, and IPC hearts received 5-min ischemia and 5-min reperfusion before ischemia. APC hearts received a bolus injection of adenosine coincident with IPC. Adenosine receptor (A(1), A(2), and A(3)) antagonists were used with APC before ischemia and/or during reperfusion. GR-69019X (A(1)/A(3)) and MRS-1191/MRS-1220 (A(3)) significantly increased infarct size in APC hearts when administered before ischemia and significantly decreased functional recovery when administered during both ischemia and reperfusion (P < 0.05 vs. APC). DPCPX (A(1)) administered either before ischemia and/or during reperfusion had no effect on APC cardioprotection. APC-enhanced infarct size reduction is modulated by adenosine receptors primarily during ischemia, whereas APC-enhanced postischemic functional recovery is modulated by adenosine receptors during both ischemia and reperfusion.     

Calcitonin gene-related peptide-mediated cardioprotection of postconditioning in isolated rat hearts

Previous studies have demonstrated that endogenous calcitonin gene-related peptide (CGRP) plays an important role in mediation of ischemic preconditioning. In the present study, we tested whether CGRP is also involved in mediation of the protective effects of postconditioning in isolated rat hearts. Sixty minutes of left coronary artery occlusion and followed by 60 min of reperfusion caused a significant decrease in cardiac function and a significant increase in creatine kinase (CK) release and infarct size. Postconditioning with three cycles of 1-min ischemia and 1-min reperfusion produced a marked improvement of cardiac function and decreased CK release and infarct size, concomitantly with an increase in the release of CGRP release in coronary effluent. However, the cardioprotection afforded by postconditioning was abolished by CGRP 8-37 (10^− 7 M), a selective CGRP receptor antagonist, or pretreatment with capsaicin (50 mg/kg, s.c.), which depletes transmitters in sensory nerves. Exogenous CGRP (5 × 10^− 9 M) administration of CGRP reappeared postconditioning-like cardioprotection in the rats pretreated with capsaicin. These results suggest that the protective effects of ischemic postconditioning are related to stimulation of endogenous CGRP release in rat hearts.     

Hypercholesterolemic Myocardium Is Vulnerable to Ischemia-Reperfusion Injury and Refractory to Sevoflurane-Induced Protection

Recent studies have demonstrated that volatile anesthetic postconditioning confers myocardial protection against ischemia-reperfusion (IR) injury through activation of the reperfusion injury salvage kinase (RISK) pathway. As RISK has been shown to be impaired in hypercholesterolemia. Therefore, we investigate whether anesthetic-induced cardiac protection was maintained in hypercholesterolemic rats. In the present study, normocholesteolemic or hypercholesterolemic rat hearts were subjected to 30 min of ischemia and 2 h of reperfusion. Animals received 2.4% sevoflurane for 5 min or 3 cycles of 10-s ischemia/10-s reperfusion. The hemodynamic parameters, including left ventricular developed pressure, left ventricular end-diastolic pressure and heart rate, were continuously monitored. The infarct size, apoptosis, p-Akt, p-ERK1/2, p-GSK3β were determined. We found that both sevoflurane and ischemic postconditioning significantly improved heart pump function, reduced infarct size and increased the phosphorylation of Akt, ERK1/2 and their downstream target of GSK3β in the healthy rats. In the hypercholesterolemic rats, neither sevoflurane nor ischemic postconditioning improved left ventricular hemodynamics, reduced infarct size and increased the phosphorylated Akt, ERK1/2 and GSK3β. In contrast, GSK inhibitor SB216763 conferred cardioprotection against IR injury in healthy and hypercholesterolemic hearts. In conclusions, hyperchoesterolemia abrogated sevoflurane-induced cardioprotection against IR injury by alteration of upstream signaling of GSK3β and acute GSK inhibition may provide a novel therapeutic strategy to protect hypercholesterolemic hearts against IR injury.     

Noninvasive delayed limb ischemic preconditioning attenuates myocardial ischemia-reperfusion injury in rats by a mitochondrial K(ATP) channel-dependent mechanism

We previously demonstrated in rats that noninvasive delayed limb ischemic preconditioning (LIPC) induced by three cycles of 5-min occlusion and 5-min reperfusion of the left hind limb per day for three days confers the same cardioprotective effect as local ischemic preconditioning of the heart, but the mechanism has not been studied in depth. The aim of this project was to test the hypothesis that delayed LIPC enhances myocardial antioxidative ability during ischemia-reperfusion by a mitochondrial K(ATP) channel (mito K(ATP))-dependent mechanism. Rats were randomized to five groups: ischemia-reperfusion (IR)-control group, myocardial ischemic preconditioning (MIPC) group, LIPC group, IR-5HD group and LIPC-5HD group. The MIPC group underwent local ischemic preconditioning induced by three cycles of 5-min occlusion and 5-min reperfusion of the left anterior descending coronary arteries. The LIPC and LIPC-5HD groups underwent LIPC induced by three cycles of 5-min occlusion and 5-min reperfusion of the left hind limb using a modified blood pressure aerocyst per day for three days. All rats were subjected to myocardial ischemia-reperfusion injury. The IR-5HD and LIPC-5HD groups received the mito K(ATP) channel blocker 5-hydroxydecanoate Na (5-HD) before and during the myocardial ischemia-reperfusion injury. Compared with the IR-control group, both the LIPC and MIPC groups showed an amelioration of ventricular arrhythmia, reduced myocardial infarct size, increased activities of total superoxide dismutase, manganese-superoxide dismutase (Mn-SOD) and glutathione peroxidase, increased expression of Mn-SOD mRNA and decreased xanthine oxidase activity and malondialdehyde concentration. These beneficial effects of LIPC were prevented by 5-HD. In conclusion, delayed LIPC offers similar cardioprotection as local IPC. These results support the hypothesis that the activation of mito K(ATP) channels enhances myocardial antioxidative ability during ischemia-reperfusion, thereby contributing, at least in part, to the anti-arrhythmic and anti-infarct effects of delayed LIPC.     

Myocardial preconditioning against ischemia-reperfusion injury is abolished in Zucker obese rats with insulin resistance

Insulin resistance (IR) precedes the onset of Type 2 diabetes, but its impact on preconditioning against myocardial ischemia-reperfusion injury is unexplored. We examined the effects of diazoxide and ischemic preconditioning (IPC; 5-min ischemia and 5-min reperfusion) on ischemia (30 min)-reperfusion (240 min) injury in young IR Zucker obese (ZO) and lean (ZL) rats. ZO hearts developed larger infarcts than ZL hearts (infarct size: 57.3 +/- 3% in ZO vs. 39.2 +/- 3.2% in ZL; P < 0.05) and also failed to respond to cardioprotection by IPC or diazoxide (47.2 +/- 4.3% and 52.5 +/- 5.8%, respectively; P = not significant). In contrast, IPC and diazoxide treatment reduced the infarct size in ZL hearts (12.7 +/- 2% and 16.3 +/- 6.7%, respectively; P < 0.05). The mitochondrial ATP-activated potassium channel (K(ATP)) antagonist 5-hydroxydecanoic acid inhibited IPC and diazoxide-induced preconditioning in ZL hearts, whereas it had no effect on ZO hearts. Diazoxide elicited reduced depolarization of isolated mitochondria from ZO hearts compared with ZL (73 +/- 9% in ZL vs. 39 +/- 9% in ZO; P < 0.05). Diazoxide also failed to enhance superoxide generation in isolated mitochondria from ZO compared with ZL hearts. Electron micrographs of ZO hearts revealed a decreased number of mitochondria accompanied by swelling, disorganized cristae, and vacuolation. Immunoblots of mitochondrial protein showed a modest increase in manganese superoxide dismutase in ZO hearts. Thus obesity accompanied by IR is associated with the inability to precondition against ischemic cardiac injury, which is mediated by enhanced mitochondrial oxidative stress and impaired activation of mitochondrial K(ATP).     

Involvement of CCR-2 chemokine receptor activation in ischemic preconditioning and postconditioning of brain in mice

The present study has been designed to investigate the potential role of CCR-2 chemokine receptor in ischemic preconditioning as well as postconditioning induced reversal of ischemia-reperfusion injury in mouse brain. Bilateral carotid artery occlusion of 17min followed by reperfusion for 24h was employed in present study to produce ischemia and reperfusion induced cerebral injury in mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining. Memory was evaluated using elevated plus-maze test and Morris water maze test. Rota rod test was employed to assess motor incoordination. Bilateral carotid artery occlusion followed by reperfusion produced cerebral infarction and impaired memory and motor co-ordination. Three preceding episodes of bilateral carotid artery occlusion for 1min and reperfusion of 1min were employed to elicit ischemic preconditioning of brain, while three episodes of bilateral carotid artery occlusion for 10s and reperfusion of 10s immediately after the completion of were employed to elicit ischemic postconditioning of brain. Both prior ischemic preconditioning as well as ischemic postconditioning immediately after global cerebral ischemia prevented markedly ischemia-reperfusion-induced cerebral injury as measured in terms of infarct size, loss of memory and motor coordination. RS 102895, a selective CCR-2 chemokine receptor antagonist, attenuated the neuroprotective effect of both the ischemic preconditioning as well as postconditioning. It is concluded that the neuroprotective effect of both ischemic preconditioning as well as ischemic postconditioning may involve the activation of CCR-2 chemokine receptors.     

Prolonged transient acidosis during early reperfusion contributes to the cardioprotective effects of postconditioning

We have previously reported that the prolonged transient acidosis during early reperfusion mediates the cardioprotective effects in canine hearts. Recently, postconditioning has been shown to be one of the novel strategies to mediate cardioprotection. We tested the contribution of the prolonged transient acidosis to the cardioprotection of postconditioning. Open-chest anesthetized dogs subjected to 90-min occlusion of the left anterior descending coronary artery and 6-h reperfusion were divided into four groups: 1) control group; no intervention after reperfusion (n = 6); 2) postconditioning (Postcon) group; four cycles of 1-min reperfusion and 1-min reocclusion (n = 7); 3) Postcon + sodium bicarbonate (NaHCO(3)) group; four cycles of 1-min reperfusion and 1-min reocclusion with the administration of NaHCO(3) (n = 8); and 4) NaHCO(3) group; administration of NaHCO(3) without postconditioning (n = 6). Infarct size, the area at risk (AAR), collateral blood flow during ischemia, and pH in coronary venous blood were measured. The phosphorylation of Akt and extracellular signal-regulated kinase (ERK) in ischemic myocardium was assessed by Western blot analysis. Systemic hemodynamic parameters, AAR, and collateral blood flow were not different among the four groups. Postconditioning induced prolonged transient acidosis during the early reperfusion phase. Administration of NaHCO(3) completely abolished the infarct size-limiting effects of postconditioning. Furthermore, the phosphorylation of Akt and ERK in ischemic myocardium induced by postconditioning was also blunted by the cotreatment of NaHCO(3). In conclusion, postconditioning mediates its cardioprotective effects possibly via prolonged transient acidosis during the early reperfusion phase with the activation of Akt and ERK.     

Effect of estrogen replacement treatment on ischemic preconditioning in isolated rat hearts

In the present study, we tested the effects of long-term estrogen replacement treatment on myocardial ischemia-reperfusion injury and on the cardioprotection of ischemic preconditioning in isolated hearts from ovariectomized rats. Ovariectomized rats were treated with 17beta-estradiol (30 micro g/kg/d, s.c.) for 12 weeks. Isolated rat hearts were perfused in the Langendorff mode. Heart rate, coronary flow, left ventricular pressure and its first derivative (+/-LVdp/dtmax) were recorded. Fifteen-min global ischemia and 30-min reperfusion caused a significant decrease of cardiac mechanical function, which were not affected by ovariectomy or estrogen replacement treatment. The isolated hearts in all groups could be preconditioned, and the cardioprotection afforded by preconditioning in the sham-operated rats was greater compared with ovariectomized rats with or without estrogen treatment. These results suggest that long-term estrogen replacement treatment exerts no effect on the inhibition of mechanical function after ischemia-reperfusion, and this study also suggests that estrogen does not affect ischemic preconditioning in isolated hearts of ovariectomized rats.     

Impact of 6-mo caloric restriction on myocardial ischemic tolerance: possible involvement of nitric oxide-dependent increase in nuclear Sirt1

Ischemic tolerance decreases with aging, and the cardioprotective effect of ischemic preconditioning (IPC) is impaired in middle-aged animals. We have demonstrated that short-term caloric restriction (CR) improves myocardial ischemic tolerance in young and old animals via the activation of adiponectin-AMP-activated protein kinase (AMPK)-mediated signaling. However, it is unknown whether prolonged CR confers cardioprotection in a similar manner. Furthermore, little is known regarding the myocardial expression of silent information regulator 1 (Sirt1; which reportedly mediates various aspects of the CR response) with prolonged CR. Thus, 6-mo-old male Fischer-344 rats were randomly divided into ad libitum (AL) and CR groups. Six months later, isolated perfused hearts were subjected to 25 min of global ischemia followed by 120 min of reperfusion with or without IPC. CR improved the recovery of left ventricular function and reduced infarct size after ischemia-reperfusion and restored the IPC effect. Serum adiponectin levels increased, but myocardial levels of total and phosphorylated AMPK did not change with prolonged CR. Total levels of Sirt1 did not change with CR; however, in the nuclear fraction, CR significantly increased Sirt1 and decreased acetyl-histone H3. Eleven rats from each group were given N-nitro-l-arginine methyl ester in their drinking water for 4 wk before death. In these hearts, chronic inhibition of nitric oxide synthase prevented the increase in nuclear Sirt1 content by CR and abrogated CR-induced cardioprotection. These results demonstrate that 1) prolonged CR improves myocardial ischemic tolerance and restores the IPC effect in middle-aged rats and 2) CR-induced cardioprotection is associated with a nitric oxide-dependent increase in nuclear Sirt1 content.     

Attenuation of ischemia–reperfusion injury by sevoflurane postconditioning involves protein kinase B and glycogen synthase kinase 3 beta activation in isolated rat hearts

Volatile anesthetic ischemic postconditioning reduces infarct size following ischemia/reperfusion. Whether phosphorylation of protein kinase B (PKB/Akt) and glycogen synthase kinase 3 beta (GSK3β) is causal for cardioprotection by postconditioning is controversial. We therefore investigated the impact of PKB/Akt and GSK3β in isolated perfused rat hearts subjected to 40 min of ischemia followed by 1 h of reperfusion. 2.0% sevoflurane (1.0 minimum alveolar concentration) was administered at the onset of reperfusion in 15 min as postconditioning. Western blot analysis was used to determine phosphorylation of PKB/Akt and its downstream target GSK3β after 1 h of reperfusion. Mitochondrial and cytosolic content of cytochrome C checked by western blot served as a marker for mitochondrial permeability transition pore opening. Sevoflurane postconditioning significantly improved functional cardiac recovery and decreased infarct size in isolated rat hearts. Compared with unprotected hearts, sevoflurane postconditioning-induced phosphorylation of PKB/Akt and GSK3β were significantly increased. Increase of cytochrome C in mitochondria and decrease of it in cytosol is significant when compared with unprotected ones which have reversal effects on cytochrome C. The current study presents evidence that sevoflurane-induced cardioprotection at the onset of reperfusion are partly through activation of PKB/Akt and GSK3β.     

Increased mitochondrial calcium coexists with decreased reperfusion injury in postconditioned (but not preconditioned) hearts

Ca(2+) is the main trigger for mitochondrial permeability transition pore opening, which plays a key role in cardiomyocyte death after ischemia-reperfusion. We investigated whether a reduced accumulation of mitochondrial Ca(2+) might explain the attenuation of lethal reperfusion injury by postconditioning. Anesthetized New Zealand White rabbits underwent 30 min of ischemia, followed by either 240 (infarct size protocol) or 60 (mitochondria protocol) min of reperfusion. They received either no intervention (control), preconditioning by 5-min ischemia and 5-min reperfusion, postconditioning by four cycles of 1-min reperfusion and 1-min ischemia at the onset of reflow, or pharmacological inhibition of the transition pore opening by N-methyl-4-isoleucine-cyclosporin (NIM811; 5 mg/kg iv) given at reperfusion. Area at risk and infarct size were assessed by blue dye injection and triphenyltetrazolium chloride staining. Mitochondria were isolated from the risk region for measurement of 1) Ca(2+) retention capacity (CRC), and 2) mitochondrial content of total (atomic absorption spectrometry) and ionized (potentiometric technique) calcium concentration. CRC averaged 0.73 +/- 0.16 in control vs. 4.23 +/- 0.17 mug Ca(2+)/mg proteins in shams (P < 0.05). Postconditioning, preconditioning, or NIM811 significantly increased CRC (P < 0.05 vs. control). In the control group, total and free mitochondrial calcium significantly increased to 2.39 +/- 0.43 and 0.61 +/- 0.10, respectively, vs. 1.42 +/- 0.09 and 0.16 +/- 0.01 mug Ca(2+)/mg in sham (P < 0.05). Surprisingly, whereas total and ionized mitochondrial Ca(2+) decreased in preconditioning, it significantly increased in postconditioning and NIM811 groups. These data suggest that retention of calcium within mitochondria may explain the decreased reperfusion injury in postconditioned (but not preconditioned) hearts.     

The cardioprotection of the late phase of ischemic preconditioning is enhanced by postconditioning via a COX-2-mediated mechanism in conscious rats

The present study sought to determine whether the combination of late preconditioning (PC) with postconditioning enhances the reduction in infarct size. Chronically instrumented rats were assigned to a 45-min (subset 1) or 60-min (subset 2) coronary occlusion followed by 24 h of reperfusion. In each subset, rats received no further intervention (control) or were preconditioned 24 h before occlusion (PC), postconditioned at the onset of reperfusion following occlusion, or preconditioned and postconditioned without (PC + postconditioning) or with the COX-2 inhibitor celecoxib (3 mg/kg ip; PC + postconditioning + celecoxib) 10 min before postconditioning. Myocardial cyclooxygenase-2 (COX-2) protein expression and COX-2 activity (assessed as myocardial levels of PGE(2)) were measured 6 min after reperfusion in an additional five groups (control, PC, postconditioning, PC + postconditioning, and PC + postconditioning + celecoxib) subjected to a 45-min occlusion. PC alone reduced infarct size after a 45-min occlusion but not after a 60-min occlusion. Postconditioning alone did not reduce infarct size in either setting. However, the combination of late PC and postconditioning resulted in a robust infarct-sparing effect in both settings, suggesting additive cardioprotection. Celecoxib completely abrogated the infarct-sparing effect of the combined interventions in both settings. Late PC increased COX-2 protein expression and PGE(2) content. PGE(2) content (but not COX-2 protein) was further increased by the combination of both interventions, suggesting that postconditioning increases the activity of COX-2 induced by late PC. In conclusion, the combination of late PC and postconditioning produces additive protection, likely due to a postconditioning-induced enhancement of COX-2 activity.     

Sphingosine kinase and sphingosine 1-phosphate in the heart: A decade of progress

Activation of sphingosine kinase/sphingosine 1-phosphate (SK/S1P)‐mediated signaling has emerged as a critical cardioprotective pathway in response to acute ischemia/reperfusion injury. S1P is released in both ischemic pre- and post-conditioning. Application of exogenous S1P to cultured cardiac myocytes subjected to hypoxia or treatment of isolated hearts either before ischemia or at the onset of reperfusion exerts prosurvival effects. Synthetic congeners of S1P such as FTY720 mimic these responses. Gene targeted mice null for the SK1 isoform whose hearts are subjected to ischemia/reperfusion injury exhibit increased infarct size and respond poorly either to ischemic pre- or postconditioning. Measurements of cardiac SK activity and S1P parallel these observations. Experiments in SK2 knockout mice have revealed that this isoform is necessary for survival in the heart. High density lipoprotein (HDL) is a major carrier of S1P, and studies of hearts in which selected S1P receptors have been inhibited implicate the S1P cargo of HDL in cardioprotection. Inhibition of S1P lyase, an endogenous enzyme that degrades S1P, also leads to cardioprotection. These observations have considerable relevance for future therapeutic approaches to acute and chronic myocardial injury. This article is part of a Special Issue entitled Advances in Lysophospholipid Research.     

Sevoflurane postconditioning protects isolated rat hearts against ischemia-reperfusion injury: the role of radical oxygen species,extracellular signal-related kinases 1/2 and mitochondrial permeability transition pore

The roles of reactive oxygen species (ROS), extracellular signal-regulated kinase 1/2 (ERK 1/2) and mitochondrial permeability transition pore (mPTP) in sevoflurane postconditioning induced cardioprotection against ischemia-reperfusion injury in Langendorff rat hearts were investigated. When compared with the unprotected hearts subjected to 30 min of ischemia followed by 1 h of reperfusion, exposure of 3% sevoflurane during the first 15 min of reperfusion significantly improved functional recovery, decreased infarct size, reduced lactate dehydrogenase and creatine kinase-MB release, and reduced myocardial malondialdehyde production. However, these protective effects were abolished in the presence of either ROS scavenger N-acetylcysteine or ERK 1/2 inhibitor PD98059, and accompanied by prevention of ERK 1/2 phosphorylation and elimination of inhibitory effect on mPTP opening. These findings suggested that sevoflurane postconditioning protected isolated rat hearts against ischemia-reperfusion injury via the recruitment of the ROS-ERK 1/2-mPTP signaling cascade.     

P2X7 receptor agonists pre- and postcondition the heart against ischemia-reperfusion injury by opening pannexin-1/P2X₇ channels

Protection of the heart from ischemia-reperfusion injury can be achieved by ischemic preconditioning and ischemic postconditioning. Previous studies revealed that a complex of pannexin-1 with the P2X(7) receptor forms a channel during ischemic preconditioning and ischemic postconditioning that results in the release of endogenous cardioprotectants. ATP binds to P2X(7) receptors, inducing the formation of a channel in association with pannexin-1. We hypothesized that this channel would provide a pathway for the release of these same cardioprotectants. Preconditioning-isolated perfused rat hearts with 0.4 μM ATP preceding 40 min of ischemia minimized infarct size upon subsequent reperfusion (5% of risk area) and resulted in >80% recovery of left ventricular developed pressure. Postconditioning with ATP after ischemia during reperfusion was also protective (6% infarct and 72% recovery of left ventricular developed pressure). Antagonists of both pannexin-1 (carbenoxolone and mefloquine) and P2X(7) receptors (brilliant blue G and A438079) blocked ATP pre- and postconditioning, indicating that ATP protection was elicited via the opening of a pannexin-1/P2X(7) channel. An antagonist of binding of the endogenous cardioprotectant sphingosine 1-phosphate to its G protein-coupled receptor diminished protection by ATP, which is also consistent with an ATP-dependent release of cardioprotectants. Suramin, an antagonist of binding of ATP (and ADP) to P2Y receptors, was without effect on ATP protection. Benzoyl benzoyl-ATP, a more specific P2X(7) agonist, was also a potent pre- and postconditioning agent and sensitive to blockade by pannexin-1/P2X(7) channel antagonists. The data point out for the first time the potential of P2X(7) agonists as cardioprotectants.     

Effect of age on alpha-calcitonin gene-related peptide-mediated delayed cardioprotection induced by intestinal preconditioning in rats

In the present study, we examined whether age-related reduction in cardioprotection of intestinal ischemic preconditioning is related to stimulation of the release and synthesis of calcitonin gene-related peptide (CGRP) in rats. Ischemia-reperfusion injury was induced by a 45-min coronary artery occlusion and 180-min reperfusion, and ischemic preconditioning was induced by six cycles of 4-min ischemia and 4-min reperfusion of the small intestine. The serum concentration of creatine kinase, infarct size, the expression of CGRP isoforms (alpha- and beta-CGRP) mRNA in lumbar dorsal root ganglia and CGRP concentration in plasma were measured. Pretreatment with intestinal ischemic preconditioning for 24 h significantly reduced infarct size and creatine kinase release concomitantly with a significant increase in the expression of alpha-CGRP mRNA, but not beta-CGRP mRNA, and plasma concentrations of CGRP at 6 months of age but not at 24 months of age. These results suggest that the delayed cardioprotective effect of intestinal ischemic preconditioning is decreased in senescent rats, and the age-related change is related to reduction of the synthesis and release of alpha-CGRP.     

Involvement of the mitochondrial calcium uniporter in cardioprotection by ischemic preconditioning

The objective of the present study was to determine whether the mitochondrial calcium uniporter plays a role in the cardioprotection induced by ischemic preconditioning (IPC). Isolated rat hearts were subjected to 30 min of regional ischemia by ligation of the left anterior descending artery followed by 120 min of reperfusion. IPC was achieved by two 5-min periods of global ischemia separated by 5 min of reperfusion. IPC reduced the infarct size and lactate dehydrogenase release in coronary effluent, which was associated with improved recovery of left ventricular contractility. Treatment with ruthenium red (RR, 5 μM), an inhibitor of the uniporter, or with Ru360 (10 μM), a highly specific uniporter inhibitor, provided cardioprotective effects like those of IPC. The cardioprotection induced by IPC was abolished by spermine (20 μM), an activator of the uniporter. Cyclosporin A (CsA, 0.2 μM), an inhibitor of the mitochondrial permeability transition pore, reversed the effects caused by spermine. In mitochondria isolated from untreated hearts, both Ru360 (10 μM) and RR (1 μM) decreased pore opening, while spermine (20 μM) increased pore opening which was blocked by CsA (0.2 μM). In mitochondria from preconditioned hearts, the opening of the pore was inhibited, but this inhibition did not occur in the mitochondria from hearts treated with IPC plus spermine. These results indicate that the mitochondrial calcium uniporter is involved in the cardioprotection conferred by ischemic preconditioning.     

Ischemic postconditioning during reperfusion activates Akt and ERK without protecting against lethal myocardial ischemia-reperfusion injury in pigs

Transient episodes of ischemic preconditioning (PC) render myocardium protected against subsequent lethal injury after ischemia and reperfusion. Recent studies indicate that application of short, repetitive ischemia only during the onset of reperfusion after the lethal ischemic event may obtain equivalent protection. We assessed whether such ischemic postconditioning (Postcon) is cardioprotective in pigs by limiting lethal injury. Pentobarbital sodium-anesthetized, open-chest pigs underwent 30 min of complete occlusion of the left anterior descending coronary artery and 3-h reflow. PC was elicited by two cycles of 5-min occlusion plus 10-min reperfusion before the 30-min occlusion period. Postcon was elicited by three cycles of 30-s reperfusion, followed by 30-s reocclusion, after the 30-min occlusion period and before the 3-h reflow. Infarct size (%area-at-risk using triphenyltetrazolium chloride macrochemistry; means +/- SE) after 30 min of ischemia was 26.5 +/- 5.2% (n = 7 hearts/treatment group). PC markedly limited myocardial infarct size (2.8 +/- 1.2%, n = 7 hearts/treatment group, P < 0.05 vs. controls). However, Postcon had no effect on infarct size (37.8 +/- 5.1%, n = 7 hearts/treatment group). Within the subendocardium, Postcon increased phosphorylation of Akt (74 +/- 12%) and ERK1/2 (56 +/- 10%) compared with control hearts subjected only to 30-min occlusion and 15-min reperfusion (P < or = 0.05), and these changes were not different from the response triggered by PC (n = 5 hearts/treatment group). Phosphorylation of downstream p70S6K was also equivalent in PC and Postcon groups. These data do not support the hypothesis that application of 30-s cycles of repetitive ischemia during reperfusion exerts a protective effect on pig hearts subjected to lethal ischemia, but this is not due to a failure to phosphorylate ERK and Akt during early reperfusion.     

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.     

肾神经在肾缺血预处理对麻醉家兔心脏保护中的作用

在氨基甲酸乙酯麻醉家兔上,观察肾脏缺血预处理(RIP)对缺血-再灌注心肌的影响,旨在证实RIP对心肌有无保护效应,并明确肾神经在其中的作用。所得结果如下(1)在心脏45min缺血和180min再灌注过程中,血压、心率和心肌耗氧量呈进行性下降;心外膜电图ST段在缺血期明显抬高,再灌注过程中逐渐恢复到基础对照值。心肌梗塞范围占缺血心肌的55.80±1.25%。(2)RIP时心肌梗塞范围为36.51±2.8%,较单纯心肌缺血-再灌注显著减少(P<0.01),表明RIP对心肌有保护作用。(3)肾神经切断可取消RIP对心肌的保护效应,但肾神经切断本身对单纯缺血-再灌注所致的心肌梗死范围无明显影响。(4)肾缺血(10min)时,肾传入神经放电活动由0.14±0.08增至0.65±0.12imp/s(P<0.01)。(5)预先应用腺苷受体拮抗剂8-苯茶碱可明显减弱肾缺血所激活的肾传入神经活动,提示肾传入活动的增强是由肾缺血产生的腺苷所介导。以上结果表明,肾短暂缺血-再灌注所诱发的肾神经传入活动在RIP心肌保护效应中起重要作用。