Sustained Ischemia Research Articles

The Protective Effect of Ischemic Preconditioning Associated with Altered Gene Expression Profiles in Rat Lung after Reperfusion

Ischemia-reperfusion (IR) injury remains an important clinical problem after lung transplantation. It is well-known that ischemic preconditioning (IPC) markedly enhances the ability of organs to withstand a sustained ischemia and reperfusion injury. But the molecular mechanisms of this protective effect are poorly understood. Therefore, we used gene microarrays to profile gene expression patterns involved in ischemia precondition. Three groups of rats were studied: IR group (IR 0, 1, 3, 6, 24, n = 5 each group, IPC group (IP 1, IP 3, 6, 24 h, n = 5 each group) and Sham group (n = 5). Samples were collected from different groups. The RatRef-12 expression Beadchip (22,226 gene probes per array) was used to analyze the pattern of gene expression in all groups. Microarray analysis showed that 648, 340, 711, 1279, and 641 genes were differentially expressed in IR 0-, 1-, 3-, 6- and 24-h groups, respectively. The profile revealed that IPC induced significant alterations of expression of many genes encoding inflammatory associated factors, oxidation and antioxidant molecules, apoptosis regulatory protein, metabolic enzyme, and ion channel protein. The damage and repair after lung IR are a dynamic process. IPC affects gene expression profiles in IR lung tissue mainly within 6 h, and sustains until 24 h later. IPC reduces lung IR injury mainly through anti-inflammatory response, antioxidative stress, and regulating cell energy metabolism and ion channels.

Acute memory phase of sevoflurane preconditioning is associated with sustained translocation of protein kinase C-α and ϵ, but not δ, in isolated guinea pig hearts

Anaesthetic preconditioning (APC) exerts cardioprotective effects by reducing infarct size and improving recovery of contractile function after ischaemia-reperfusion. The interval between brief exposure to volatile anaesthetic and sustained ischaemia, the acute memory phase, is dependent on intracellular signalling mediating this cardioprotection. Intramyocyte translocation of protein kinase C (PKC) is known to be a key mediator in APC. We examined the relationship between the time frame of the acute memory phase of sevoflurane preconditioning and intramyocyte translocation of PKC-alpha, delta and epsilon to the particulate fraction. Isolated perfused guinea pig hearts were subjected to 30 min ischaemia and 120 min reperfusion. APC was elicited with one minimum alveolar concentration sevoflurane for 10 min. Washout times of 10, 30, 60 and 90 min were studied. Contractile recovery was assessed by monitoring left ventricular developed pressures. Infarct size was determined by triphenyltetrazolium chloride staining. Translocation of PKC was examined by western blot analysis. After ischaemia-reperfusion, left ventricular developed pressure recovered to a greater degree with APC compared with control for washout times of 10 and 30 min, but not 60 and 90 min. Similarly, infarct size was reduced for washout times of 10 and 30 min, but not 60 and 90 min. Sustained translocation of PKC-alpha and epsilon, but not delta, was associated with the time frame of the acute memory phase. The acute memory phase of sevoflurane preconditioning is limited to less than 60 min. Sustained translocation of PKC-alpha and epsilon, but not delta, correlates with this acute memory phase of sevoflurane preconditioning.

Altitude-Induced Changes in Muscle Contractile Properties

Because of its high energetic demand, skeletal muscle is sensitive to changes in the partial pressure of oxygen. Most human studies on in vivo skeletal muscle function during hypoxia were performed with voluntary contractions. However, skeletal muscle function is not only characterized by voluntary maximal or repeated force- generating capacity, but also by force generated by evoked muscle contractions (i.e., force-frequency properties). This mini-review reports on the effects of acute or prolonged exposure to hypoxia on human skeletal muscle performance and contractile properties. The latter depend on both the amount and type of contractile proteins and the efficiency of the cellular mechanism of excitation-contraction coupling. Observations on humans indicate that hypoxia (during simulated ascent or brief exposure) exerts modest influences on the membrane propagation of the muscle action potentials during voluntary contractions. Overall in humans, in physiological conditions, including that of climbing Mt. Everest, there is extraordinarily little that changes with regard to maximal force-generating capacity. Interestingly, it appears that the adaptations to chronic hypoxia minimize the effects on skeletal muscle dysfunction (i.e., impairment during fatigue resistance exercise and in muscle contractile properties) that may occur during acute hypoxia for some isolated muscle exercises. Only sustained isometric exercise exceeding a certain intensity (30% MVC) and causing substantial and sustained ischemia is not affected by acute hypoxia.

Postconditioning during primary percutaneous coronary intervention: A review and meta-analysis

BackgroundMyocardial postconditioning (POC), defined as intermittent interruptions of blood flow at the onset of reperfusion after sustained ischemia, may attenuate reperfusion injury. In order to weigh the current evidence linking POC to reduction of myocardial infarct size in patients with ST-segment elevation myocardial infarction (STEMI) undergoing primary percutaneous coronary intervention (pPCI), we performed a systematic review and meta-analysis of the available data from small randomised trials. MethodsA Web-based search was performed for relevant studies, and retrieved data were subjected to systematic review and meta-analysis. ResultsSix studies were identified and the meta-analysis included a total of 244 patients with STEMI undergoing pPCI with or without POC. Pooled analysis of all studies demonstrated a significant reduction of peak creatine kinase with POC relative to standard care (weighted mean peak creatine kinase difference −609.59 IU/L, 95% confidence interval [CI] −1030 to −189; p=0.005). A secondary analysis of 4 of the studies also showed an improvement of left ventricular ejection fraction with POC (weighted mean ejection fraction difference 4.2%, 95% CI 2.1% to 6.2%; p=0.0001). ConclusionThis first systematic review and meta-analysis of randomized trials of POC in patients with STEMI undergoing primary PCI, demonstrated a significant benefit of POC over standard care for reduction of myocardial infarct size as determined by peak creatine kinase release and left ventricular ejection fraction. The effects of POC on clinical outcomes remain to be determined.

Dexamethasone-induced cardioprotection: A role for the phosphatase MKP-1?

Aims Previous studies suggested that p38 MAPK activation during sustained myocardial ischaemia and reperfusion was harmful. We hypothesize that attenuation of p38MAPK activity via dephosphorylation by the dual-specificity phosphatase MKP-1 should be protective against ischaemia/reperfusion injury. Since the glucocorticoid, dexamethasone, induces the expression of MKP-1, the aim of this study was to determine whether upregulation of this phosphatase by dexamethasone protects the heart against ischaemia/reperfusion injury. Main methods Male Wistar rats were treated with dexamethasone (3 mg/kg/day ip) for 10 days, before removal of the hearts for Western blot (ip Dex − P) or perfusion in the working mode (ip Dex + P). Hearts were subjected to 20 min global or 35 min regional ischaemia (36.5 °C) and 30 or 120 min reperfusion. In a separate series, dexamethasone (1 µM) was added to the perfusate for 10 min (Pre + Dex) before or after (Rep + Dex) ischaemia. Key findings Dexamethasone, administered intraperitoneally or added directly to the perfusate, significantly improved post-ischaemic functional recovery and reduced infarct size compared to untreated controls ( p < 0.05). These were associated with enhanced up-regulation of MKP-1 protein expression (arbitrary units (mean ± SD): Untreated: 1; ip Dex − P: 2.59 ± 0.22; ip Dex + P: 1.51 ± 0.22; Pre + Dex: 4.11 ± 0.73, Rep + 15′Dex: 1.51 ± 0.14; untreated vs. all groups, p < 0.05) and attenuation of p38 MAPK activation ( p < 0.05) in all dexamethasone-treated groups, except for Rep + 10′Dex. ERK and PKB/Akt activation were unchanged. Significance Dexamethasone-induced cardioprotection was associated with upregulation of the phosphatase MKP-1 and inactivation of pro-apoptotic p38 MAPK.

Double-Blind Treatment of Apathy in Patients with Poststroke Depression Using Nefiracetam

Nefiracetam is a novel pyrrolidone-type nootropic compound shown in preliminary trials to increase blood flow and improve patient outlook and energy following stroke. Of 137 stroke patients with major depression, 70 also met published diagnostic criteria for apathy (51.1%) and were randomly assigned either to placebo or 600 mg or 900 mg of nefiracetam per day, and received at least 4 weeks of treatment. Using the group with at least 4 weeks of treatment as the intention-to-treat sample with last observation carried forward, repeated measures analysis of variance of Apathy Scale scores demonstrated a significant time-by-treatment interaction. Patients taking 900 mg nefiracetam had a significantly greater change in Apathy Scale scores compared to 600 mg of nefiracetam or placebo. Future studies should assess whether apathy without depression may respond to this novel treatment.

The Rocky Road from Bench to Bedside

The Rocky Road from Bench to Bedside

The Strategy of Combined Ischemia Preconditioning and Salvianolic Acid-B Pretreatment to Prevent Hepatic Ischemia-Reperfusion Injury in Rats

Ischemia-reperfusion injury (IRI) is a serious complication of liver surgery, especially for extended hepatectomy and liver transplantation. The aim of this study was to evaluate the protective effect of combined ischemic preconditioning (IPC) and salvianolic acid-B (Sal-B) pretreatment against IRI-induced hepatocellular injury. Sixty male Wistar rats weighing around 200 g were randomized into five groups (n=12): sham group: only anesthesia and laparotomy; IR group: 90 min sustained ischemia by blocking the left ortal vessels; IPC group: 10 min ischemia and 10 min reperfusion prior to the sustained ischemia; Sal-B group: 10 mg/kg injection of Sal-B intravenously 10 min prior to the sustained ischemia; IPC+Sal-B group: same IPC procedure as in IPC group, but proceeded by intravenous administration of Sal-B 10 min prior to sustained ischemia. After 5 h of reperfusion, serum levels of ALT and AST were measured; the amount of malondialdehyde (MDA) and adenine nucleotides in liver tissue was determined; the expression of Bcl-2 and caspase-3 was detected by immunofluorescent and western blotting techniques; the severity of apoptosis and pathological alterations was evaluated by TUNEL and H&E staining, respectively. The serum aminotransferases, hepatic MDA concentration, and apoptotic index in groups IPC, Sal-B, and IPC+Sal-B were significantly lower than those in the IR group (P<0.001), while the IPC+Sal-B group had the lowest values among these groups (P<0.05). Compared with the IR group, groups IPC and Sal-B not only had statistically higher ATP levels and energy charge (EC) values (P<0.01), but also had upregulated Bcl-2 expression and downregulated cleaved caspase-3 expression in liver tissue. All these effects were further augmented in the IPC+Sal-B group. Liver histopathological findings were consistent with these results. Based on these results, the combined IPC and Sal-B pretreatment had a synergistically protective effect on liver tissue against IRI, which might be due to decreased post-ischemic oxidative stress, improved energy metabolism, and reduced hepatocellular apoptosis.

Intestinal Preconditioning Ameliorates Ischemia-Reperfusion Induced Acute Lung Injury in Rats: An Experimental Study

Phospholipases A(2) (PLA(2)) have been implicated in the pathogenesis of acute respiratory distress syndrome (ARDS) induced by intestinal ischemia-reperfusion (IIR). Intestinal ischemic preconditioning (IIP) has been shown to improve intestinal tolerance to subsequent sustained ischemia and limit the systemic inflammatory response. We tested the effect of IIP on the intestinal ischemia-reperfusion-induced ARDS, with particular focus on PLA(2). Rats were randomized into three groups: (1) sham surgery group (sGroup), 45 min sham intestinal ischemia-4 h reperfusion, (2) IIR group (IIRGroup), 45 min intestinal ischemia-4 h reperfusion, (3) IIP group (ipGroup), three cycles of intestinal ischemia for 4 min and reperfusion for 10 min followed by 45 min intestinal ischemia-4 h reperfusion. At the end of each experiment, blood gases were obtained and bronchoalveolar lavage (BAL) followed. Biochemical (total protein, PLA(2), PAF-AcH) and cytological parameters of the BAL fluid were quantified. Plasma MDA was measured as an indicator of systemic oxidative stress. Comparisons between groups were made using one-way ANOVA followed by post hoc comparison with a Tukey test or Mann-Whitney test when appropriate. Differences were considered significant if P < 0.05. Alveolar-arterial O(2) gradient values and wet to dry lung ratio were significantly (P < 0.05) increased in the IIRGroup and this increase was prevented in the ipGroup. Following the same pattern, BAL total protein, PLA(2), and PAF-AcH were significantly lower in the ipGroup. Ischemic preconditioning significantly abolished neutrophil count in BAL fluid. Plasma MDA was significantly lower in the ipGroup. Despite a significant tissue polymorphonuclear reduction, no significant lung or intestinal histologic damage score changes were revealed. Intestinal preconditioning protects IIR-induced lung injury, partly by modulating the arachidonic acid cascade.

Extraction of Erythrocyte Enzymes for the Preparation of Polyhemoglobin-catalase-superoxide Dismutase

In sustained severe ischemia, reperfusion with oxygen carriers may result in ischemia-reperfusion injuries because of the release of damaging oxygen radicals. A nanobiotechnology-based polyhemogloin-calatase-superoxide dismutase can prevent this because the oxygen carrier, polyhemoglobin, is linked to antioxidant enzymes, catalase and superoxide dismutase. However, these antioxidant enzymes come from nonhuman sources and recombinant human enzymes are expensive. This paper describes our study on extracting these enzymes from red blood cells and analyzing the amount of enzymes needed for adequate protection from ischemia-reperfusion.

Functional protection of pentoxifylline against spinal cord ischemia/reperfusion injury in rabbits: necrosis and apoptosis effects

Little is known about neuronal death mechanisms following spinal cord ischemia. The present study aimed to investigate the protective effect of pentoxifylline (PTX) against spinal cord ischemia/reperfusion (I/R) injury. Rabbits sustained spinal cord ischemia following 45 minutes cross-clamping of the infrarenal aorta. Experimental groups were as follows: the first group of animals (sham, n = 8) underwent laparotomy alone and served as the sham group; the second group (I/R, n = 20) received carrier (3 ml saline solution) and served as the control group; the third group (PTX-A, n = 20) received PTX intravenously 10 minutes prior to ischemia; and the fourth group (PTX-B, n = 20) received PTX intravenously at the onset of reperfusion. Rabbits were evaluated for hind-limb motor function with the Tarlov scoring system at 48 hours. Serum was assayed with enzyme-linked immunosorbent assay for tumor necrosis factor alpha (TNF-alpha) and spinal cords were harvested for myeloperoxidase (MPO) activity, histopathological analysis, terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling staining, platelet/endothelial cell adhesion molecule-1 (PECAM-1) and caspase-3 immunohistochemistry, and the number of necrotic and apoptotic neuron were counted and data analyzed at 12, 24, 48 and 72 hours of reperfusion. Spinal cords were studied by electron microscopy. Improved Tarlov scores were seen in PTX-treated rabbits as compared with ischemic control rabbits at 48 hours. A significant reduction was found in TNF-alpha in serum, activity of MPO and immunoreactivity of the PECAM-1 and caspase-3 in PTX-treated rabbits. There were fewer apoptotic neurons than necrotic neurons (P < 0.05). A significant decrease in both necrotic and apoptotic neurons was observed in the PTX-treated groups (PTX-A and PTX-B) compared with the I/R group (P < 0.05). Both necrotic and apoptotic neurons were found with the electron microscope. PTX may induce protection against ischemia injury in the spinal cord, thereby preventing both necrosis and apoptosis. A major mode of cell death in spinal cord ischemia/reperfusion injury is necrosis while apoptosis is not dominant.

Na+/H+exchange inhibitor cariporide attenuates skeletal muscle infarction when administered before ischemia or reperfusion

Administration of Na(+)/H(+) exchange isoform-1 (NHE-1) inhibitors before ischemia has been shown to attenuate myocardial infarction in several animal models of ischemia-reperfusion injury. However, controversy still exists as to the efficacy of NHE-1 inhibitors in protection of myocardial infarction when administered at the onset of reperfusion. Furthermore, the efficacy of NHE-1 inhibition in protection of skeletal muscle from infarction (necrosis) has not been studied. This information has potential clinical applications in prevention or salvage of skeletal muscle from ischemia-reperfusion injury in elective and trauma reconstructive surgery. The objective of this research project is to test our hypothesis that the NHE-1 inhibitor cariporide is effective in protection of skeletal muscle from infarction when administered at the onset of sustained ischemia or reperfusion and to study the mechanism of action of cariporide. In our studies, we observed that intravenous administration of cariporide 10 min before ischemia (1 or 3 mg/kg) or reperfusion (3 mg/kg) significantly reduced infarction in pig latissimus dorsi muscle flaps compared with the control, when these muscle flaps were subjected to 4 h of ischemia and 48 h of reperfusion (P < 0.05; n = 5 pigs/group). Both preischemic and postischemic cariporide treatment (3 mg/kg) induced a significant decrease in muscle myeloperoxidase activity and mitochondrial-free Ca(2+) content and a significant increase in muscle ATP content within 2 h of reperfusion (P < 0.05; n = 4 pigs/group). Preischemic and postischemic cariporide treatment (3 mg/kg) also significantly inhibited muscle NHE-1 protein expression within 2 h of reperfusion after 4 h of ischemia, compared with the control (P < 0.05; n = 3 pigs/group). These observations support our hypothesis that cariporide attenuates skeletal muscle infarction when administered at the onset of ischemia or reperfusion, and the mechanism involves attenuation of neutrophil accumulation and mitochondrial-free Ca(2+) overload and preservation of ATP synthesis in the early stage of reperfusion.

Inducible Nitric Oxide Synthase Expression and Cardiomyocyte Dysfunction During Sustained Moderate Ischemia in Pigs

In acute myocardial ischemia, regional blood flow and function are proportionally reduced. With prolongation of ischemia, function further declines at unchanged blood flow. We studied the involvement of an inflammatory signal cascade in such progressive dysfunction and whether dysfunction is intrinsic to cardiomyocytes. In 10 pigs, ischemia was induced by adjusting inflow into the cannulated left anterior coronary artery to reduce coronary arterial pressure to 45 mm Hg (ISCH); 4 pigs received the inducible nitric oxide synthase (iNOS) inhibitors aminoguanidine or L-N(6)-(1-iminoethyl)-lysine during ISCH (ISCH+iNOS-Inhib); 6 pigs served as controls (SHAM). Anterior (AW) and posterior (PW) systolic wall thickening (sonomicrometry) were measured. After 6 hours, nitric oxide (NO) synthase (NOS) protein expression, NOS activity, and NO metabolites (nitrite/nitrate/nitroso species) were quantified in biopsies isolated from AW and PW. Cardiomyocyte shortening and intracellular calcium (Indo-1 acetoxymethyl ester) were measured without and with the NOS substrate L-arginine (100 micromol/L). In ISCH, AW wall thickening decreased from 42+/-4% (baseline) to 16+/-3% (6 hours). Wall thickening remained unchanged in ISCH-PW and SHAM-AW/PW. NOS2 (iNOS) protein expression and activity, but not NOS3 (endothelial NO synthase), were increased in ISCH-AW and ISCH-PW. iNOS expression correlated with increased nitrite contents. Cardiomyocyte shortening was reduced in ISCH-AW versus SHAM-AW (4.4+/-0.3% versus 5.6+/-0.3%). L-Arginine reduced cardiomyocyte shortening further in ISCH-AW (to 2.8+/-0.2%) and ISCH-PW (3.4+/-0.4% versus 5.4+/-0.4%) but not in SHAM or in ISCH+iNOS-Inhib; intracellular [Ca(2+)] remained unchanged. With L-arginine, in vitro AW cardiomyocyte shortening correlated with in vivo AW wall thickening (r=0.72). In conclusion, sustained regional ischemia induces myocardial iNOS expression in pigs, which contributes to contractile dysfunction at the cardiomyocyte level.

Signalosomes: delivering cardioprotective signals from GPCRs to mitochondria

Signalosomes: delivering cardioprotective signals from GPCRs to mitochondria

Ischemic preconditioning accelerates the fatty acid oxidation of rat hearts

Background Ischemic preconditioning (IPC) reduced myocardial ATP depletion during sustained ischemia and has a powerful protective effect on the myocardium. The purpose of the present study was to clarify the effects of IPC on myocardial accumulation of fatty acid (FA) tracer and its intracellular metabolism. Methods Myocardial ischemia-reperfusion (MI-R) injury was induced by the left coronary artery ligation for 15 min followed by reperfusion in Wistar rats. IPC was achieved with a single cycle of 5-minute coronary ligation followed by 5-minute reperfusion before MI-R. Three days after ischemia-reperfusion, FA metabolism was evaluated in rats with or without IPC using 131I- and 125I-15-( p-iodophenyl)-9-methylpentadecanoic acid (9MPA) and thin-layer chromatography. Results IPC attenuated a reduction of 9MPA accumulation in ischemic region (IR). The metabolite fraction of 9MPA including both early and late metabolites was less in IR as compared to non-IR in rats without IPC. IPC increased the final metabolic product of 9MPA processed via α- and β-oxidation in both IR and non-IR. Conclusions IPC accelerated fatty acid oxidation in both IR and non-IR. This alteration in fatty acid metabolism would inhibit an intracellular accumulation of detrimental fatty acid metabolites.

Effects of hydrogen sulphide on ischaemia–reperfusion injury and ischaemic preconditioning in the isolated, perfused rat heart☆

Hydrogen sulphide (H(2)S) protects the heart against ischaemia-reperfusion injury caused by low flow or local ischaemia. We hypothesised that: (1) H(2)S protects against global ischaemia-reperfusion injury of the heart, (2) H(2)S plays a mechanistic role in ischaemic preconditioning, and (3) H(2)S acts by phosphorylation of protein kinases. Isolated, perfused rat hearts were used in two series. Series 1: group 1.1 (n=10), 40 min of ischaemia and 120 min of reperfusion, group 1.2 (n=7), like 1.1 except that 40 microM NaHS was added to the perfusate during stabilisation and throughout the experiment. Group 1.3 (n=10), like 1.1, but endogenously produced H(2)S was blocked by D,L-propargylglycine. Series 2: group 2.1 (n=10) control, 30 min of ischaemia followed by 120 min of reperfusion. Group 2.2 (n=10) ischaemic preconditioning before sustained ischaemia and 120 min of reperfusion. Group 2.3 (n=10) like 2.2 except of D,L-propargylglycine treatment like in group 1.3. Mitogen activated protein kinases including extracellular signal-regulated kinases (ERK 1/2), the stress-activated/c-Jun NH2 terminal kinases (JNK), P38, as well as protein kinase B/AKT (AKT), adenosine monophosphate dependent protein kinase (AMPK) and the inducible heat shock protein 72 were measured by Western blotting. Adenine nucleotides (ATP, ADP, and AMP) were measured by high-pressure liquid chromatography and energy charge was calculated. Infarct size was increased by D,L-propargylglycine (40+/-6 vs 27+/-10% in controls, p=0.03, Bonferroni post hoc test). There was a non-significant decrease in infarct size in the NaHS group (to 20+/-13%). Western blot analysis indicated an upregulation of heat shock protein 72 in the NaHS treated group and a reduced phosphorylation of AKT in the D,L-propargylglycine group. D,L-propargylglycine had no effect on ischaemic preconditioning or on phosphorylation of protein kinases (ERK, AKT, P38, JNK and AMPK) in preconditioned hearts. No difference in energy charge was found between groups, although ADP was increased in the NaHS-treated group. Endogenous H(2)S production protects against global ischaemia, and H(2)S may be a part of the endogenous cell defence. However, endogenous H(2)S did not appear to be important in ischaemic preconditioning, and protein kinases were not important for the effect of H(2)S. Exogenous H(2)S may provide myocardial protection, possibly acting by expression of heat shock protein 72.

Injection of neural progenitor cells improved learning and memory dysfunction after cerebral ischemia

Accumulating evidence indicates that stem cells have the ability to improve neurological deficits seen after cerebral ischemia. However, the effects of neural progenitor cells (NPCs) on cerebral ischemia-induced learning and memory dysfunction remain to be clarified. The purpose of the present study was to determine whether the injection of exogenous NPCs could prevent learning and memory dysfunction after cerebral ischemia. Sustained cerebral ischemia was produced by the injection of 700 microspheres into the right hemisphere of each rat. We demonstrated that injection of NPCs into the hippocampus at 10 min after the induction of cerebral ischemia reduced prolongation of the escape latency seen in acquisition and retention tests of the water maze task on Days 12–28 after cerebral ischemia. Injection of NPCs partially attenuated the decrease in viable areas of the ipsilateral hemisphere on Day 28 after the cerebral ischemia. We also demonstrated that injection of NPCs prevented the decrease in the level of BDNF seen at the early period after cerebral ischemia. These results suggest that the injection of exogenous NPCs into the hippocampus can prevent cerebral ischemia-induced learning and memory dysfunction, possibly through maintenance of the BDNF level.

Remote periconditioning reduces myocardial no-reflow by the activation of K ATP channel via inhibition of Rho-kinase

Remote periconditioning is induced by brief cycles of ischemia and reperfusion of a remote organ applied during sustained myocardial ischemia. It remains unknown whether the remote periconditioning reduces myocardial no-reflow. The adenosine triphosphate-sensitive potassium (K ATP) channel opening and inhibition of Rho-kinase may be the important mechanism of protection against myocardial no-reflow. Therefore, this study was sought to assess the effect of remote periconditioning on myocardial no-reflow and explore the possible mechanism. Methods Coronary ligation area and area of no-reflow were determined with pathological means in 58 mini-swines randomized into 7 study groups: 9 controls, 8 in remote periconditioning, 8 in hydroxyfasudil (a specific inhibitor of Rho-kinase)-treated, 9 in glibenclamide (K ATP channel blocker)-treated, 8 in remote periconditioning and glibenclamide, 8 in hydroxyfasudil and glibenclamide and 8 sham-operated. The ischemia and reperfusion model was created with 3 h of left anterior descending artery occlusion followed by 2 h of reperfusion. Results Compared with the control group, remote periconditioning decreased Rho-kinase activity ( P < 0.01), increased coronary blood volume ( P < 0.05), decreased area of no-reflow (from 82.3 ± 3.9% to 45.5 ± 5.7% of ligation area, P < 0.01) and reduced necrosis size (from 98.5 ± 1.3% to 74.7 ± 6.3% of ligation area, P < 0.05). Hydroxyfasudil had the same effect on the above parameters as remote periconditioning. Glibenclamide abrogated the effect of remote periconditioning or hydroxyfasudil on area of no-reflow and necrosis area, but not Rho-kinase activity. Conclusion Remote periconditioning can reduce myocardial no-reflow after ischemia and reperfusion. This beneficial effect could be due to its activation of K ATP channel via inhibition of Rho-kinase.

CREB Activation and Ischaemic Preconditioning

Previous studies from our laboratory showed that activation of p38 MAPK is one of the triggers of ischaemic preconditioning. The signalling events downstream of p38 MAPK and their links to the putative final effectors of preconditioning are not clear. The cAMP responsive element-binding protein (CREB) is also phosphorylated by exposure to short episodes of ischaemia/reperfusion, suggesting a triggering action. The aim of this study was to systematically investigate (1) the signalling pathways leading to CREB phosphorylation during an ischaemic or beta-adrenergic preconditioning protocol (2) changes in CREB phosphorylation during sustained ischaemia and their significance in ischaemia/reperfusion injury. The isolated perfused working rat heart was preconditioned by 1 x 5 min global ischaemia or 3 x 5 min global ischaemia and freeze-clamped. Drugs to manipulate CREB activation were added 5 min before onset of ischaemia. Non-preconditioned and preconditioned hearts were subjected to 25 min global or 35 min regional ischaemia, followed by 30 min reperfusion. Infarct sizes were determined using tetrazolium staining. Phosphorylation of CREB was determined by Western blots. Exposure of hearts to 5 min global ischaemia followed by reperfusion, significantly increased CREB phosphorylation This is mediated by, amongst others, release of endogenous catecholamines and adenosine, as indicated by the use of receptor blockers. Events downstream of receptor stimulation were evaluated using inhibitors for PKA (H89), MSK-1 (H89, Ro318220), PKC (bisindolylmaleimide), p38 MAPK (SB203580) and ERK (PD98059). Activation of PKA, PKC, ERK and p38 MAPK is involved in preconditioning-induced CREB phosphorylation. Ischaemia-induced activation of iPLA(2) and cPLA(2) also contribute to CREB phosphorylation as indicated by the use of the inhibitors 4-bromo-enol-lactone (BEL) and AACOF(3,) respectively. Inhibition of CREB phosphorylation by either BEL or AACOF(3) during a preconditioning protocol partially attenuated cardioprotection. CREB phosphorylation was attenuated during sustained global ischaemia of both non-preconditioned and preconditioned hearts. These data suggest that CREB phosphorylation during an ischaemic preconditioning protocol may contribute to triggering preconditioning, while reduced phosphorylation during sustained ischaemia does not appear to be associated with cardioprotection.

Neuroprotectionby MK-801 following cerebral ischemia in Mongolian gerbils

Global cerebral ischemia in Mongolian gerbils is an established model in experimental research on cerebral ischemia, which is characterized morphologically by selective neuronal damage in the hippocampus, striatum, and cortex. Elevated glutamate levels are thought to be a primary cause of neuronal death after global cerebral ischemia. The purpose of this study was to investigate the potential neuroprotective effects of dizocilpine malate (MK-801), a non-competitive glutamate antagonist, in the model of 10-min gerbil cerebral ischemia. Gerbils were given MK-801(3 mg/kg i.p.)or saline immediately after the occlusion. On day 4 after reperfusion, neuronal damage was examined in the hippocampus (30 ?m)and striatum slices (5 ?m)stained with hematoxylin/eosin, fluorescent Nissl staining and membrane tracer DiI. The striatum and C3 regions of the hippocampus were analyzed by confocal microscopy. Neuroprotection was determined by quantifying the degree of cell loss, reduction of morphologically damaged cells, and the degree of preservation of recog?nizable neuroanatomical pathways after the ischemic insult. Our results demonstrate that the neuronal damage induced by sustained ischemia is related to abnormalities in glutamatergic function associated with NMDA receptors. MK-801significantly prevented neuronal loss in the tested brain structures. All of this contributes to a better understanding of the given pathophysiological process causing ischemic neuronal damage. &lt;br&gt;&lt;br&gt;&lt;font color="red"&gt;&lt;b&gt; This article has been corrected. Link to the correction &lt;u&gt;&lt;a href="http://dx.doi.org/10.2298/ABS160524054E"&gt;10.2298/ABS160524054E&lt;/a&gt;&lt;u&gt;&lt;/b&gt;&lt;/font&gt;