Involvement of src-kinase activation in ischemic preconditioning induced protection of mouse brain

AimsTo investigate the role of src-kinase in ischemic preconditioning induced reversal of ischemia and reperfusion induced cerebral injury in mice.Main methodsBilateral carotid artery occlusion of 17 min followed by reperfusion for 24 h was employed to produce ischemia and reperfusion induced cerebral injury in mice. Cerebral infarct size was measured using triphenyltetrazolium chloride staining using both by volume and by weight methods differently. Memory was evaluated using elevated plus maze test. Rota rod test was employed to assess motor incoordination.Key findingsBilateral 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 1 min and reperfusion of 1 min (ischemic preconditioning) prevented markedly ischemia–reperfusion-induced cerebral injury measured in terms of infarct size (38.5 ± 1.3% and 38.5 ± 2.9% mean infarct of control animals was reduced to 24.3 ± 1.2% and 23.5 ± 1.8% of the preconditioning groups respectively), loss of memory (72.2 ± 3.6 mean transfer latency time of control animals was reduced to 25.6 ± 5.2 of the preconditioning group respectively) and motor coordination (78.3 ± 17.6 s mean falling down latency time of control animals was increased to a mean value of 180.9 ± 6.5 s of the preconditioning groups respectively). SU6656 (2 mg/kg, ip) and PP1 (0.1 mg/kg, ip), highly selective src-kinase inhibitors, attenuated this neuroprotective effect of ischemic preconditioning.SignificanceTherefore, neuroprotective effect of ischemic preconditioning may be due to src-kinase linked mechanism.     

Inhibition of myocardial injury by ischemic postconditioning during reperfusion: comparison with ischemic preconditioning

Ischemic preconditioning (Pre-con) is an adaptive response triggered by a brief ischemia applied before a prolonged coronary occlusion. We tested the hypothesis that repetitive ischemia applied during early reperfusion, i.e., postconditioning (Post-con), is cardio-protective by attenuating reperfusion injury. In anesthetized open-chest dogs, the left anterior descending artery (LAD) was occluded for 60 min and reperfused for 3 h. In controls (n = 10), there was no intervention. In Pre-con (n = 9), the LAD was occluded for 5 min and reperfused for 10 min before the prolonged occlusion. In Post-con (n = 10), at the start of reperfusion, three cycles of 30-s reperfusion and 30-s LAD reocclusion preceded the 3 h of reperfusion. Infarct size was significantly less in the Pre-con (15 +/- 2%, P < 0.05) and Post-con (14 +/- 2%, P < 0.05) groups compared with controls (25 +/- 3%). Tissue edema (% water content) in the area at risk was comparably reduced in Pre-con (78.3 +/- 1.2, P < 0.05) and Post-con (79.7 +/- 0.6, P < 0.05) versus controls (81.5 +/- 0.4). Polymorphonuclear neutrophil (PMN) accumulation (myeloperoxidase activity, Deltaabsorbance.min-1.g tissue-1) in the area at risk myocardium was comparably reduced in Post-con (10.8 +/- 5.5, P < 0.05) and Pre-con (13.4 +/- 3.4, P < 0.05) versus controls (47.4 +/- 15.3). Basal endothelial function measured by PMN adherence to postischemic LAD endothelium (PMNs/mm2) was comparably attenuated by Post-con and Pre-con (15 +/- 0.6 and 12 +/- 0.6, P < 0.05) versus controls (37 +/- 1.5), consistent with reduced expression of P-selectin on coronary vascular endothelium in Post-con and Pre-con. Endothelial function assessed by the maximal vasodilator response of postischemic LAD to acetylcholine was significantly greater in Post-con (104 +/- 6%, P < 0.05) and Pre-con (109 +/- 5%, P < 0.05) versus controls (71 +/- 8%). Plasma malondialdehyde (microM/ml), a product of lipid peroxidation, was significantly less at 1 h of reperfusion in Post-con (2.2 +/- 0.2, P < 0.05) versus controls (3.2 +/- 0.3) associated with a decrease in superoxide levels revealed by dihydroethidium staining in the myocardial area at risk. These data suggest that Post-con is as effective as Pre-con in reducing infarct size and preserving endothelial function. Post-con may be clinically applicable in coronary interventions, coronary artery bypass surgery, organ transplantation, and peripheral revascularization where reperfusion injury is expressed.     

Protease-activated receptor-2 activation improves efficiency of experimental ischemic preconditioning

Protease-activated receptor-2 (PAR-2) is a member of seven transmembrane domain G protein-coupled receptors activated by proteolytic cleavage. PAR-2 is involved in inflammatory events and cardiac ischemic reperfusion injury. The objective of this study was to investigate the effects of PAR-2 in experimental myocardial ischemic preconditioning. To monitor the effects of PAR-2, Langendorff-perfused rat hearts were used. These hearts were treated with PAR-2-activating peptide (PAR-2AP) in various protocols. Hemodynamic parameters (left ventricular developed pressure, left ventricular diastolic pressure, coronary flow rate, and heart rate), several indexes of oxidative injury, and neutrophil accumulation were evaluated. We show for the first time that enhanced PAR-2 activation improves efficiency of ischemic preconditioning and reduces cardiac inflammation in the rat heart. Indeed, after PAR-2AP infusion we found that hemodynamic parameters, oxidative injury, infarct size, and neutrophil accumulation were involved. These data support the concept that PAR-2-dependent cell trafficking may regulate signaling responses to cardiac ischemia and inflammation.     

Combined preconditioning and postconditioning provides synergistic protection against liver ischemic reperfusion injury

Hepatic Ischemia and Reperfusion Injury (IRI) is a major cause of liver damage during liver surgery and transplantation. Ischemic preconditioning and postconditioning are strategies that can reduce IRI. In this study, different combined types of pre- and postconditioning procedures were tested in a murine warm hepatic IRI model to evaluate their protective effects. Proanthocyanidins derived from grape seed was used before ischemia process as pharmacological preconditioning to combine with technical preconditioning and postconditioning. Three pathways related to IRI, including reactive oxygen species (ROS) generation, pro-inflammatory cytokines release and hypoxia responses were examined in hepatic IRI model. Individual and combined pre- and postconditioning protocols significantly reduce liver injury by decreasing the liver ROS and cytokine levels, as well as enhancing the hypoxia tolerance response. Our data also suggested that in addition to individual preconditioning or postconditioning, the combination of these two treatments could reduce liver ischemia/reperfusion injury more effectively by increasing the activity of ROS scavengers and antioxidants. The utilization of grape seed proanthocyanidins (GSP) could improve the oxidation resistance in combined pre- and postconditioning groups. The combined protocol also further increased the liver HIF-1 alpha protein level, but had no effect on pro-inflammatory cytokines release compared to solo treatment.     

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.     

Role of receptor transactivation in the cardioprotective effects of preconditioning and postconditioning

Analysis of published data indicates that the activity of receptors for adenosine, opioids, bradykinin, calcitonin-gene related peptides (CGRP) and epidermal growth factor (EGF) play important role in triggering the cardioprotective effects of ischemic preconditioning. Cannabinoids mimic the infarct-sparing effects of preconditioning. Endogenous adenosine, opioids, bradykinin and CGRP have also been implicated in infarct-reduction with ischemic postconditioning. Again, cannabinoids also mimic the protective effect of postconditioning. Recent works support heterodimerization of G-protein coupled receptors (GPCRs), and GPCR transactivation of EGF receptors. It was found that cross-talk between delta(j)-opioid receptors and adenosine A(1)-receptors is essential to cardiac protection. Furthermore, evidence implicates EGF receptor transactivation in cardioprotective effect of multiple GPCrs including adenosine, acetylcholine, bradykinin, and opioid receptors. Such findings support a convergent pathway in which multiple GPCRs may interact (or function independently) to transactivate EGF receptor-dependent kinase signaling and cytoprotection.     

Involvement of metallothionein in the protection of lung ischemic preconditioning

The aim of the present study was to investigate whether metallothionein was involved in the protection of lung ischemic preconditioning (IP) against lung ischemia-reperfusion (I/R) injury. Adult male Sprague-Dawley rats were randomly divided into 3 groups based upon the intervention (n=8): control group (C), lung I/R group (I/R), lung I/R+IP group (IP). At the end of the experiment, the content of metallothionein was tested in lung tissue. Blood specimens collected from the arteria carotis were tested for the contents of malondialdehyde (MDA), the activities of superoxide dismutase (SOD) and myeloperoxidase (MPO). The pneumocyte apoptosis index (AI) was determined by terminal deoxynucleotidyl transferase mediated dUTP nick end labeling (TUNEL). Ultrastructural changes of lung tissue were observed by using transmission electron microscope. The results showed that in I/R group, the content of metallothionein was decreased (P<0.05), the content of MDA and MPO activity were increased (P<0.01), and SOD activity was decreased (P<0.01), compared with those in control group. IP treatment significantly increased the content of metallothionein (P<0.01), attenuated the MDA level (P<0.05) and MPO activity (P<0.01), and improved SOD activity (P<0.01) in blood serum. The number of TUNEL-positive cells in IP group was significantly reduced compared with that in I/R group (P<0.01). There were abnormal ultrastructural changes in I/R group, which were markedly reversed in IP group. In conclusion, IP may protect lung against I/R injury by inducing the expression of metallothionein.     

Involvement of MAPK activation in chemokine or COX-2 productions by Toxoplasma gondii

This experiment focused on MAPK activation in host cell invasion and replication of T. gondii, as well as the expression of CC chemokines, MCP-1 and MIP-1 alpha , and enzyme, COX-2/prostaglandin E2 (PGE2) in infected cells via western blot, [3H]-uracil incorporation assay, ELISA and RT-PCR. The phosphorylation of ERK1/2 and p38 in infected HeLa cells was detected at 1 hr and/or 6 hr postinfection (PI). Tachyzoite proliferation was reduced by p38 or JNK MAPK inhibitors. MCP-1 secretion was enhanced in infected peritoneal macrophages at 6 hr PI. MIP-1 alpha mRNA was increased in macrophages at 18 hr PI. MCP-1 and MIP-1 alpha were reduced after treatment with inhibitors of ERK1/2 and JNK MAPKs. COX-2 mRNA gradually increased in infected RAW 264.7 cells and the secretion of COX-2 peaked at 6 hr PI. The inhibitor of JNK suppressed COX-2 expression. PGE2 from infected RAW 264.7 cells was increased and synthesis was suppressed by PD98059, SB203580, and SP600125. In this study, the activation of p38, JNK and/or ERK1/2 MAPKs occurred during the invasion and proliferation of T. gondii tachyzoites in HeLa cells. Also, increased secretion and expression of MCP-1, MIP-1 alpha , COX-2 and PGE2 were detected in infected macrophages, and appeared to occur via MAPK signaling pathways.     

Hypoxic preconditioning protects human brain endothelium from ischemic apoptosis by Akt-dependent survivin activation

Preconditioning-induced ischemic tolerance is well documented in the brain, but cell-specific responses and mechanisms require further elucidation. The aim of this study was to develop an in vitro model of ischemic tolerance in human brain microvascular endothelial cells (HBMECs) and to examine the roles of phosphatidylinositol 3-kinase (PI3-kinase)/Akt and the inhibitor-of- apoptosis protein, survivin, in the ability of hypoxic preconditioning (HP) to protect endothelium from apoptotic cell death. Cultured HBMECs were subjected to HP, followed 16 h later by complete oxygen and glucose deprivation (OGD) for 8 h; cell viability was quantified at 20 h of reoxygenation (RO) by the 3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide assay. HBMECs were examined at various times after HP or OGD/RO using immunoblotting and confocal laser scanning immunofluorescence microscopy for appearance of apoptotic markers and expression of phosphorylated (p)-Akt and p-survivin. Causal evidence for the participation of the PI3-kinase/Akt pathway in HP-induced protection and p-survivin upregulation was assessed by the PI3-kinase inhibitor LY-294002. HP significantly reduced OGD/RO-induced injury by 50% and also significantly reduced the OGD-induced translocation of apoptosis-inducing factor (AIF) from mitochondria to nucleus and the concomitant cleavage of poly(ADP-ribose) polymerase-1 (PARP-1). PI3-kinase inhibition blocked HP-induced increases in Akt phosphorylation, reversed the effects of HP on OGD-induced AIF translocation and PARP-1 cleavage, blocked HP-induced survivin phosphorylation, and ultimately attenuated HP-induced protection of HBMECs from OGD. Thus HP promotes an antiapoptotic phenotype in HBMECs, in part by activating survivin via the PI3-kinase/Akt pathway. Survivin and other phosphorylation products of p-Akt may be therapeutic targets to protect cerebrovascular endothelium from apoptotic injury following cerebral ischemia.     

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.     

Co-application of ischemic preconditioning and postconditioning provides additive neuroprotection against spinal cord ischemia in rabbits

Postconditioning can induce cardioprotection against ischemia. However, the data on postconditioning of the spinal cord is very limited. We investigated here whether co-application of ischemic preconditioning (IPC) and postconditioning can provide additive neuroprotection against prolonged spinal cord ischemia. Spinal cord ischemia was produced in rabbits by infrarenal aortic occlusion with a balloon catheter for 30 min. The four treatment groups were control (n=10): no intervention; IPC (n=10): a 5-minute aortic occlusion was performed 20 min before the prolonged ischemia; Postcon (n=10): postconditioning comprised of four cycles of 1-minute occlusion/1-minute reperfusion was applied one minute after the start of reperfusion. IPC+postcon (n=11): both IPC and postconditioning were applied. Functional evaluation with Tarlov score was performed during a 14-day observation period. Neurologic impairment was noticeably attenuated in the IPC+postcon group (compared with the control group, P<0.01, at day 1, day 2, day 7 and day 14, respectively), but not in either the IPC or Postcon group. Plasma malondialdehyde levels after reperfusion were significantly decreased to a similar extent in the IPC, Postcon and IPC+Postcon groups (compared with the control group (P<0.01). In the IPC+Postcon group, many more large motor neurons were preserved than in the control group (P<0.05) and white matter injury was also markedly attenuated as evidenced by reduction of the vacuolation area of the white matter (P<0.01) and decreased amyloid precursor protein immunoreactivity (P<0.01). From this, we conclude that the combination of IPC and postconditioning induces additive neuroprotective effects for spinal cord against ischemia and reperfusion injuries.     

δ-阿片受体在缺血后处理心肌保护中的作用

目的：探讨δ-阿片受体是否参与缺血后处理对抗心肌缺血/复灌（I/R）损伤和心肌细胞低氧/复氧（H/R）损伤作用及其机制。方法：采用离体大鼠心脏Langendorff灌流模型,全心停灌30 min、复灌120 min复制I/R模型。测定心室力学指标和复灌时冠脉流出液中乳酸脱氢酶（lactate dehydrogenase,LDH）活性,实验结束测定心肌组织formazan含量。酶解分离的心肌细胞采用低氧60min、复氧60min复制H/R模型,测定心肌细胞存活率。结果：在离体心脏模型上,与I/R组相比,缺血后处理组（停灌后复灌即刻立即给予6次全心停灌/复灌循环）心肌组织的formazan含量明显增高,复灌期间冠脉流出液中LDH明显降低,同时缺血后处理明显改善心室力学指标,缓解冠脉流量的减少 在分离心肌细胞模型上,低氧后处理明显提高心肌细胞存活率。δ-阿片受体阻断剂naltrindole（NTI）和线粒体钙激活钾通道（KCa）阻断剂paxilline（Pax）在离体大鼠心脏模型和分离心肌细胞模型上均能明显减弱缺血后处理的作用。在心肌细胞模型上,与H/R组相比,δ-阿片受体激动剂DADLE明显提高心肌细胞存活率,其作用可被paxilline所阻断。结论：缺血后处理具有抗心肌缺血/复灌损伤的作用,这种保护作用可能与其激活δ-阿片受体和开放KCa有关。     

Role of P2X7 purinoceptors in neuroprotective mechanism of ischemic postconditioning in mice

Cerebral ischemia–reperfusion (I–R) injury is one of the primary causes of ischemic stroke. Ischemic postconditioning (iPoCo) is evolving as an important adaptive technique to contain I–R injury. Some recent studies have shown neuroprotective effect of iPoCo. However, neuroprotective mechanism of iPoCo is not clear. So, the present study has been undertaken to investigate the possible role of P2X7 purinoceptors in neuroprotective mechanism of iPoCo in mice. Bilateral carotid artery occlusion for 12 min followed by R for 24 h produced a significant rise in cerebral infarct size and neurological severity score (NSS) along with impairment of memory and motor coordination. iPoCo, involving three episodes of 10-s carotid artery occlusion with intermittent R of 10 s applied just after ischemic insult of 12 min, produced a significant decrease in cerebral infarct size and NSS along with reversal of I–R-induced impairment of memory and motor coordination. iPoCo induced neuroprotective effects were significantly abolished by pretreatment with selective purinergic P2X7 receptor blocker Brilliant Blue G (40 mg/kg intraperitoneal). It may be concluded that neuroprotective effect of iPoCo probably involves in activation of purinergic P2X7 receptors.     

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.     

Neuroprotective effects of preconditioning ischemia on ischemic brain injury through down-regulating activation of JNK1/2 via N-methyl-D-aspartate receptor-mediated Akt1 activation

Our previous studies have demonstrated that the JNK signaling pathway plays an important role in ischemic brain injury and is mediated via glutamate receptor 6. Others studies have shown that N-methyl-d-aspartate (NMDA) receptor is involved in the neuroprotection of ischemic preconditioning. Here we examined whether ischemic preconditioning down-regulates activation of the mixed lineage kinase-JNK signaling pathway via NMDA receptor-mediated Akt1 activation. In our present results, ischemic preconditioning could not only inhibit activations of mixed lineage kinase 3, JNK1/2, and c-Jun but also enhanced activation of Akt1. In addition, both NMDA (an agonist of NMDA receptor) and preconditioning showed neuroprotective effects. In contrast, ketamine, an antagonist of NMDA receptor, prevented the above effects of preconditioning. Further studies indicated that LY294002, an inhibitor of phosphoinositide 3-kinase that is an upstream signaling protein of Akt1, could block neuroprotection of preconditioning, and KN62, an inhibitor of calmodulin-dependent protein kinase, also achieved the same effects as LY294002. Therefore, both phosphoinositide 3-kinase and calmodulin-dependent protein kinase are involved in the activation of Akt1 in ischemic tolerance. Taken together, our results indicate that preconditioning can inhibit activation of JNK signaling pathway via NMDA receptor-mediated Akt1 activation and induce neuroprotection in hippocampal CA1 region.     

No ischemic preconditioning in heterozygous connexin43-deficient mice

Protein kinase Cepsilon (PKCepsilon) plays a central role in ischemic preconditioning (IP) in mice and rabbits, and activated PKCepsilon colocalizes with and phosphorylates connexin43 (Cx43) in rats and humans. Whether or not Cx43 contributes to the mechanism(s) of IP in vivo is yet unknown. Therefore, wild-type (n = 8) and heterozygous Cx43-deficient mice (n = 8) were subjected to 30 min occlusion and 120 min reperfusion of the left anterior descending coronary artery. IP was induced by one cycle of 5 min occlusion and 10 min reperfusion (n = 8/8 mice) before the sustained occlusion. Infarct size was reduced by IP in wild-type mice [11.3 +/- 3.4% vs. 23.7 +/- 7.2% of the left ventricle (LV), P < 0.05] but not in Cx43-deficient mice (26.0 +/- 6.0% vs. 25.1 +/- 3.8% of LV). Also, three cycles of 5 min occlusion and 10 min reperfusion (n = 5) did not induce protection in Cx43-deficient mice (27.6 +/- 5.5 % of LV). Thus Cx43 contributes to the protection of IP in mice in vivo.     

Myocardial ischemic postconditioning against ischemia-reperfusion is impaired in ob/ob mice

Ischemic postconditioning (IPCD) significantly reduces infarct size in healthy animals and protects the human heart. Because obesity is a major risk factor of cardiovascular diseases, the effects of IPCD were investigated in 8- to 10-wk-old leptin-deficient obese (ob/ob) mice and compared with wild-type C57BL/6J (WT) mice. All animals underwent 30 min of coronary artery occlusion followed by 24 h of reperfusion associated or not with IPCD (6 cycles of 10-s occlusion, 10-s reperfusion). Additional mice were killed at 10 min of reperfusion for Western blotting. IPCD reduced infarct size by 58% in WT mice (33+/-1% vs. 14+/-3% for control and IPCD, respectively, P<0.05) but failed to induce cardioprotection in ob/ob mice (53+/-4% vs. 56+/-5% for control and IPCD, respectively). In WT mice, IPCD significantly increased the phosphorylation of Akt (+77%), ERK1/2 (+41%), and their common target p70S6K1 (+153% at Thr389 and +57% at Thr421/Ser424). In addition, the phosphorylated AMP-activated protein kinase (AMPK)-to-total AMPK ratio was also increased by IPCD in WT mice (+64%, P<0.05). This was accompanied by decreases in phosphatase and tensin homolog deleted on chromosome 10 (PTEN), MAP kinase phosphatase (MKP)-3, and protein phosphatase (PP)2C levels. In contrast, IPCD failed to increase the phosphorylation state of all these kinases in ob/ob mice, and the level of the three phosphatases was significantly increased. Thus, although IPCD reduces myocardial infarct size in healthy animals, its cardioprotective effect vanishes with obesity. The lack of enhanced phosphorylation by IPCD of Akt, ERK1/2, p70S6K1, and AMPK might partly explain the loss of cardioprotection in this experimental model of obese mice.     

Sumoylation of sodium/calcium exchanger in brain ischemia and ischemic preconditioning

The small ubiquitin-like modifier (SUMO) conjugation (or SUMOylation) is a post-translational protein modification mechanism activated by different stress conditions that has been recently investigated in experimental models of cerebral ischemia. The expression of SUMOylation enzymes and substrates is not restricted to the nucleus, since they are present also in the cytoplasm and on plasma membrane and are involved in several physiological and pathological conditions.In the last decades, convincing evidence have supported the idea that the increased levels of SUMOylated proteins may induce tolerance to ischemic stress. In particular, it has been established that protein SUMOylation may confer neuroprotection during ischemic preconditioning.Considering the increasing evidence that SUMO can modify stability and expression of ion channels and transporters and the relevance of controlling ionic homeostasis in ischemic conditions, the present review will resume the main aspects of SUMO pathways related to the key molecules involved in maintenance of ionic homeostasis during cerebral ischemia and ischemic preconditioning, with a particular focus on the on Na⁺/Ca²⁺ exchangers.