Search

Objectives: Aminosteroids of the lazaroid type protect organs from ischemia-reperfusion damage. The authors hypothesized that lazaroid U-83836E may be beneficial in a shock model with hemorrhage combined with limb ischemia. Furthermore, the authors hypothesized that lazaroids induce expression of heat shock proteins (HSPs) of the 72-kDa family., Methods: Rats were divided into two groups (lazaroid and control groups, n = 8 each) and pretreated with the lazaroid U-83836E (5 mg/kg) or with vehicle intraperitoneally at 12 and 24 hours before experiments. At the time of the experiment, rats were anesthetized, and the femoral artery of each rat was cannulated. After 20 minutes of stabilization, blood was shed from each rat to bring its mean arterial pressure to 24-28 mmHg for 2 hours. Bilateral tourniquets were tightened proximally on the rat thighs during those 2 hours and then released. Shed blood plus equal amounts of Ringer acetate then were infused to restore normal blood pressure, followed by a continuous infusion of Ringer acetate, the rate of which was regulated to maintain blood pressure, until 30 minutes after start of resuscitation. Fluid resuscitation was stopped, and rats were observed for another 3.5 hours. At the end of the observation period, the rats' hearts were collected for immunoblot analysis of HSP72. Additional hearts were collected from similarly pretreated rats not undergoing the episode of hemorrhagic shock and fluid resuscitation., Results: Pretreatment with U-83836E improved mean arterial blood pressure after hemorrhagic shock and fluid resuscitation (p = 0.02), combined with improvements in acid-base balance (improved base excess and standard bicarbonate; p = 0.02 and p = 0.01, respectively). Western blot of cardiac protein extracts demonstrated that lazaroid pretreatment increased expression of HSP72., Conclusions: Pretreatment with the lazaroid U-83836E improved outcome markers in this hemorrhagic shock model. The observed protection may be caused by increased expression of HSP72.

OBJECTIVES: Several studies have reported superior post-cardioplegic recovery after glutamate supplementation. The optimum dose of glutamate supplementation is unknown. The purpose of this study was to find the optimal protective concentration of glutamate supplementation in a model of ischaemia/cardioplegia and reperfusion. METHODS: Isolated rat hearts (n = 77) were perfused with the Krebs-Henseleit buffer. After stabilization, the hearts were subjected to 25 min of normothermic ischaemia followed by a single 3-min infusion of cold (4-6 °C) St. Thomas' Hospital II cardioplegia and 87 min of cardioplegic ischaemic arrest and 60 min of reperfusion. Sodium-L-glutamate was added to the perfusate (control group had zero glutamate) in increasing concentrations (0.01, 0.1, 1, 10, 20, 30 and 100mM) and given throughout perfusion. Corresponding concentrations were added to the cardioplegic solution. A balloon in the left ventricle inserted via the left atrium measured left ventricular pressures isometrically. Left ventricular developed pressure was calculated. Myocardial exchange of glucose and lactate was measured prior to ischaemia and during reperfusion. Myocardial content of glycogen and glutamate was measured at the end of reperfusion. RESULTS: During reperfusion left ventricular developed pressure increased (P < 0.0001) in groups supplemented with 0.1, 1.0, 10, 20 and 30mM glutamate, whereas left ventricular end-diastolic pressure was attenuated (P = 0.008) when compared with the controls. No additional benefit on the continuous data left ventricular developed pressure and left ventricular end-diastolic pressure was observed with glutamate concentrations above 1 mM. Onset of LV pressure rise during the period of ischaemia was delayed by 100mM of glutamate (P = 0.02). Myocardial content of glutamate was increased in a dose-related manner in Groups 10, 20, 30 and 100 compared with the control hearts (P < 0.0001). Glycogen was increased in the hearts supplemented with 100mM of glutamate (P = 0.02). CONCLUSIONS: Even low concentrations of L-glutamate improved postischaemic and post-cardioplegic heart function and 1mM seems to be optimal. [ABSTRACT FROM AUTHOR]

We aimed to study the cardiac expression of bone morphogenetic protein 2, its receptor 1 b, and connective tissue growth factor, factors implicated in cardiac embryogenesis, following ischemia/hypoxia, heart failure, and in remodeling hearts from humans and mice. Biopsies from the left ventricle of patients with end-stage heart failure due to dilated cardiomyopathy or coronary artery disease were compared with donor hearts and biopsies from patients with normal heart function undergoing coronary artery bypass grafting. Mouse model of post-infarction remodeling was made by permanent ligation of the left coronary artery. Hearts were analyzed by real-time polymerase chain reaction and Western blotting after 24 hours and after 2 and 4 weeks. Patients with dilated cardiomyopathy and mice post-infarction had increased cardiac expression of connective tissue growth factor. Bone morphogenetic protein 2 was increased in human hearts failing due to coronary artery disease and in mice post-infarction. Gene expression of bone morphogenetic protein receptor 1 beta was reduced in hearts of patients with failure, but increased two weeks following permanent ligation of the left coronary artery in mice. In conclusion, connective tissue growth factor is upregulated in hearts of humans with dilated cardiomyopathy, bone morphogenetic protein 2 is upregulated in remodeling due to myocardial infarction while its receptor 1 b in human failing hearts is downregulated. A potential explanation might be an attempt to engage regenerative processes, which should be addressed by further, mechanistic studies. [ABSTRACT FROM PUBLISHER]

Background: All-trans retinoic acid (atRA), an active derivative of vitamin A, regulates cell differentiation, proliferation and cardiac morphogenesis via transcriptional activation of retinoic acid receptors (RARs) acting on retinoic acid response elements (RARE).We hypothesized that the retinoic acid (RA) signalling pathway is activated in myocardial ischemia and postischemic remodelling. Methods and Findings: Myocardial infarction was induced through ligating the left coronary artery in mice. In vivo cardiac activation of the RARs was measured by imaging RARE-luciferase reporter mice, and analysing expression of RAR target genes and proteins by real time RT-PCR and western blot. Endogenous retinoids in postinfarcted hearts were analysed by triple-stage liquid chromatography/tandem mass spectrometry. Cardiomyocytes (CM) and cardiofibroblasts (CF) were isolated from infarcted and sham operated RARE luciferase reporter hearts and monitored for RAR activity and expression of target genes. The effect of atRA on CF proliferation was evaluated by EdU incorporation. Myocardial infarction increased thoracic RAR activity in vivo (p,0.001), which was ascribed to the heart through ex vivo imaging (p = 0.002) with the largest signal 1 week postinfarct. This was accompanied by increased cardiac gene and protein expression of the RAR target genes retinol binding protein 1 (p = 0.01 for RNA, p = 0,006 for protein) and aldehyde dehydrogenase 1A2 (p = 0.04 for RNA, p = 0,014 for protein), while gene expression of cytochrome P450 26B1 was downregulated (p = 0.007). Concomitantly, retinol accumulated in the infarcted zone (p = 0.02). CM and CF isolated from infarcted hearts had higher luminescence than those from sham operated hearts (p = 0.02 and p = 0.008). AtRA inhibited CF proliferation in vitro (p = 0.02). Conclusion: The RA signalling pathway is activated in postischemic hearts and may play a role in regulation of damage and repair during remodelling. [ABSTRACT FROM AUTHOR]

Aquaporins (AQPs) are channel-forming membrane proteins highly permeable to water. AQP4 is found in mammalian hearts; however, its expression sites, regulation and function are largely unknown. The aim was to investigate cardiac AQP4 expression in humans and mice, its regulation by ischemia and hypoxia, and in particular its role in cardiac ischemic injury using AQP4 knockout (KO) mice. Comparable levels of AQP4 were detected by Western blot and qPCR in biopsies from human donor hearts and wild type C57Bl6 mouse hearts. In mice, AQP4 was expressed on cardiomyocyte plasmalemma (qPCR, Western blot, immunogold), and its mRNA decreased following ischemia/reperfusion (isolated hearts, p = 0.02) and after normobaric hypoxia in vivo (oxygen fraction 10 % for 1 week, p < 0.001). Isolated hearts from AQP4 KO mice undergoing global ischemia and reperfusion had reduced infarct size ( p = 0.05) and attenuated left ventricular end-diastolic pressure during reperfusion ( p = 0.04). Infarct size was also reduced in AQP4 KO mice 24 h after left coronary artery ligation in vivo ( p = 0.036). AQP4 KO hearts had no compensatory change in AQP1 protein expression. AQP4 KO cardiomyocytes were partially resisted to hypoosmotic stress in the presence of hypercontracture. AQP4 is expressed in human and mouse hearts, in the latter confined to the cardiomyocyte plasmalemma. AQP4 mRNA expression is downregulated by hypoxia and ischemia. Deletion of AQP4 is protective in acute myocardial ischemia-reperfusion, and this molecule might be a future target in the treatment of acute myocardial infarction. [ABSTRACT FROM AUTHOR]

We tested if remote gene delivery of hypoxia-inducible factor 1 alpha (HIF-1α) protected hearts against induced ischemia, hypothesizing that gene delivery into skeletal muscle may lead to secretion of proteins with actions elsewhere. Murine quadriceps muscles were transfected with DNA encoding for human HIF-1α, which resulted in a local, but lasting expression (mRNA and protein, where the latter had nuclear localization). Subjection of isolated hearts to global ischemia and reperfusion 1, 4, and 8 weeks after gene delivery resulted in infarct size reduction ( p < 0.05). Supporting that this was due to paracrine effects, HL-1 cells treated with conditioned media from cells transfected with HIF-1α or serum from HIF-1α-treated mice were protected against H2O2-induced cell death ( p < 0.05, respectively). The latter protection was reduced when a heme oxygenase activity blocker was used. Taqman low-density array of 47 HIF-1α-regulated genes at the treatment site showed nine specific upregulations ( p < 0.05). Of the corresponding proteins, PDGF-B and adrenomedullin were upregulated in the heart. HIF-1α treatment induced an increased vascularization of the heart and skeletal muscle. In conclusion, remote delivery of DNAfor HIF-1α was cardioprotective, represented by consistent infarct size reduction, which may be due to release of paracrine factors from the transfected muscle. [ABSTRACT FROM AUTHOR]

Hyperoxic exposure in vivo (> 95% oxygen) attenuates ischemia-reperfusion injury, but the signaling mechanisms of this cardioprotection are not fully determined. We studied a possible role of nitric oxide (NO) and mitogen activated protein kinases (MAPK) in hyperoxic protection. Mice (n = 7–9 in each group) were kept in normoxic or hyperoxic environments for 15 min prior to harvesting the heart and Langendorff perfusion with global ischemia (45 min) and reperfusion (60 min). Endpoints were cardiac function and infarct size. Additional hearts were collected to evaluate MAPK phosphorylation (immunoblot). The nitric oxide synthase inhibitor L-NAME, the ERK1/2 inhibitor PD98059 and the p38 MAPK inhibitor FR167653 were injected intraperitoneally before hyperoxia or normoxia. Hyperoxia improved postischemic functional recovery and reduced infarct size (p < 0.05). Hyperoxic exposure caused cardiac phosphorylation of the MAPK family members p38 and ERK1/2, but not JNK. L-NAME, PD98059 and FR167653 all reduced the protection afforded by hyperoxic exposure, but did not influence performance or infarction in hearts of normoxic mice. The hyperoxia-induced phosphorylation of ERK1/2 and p38 was reduced by L-NAME and both MAPK inhibitors. Nitric oxide triggers hyperoxic protection, and ERK1/2 and p38 MAPK are involved in signaling of protection against ischemia-reperfusion injury. [ABSTRACT FROM AUTHOR]

1. Some anaesthetic agents such as barbiturates and opioids possess cardioprotective properties in rats, rabbits, dogs and pigs. The purpose of this study was to evaluate the effects of some commonly used anaesthetic agents (pentobarbital, isoflurane and a mixture of midazolam, fentanyl and fluanisone) on the tolerance of the isolated mouse heart to ischaemia-reperfusion injury. 2. The isolated, Langendorff-perfused hearts were subjected to 45 min of global ischaemia followed by 60 min of reperfusion. Left ventricular pressures, heart rate and coronary flow were measured and infarct size was determined using triphenyltetrazolium staining. 3. There were no differences in haemodynamic variables during reperfusion between groups. Infarct size was not influenced by the choice of anaesthesia. 4. None of the anaesthesia protocols exerted significant protective effects on the ischaemic-reperfused isolated mouse heart performance. In mice, isoflurane as well as pentobarbital, opioids and benzodiazepines may be safely used for anaesthesia without a risk of protective side-effects in isolated mouse heart studies. [ABSTRACT FROM AUTHOR]

Ischemic heart disease is the major cause of morbity and mortality in the Western world, with chronic heart failure as one complication. Ischemia-reperfusion injury may induce cardiomyocyte cell death by necrosis or apoptosis. The heart can be adapted to tolerate an ischemic event by preceeding brief episodes of ischemia and reperfusion, called preconditioning. Innate immunity has the latest years surfaced as important for the development of cardiovascular pathology as well as for myocardial protection. Nuclear factor kappa B (NFκB) is a redox sensitive transcription factor which contributes to the regulation of innate and adaptive immunity. NFκB regulates a battery of inflammatory genes, and has been indicated to play a role in the development of numerous pathological states. Activation of NFκB induces gene programs leading to transcription of factors which promote inflammation, among them leukocyte adhesion molecules, cytokines such as tumor necrosis factor alpha, and chemokines, but may in some situations also promote tissue remodelling, the resolution of inflammation, and transcription of some few substances with possible antiinflammatory effects. The present paper reviews the basic regulation of NFκB, and the possible role of NFκB activation in ischemia-reperfusion injury, in adaptation to ischemia-reperfusion injury, and in chronic heart failure. [ABSTRACT FROM AUTHOR]

Ischemic heart disease is the major cause of morbidity and mortality in the Western world. Ischemia-reperfusion injury may induce cardiomyocyte cell death by necrosis or apoptosis. The heart can be adapted to tolerate an ischemic event by preceding brief episodes of ischemia and reperfusion, called preconditioning. Preconditioning protects the heart when it is directed towards the heart itself either immediately before or several days before an induced ischemic event. Adaptation by preconditioning can even be achieved in other organs, and preconditioning one organ can protect another organ. Evidence suggests that preconditioning may be a naturally occurring adaptive process in vivo , and in humans unstable angina before acute myocardial infarction may represent the phenomenon. The mechanisms underlying the response are complex and intertwined, and probably differ between acute and delayed, as well as local or remote models. Based on animal experiments, it appears that preconditioning consists of a trigger phase with release of signal substance(s), a signal transduction phase where cascade reactions are induced, and an organ effector phase where mediator(s) ensure organ protection against necrosis and apoptosis. Understanding the underlying mechanisms of action is crucial to making therapeutic use of this powerful mode of myocardial protection. [ABSTRACT FROM AUTHOR]

Investigates whether pre-treatment with methylprednisolone (MP) protected the heart against ischemia-reperfusion dysfunction. Testing of the hypothesis that the protection may be due to induction of the cardioprotective heat shock protein 72 (HSP72). Cardiac function; HSP72.

Ischaemic preconditioning reduces infarct size, but the effects on cardiac function after global ischaemia are more controversial. Additionally, species differences may exist. The present study investigates the effects of preconditioning on cardiac performance in the globally ischaemic, Langendorff-perfused guinea-pig heart. Hearts were stabilized for 25 min, and divided into the following groups: (1) (n = 8) control perfusion for 16 min before 30-min global ischaemia and 30-min reperfusion, (2) (n = 7) two episodes of 3-min ischaemia and 5-min reperfusion before global ischaemia, (3) (n = 7) 5-min ischaemia and 10-min reperfusion before ischaemia, (4) (n = 8) control perfusion before 40-min ischaemia and 30-min reperfusion, (5) (n = 8) Preconditioning as group 2 before ischaemia as group 4, (6) (n = 9) Control perfusion before 50-min ischaemia and 30-min reperfusion, (7) (n = 10) Preconditioning as group 2 before ischaemia as group 6. A dose-dependent reduction of left ventricular systolic pressure, and increase of end-diastolic pressure was observed during reperfusion after 30-, 40- and 50-min ischaemia. Preconditioning did not influence these changes, nor did it attenuate the incidence of severe reperfusion arrhythmias or reduction of coronary flow. In conclusion, ischaemic preconditioning does not improve cardiac function during reperfusion of the globally ischaemic, isolated guinea-pig heart. [ABSTRACT FROM AUTHOR]

Preconditioning is in experimental studies the most powerful mode of cardioprotection known. The signal transduction pathways involve a variety of trigger substances, mediators, receptors, and effectors. The studies of preconditioning in cardiac surgery provide conflicting results but the majority of studies show that ischemic preconditiong is an effective adjunct to myocardial protection. However, ischemic preconditioning with repeated clamping of the aorta will never get widespread use. If the “preconditioning reponse” is to be exploited in cardiac surgery, targeting the underlying molecular mechanisms must provide easily applicable techniques or drugs, which are shown in large scale clinical studies to be beneficial. [Copyright &y& Elsevier]

Apoptosis or programed cell death is a continuous process of destruction of nonfunctional cells. It is a physiologic process whereby the body disposes of unwanted cells by self-destruction and is our utmost defense against damaged cells. There are several pathways leading to programed cell death. Apoptosis is seen in failing, infarcted, and hibernating human hearts, and during open heart surgery. Apoptosis appears to be induced by myocardial ischemia-reperfusion injury and this is reduced by ischemic preconditioning. Antiapoptotic interventions may be a future target for myocardial protection. [Copyright &y& Elsevier]

Abstract: Aquaporin-1 (AQP1) is expressed in human and mouse hearts, but little is known about its cellular and subcellular localization and regulation. The aim of this study was to investigate the localization of AQP1 in the mouse heart and to determine the effects of ischemia and hypoxia on its expression. Mouse myocardial cells were freshly isolated and split into cardiomyocyte and non-cardiomyocyte fractions. Isolated, Langendorff-perfused C57Bl6 mouse hearts (n=46) were harvested with no intervention, subjected to 35min of ischemia or ischemia followed by 60min of reperfusion. Eleven mouse hearts were perfusion-fixed for electron microscopy. Forty C57Bl6 mice were exposed to normobaric hypoxia for one or two weeks (n=12). Needle biopsies of human left ventricular myocardium were sampled (n=30) during coronary artery bypass surgery before cardioplegia and after 30min of reperfusion. Human umbilical vein endothelial cells (HUVECs) were subjected to 4h of hypoxia with reoxygenation for either 4 or 24h. AQP1 expression was studied by electron microscopy with immunogold labeling, Western blot, and qPCR. Expression of miR-214 and miR-320 in HUVECs with hypoxia was studied with qPCR. HUVECs were then transfected with precursors and inhibitors of miR-214. AQP1 expression was confined to cardiac endothelial cells, with no signal in cardiomyocytes or cardiac fibroblasts. Immunogold electron microscopy showed AQP1 expression in endothelial caveolae with equal distribution along the basal and apical membranes. Ischemia and reperfusion tended to decrease AQP1 mRNA expression in mouse hearts by 37±9% (p=0.06), while glycosylated AQP1 protein was reduced by 16±9% (p=0.03). No difference in expression was found between ischemia alone and ischemia-reperfusion. In human left ventricles AQP1 mRNA expression was reduced following cardioplegia and reperfusion (p=0.008). Hypoxia in mice reduced AQP1 mRNA expression by 20±7% (p<0.0001), as well as both glycosylated (−47±10%, p=0.03) and glycan-free protein (−34±16%, p=0.05). Hypoxia and reoxygenation in HUVECs downregulated glycan-free AQP1 protein (−34±24%, p=0.04) and upregulated miR-214 (+287±52%, p<0.05). HUVECs transfected with anti-miR-214 had increased glycosylated (1.5 fold) and glycan-free (2 fold) AQP1. AQP1 in mouse hearts is localized to endothelial cell membranes and caveolae. Cardioplegia, ischemia and hypoxia decrease AQP1 mRNA as well as total protein expression and glycosylation, possibly regulated by miR-214. [Copyright &y& Elsevier]

Abstract: Objective: Exposure to normobaric hyperoxia protects the heart against ischemia–reperfusion injury ex vivo. In the present study, we investigated the effect of the early and late phase of hyperoxia on in vivo myocardial infarction and apoptosis. Methods: Rats were exposed to room air preoxygenation (O2 ≥95%) followed by regional ischemia (30min) and 0, 90, 180, and 360min of reperfusion. Hyperoxic exposure was performed for 120min either immediately or 24h before coronary occlusion followed by 360-min reperfusion. Infarct size was evaluated by Evans blue/triphenyltetrazolium chloride staining. Apoptosis in the infarcted area was evaluated by terminal deoxy-nucleotidyl transferase-mediated deoxy uridine triphosphate (dUTP) nick end-labeling (TUNEL). Caspase 3 activity was measured by fluorometric enzyme assay, Bcl-2 and Bax protein expression assessed by western blotting and DNA laddering assessed with DNA gel electrophoresis. Results: The infarct size did not increase with increasing duration of reperfusion. However, apoptosis as evaluated by Bcl-2/Bax ratio, caspase 3 activity, and TUNEL-positive cells increased with increasing time of reperfusion. Both early and delayed pretreatment with hyperoxia reduced infarct size (p =0.0013, p =0.046), ameliorated ischemic arrhythmias and increased Bcl-2/Bax ratio (p =0.015, p =0.0159). Only hyperoxia immediately before coronary occlusion decreased caspase 3 activity (p =0.026) and decreased TUNEL-positive staining (p =0.046) with no visible DNA laddering. Conclusions: Detection of myocardial apoptosis increased with prolongation of reperfusion time, as opposed to infarct detection where reperfusion was essential to detect infarction, but the infarct size did not increase with time. Pretreatment with hyperoxia significantly decreased infarct size and apoptotic cell death. Pretreatment, immediately before coronary occlusion, was most cardioprotective. [ABSTRACT FROM AUTHOR]

Abstract: Aims: Lactate is transported by stereo-specific, pH-dependent monocarboxylate transporters (MCTs), of which MCT1 is expressed in abundance in rodent and human hearts. This study investigated the expression and functional role of MCT1 in mouse hearts during acute myocardial ischemia. Main methods: Mice hearts were isolated and Langendorff-perfused with induced global ischemia (40 min) and reperfusion with function and infarct size as end-points. Hearts were collected serially for protein extraction and immunoblotting with an MCT1 antibody, and for determining subcellular localization with immunogold EM. Immortalized cardiomyocytes (HL-1 cells) were injured with hydrogen peroxide, and cell death in the presence or absence of the competitive inhibitor of MCT1, d-lactate, was evaluated by Trypan blue exclusion. Key findings: MCT1 expression increased after 15 min reperfusion (p <0.05), but was not significantly increased after 60 min. MCT1 was localized in mitochondria, plasma membrane, and intercalated disks. At 15 min reperfusion, gold particle count was increased in the intercalated disks (p <0.05). d-lactate administration to isolated hearts either for 5 min before ischemia or at the first 5 min of reperfusion increased infarct size (p <0.01). A significant impairment of left ventricular performance was found when d-lactate was given before ischemia. MCT1 expression was not influenced by d-lactate. When HL-1 cells were treated with d-lactate before injury was induced with hydrogen peroxide, cell death was increased (p <0.05). Significance: Inhibition of MCT1 increases cell death. Increased MCT1 expression after ischemia and reperfusion is likely to restore cardiac pH through lactate export. [Copyright &y& Elsevier]

Abstract: Objective: Ischemic preconditioning reduces infarct size and improves hemodynamic function. Unstable angina may be a clinical analogue to ischemic preconditioning, and involve activation of gene programs. We hypothesized that preceding unstable angina and/or ischemic preconditioning activated genes regulated by nuclear factor kappa B (NFκB) or hypoxia-inducible factor 1 alpha in parallel to improved cardiac function. Methods: Patients undergoing coronary artery bypass grafting with stable or unstable angina were subjected to ischemic preconditioning or sham treatment (n =10–11 in each group). Central hemodynamics were monitored. Left ventricular and atrial biopsies were harvested before cardioplegic arrest and after 25min of reperfusion. Expression of heat shock protein 72 was evaluated by immunoblot, and activation of NFκB was detected by electrophoretic mobility shift assay. Real-time PCR was used to quantify expression of genes regulated by NFκB (inducible nitric oxide synthase, tumor necrosis factor-alpha, intercellular adhesion molecule 1) or by hypoxia-inducible factor 1 alpha (heme oxygenase-1, glucose transporter-1 and vascular endothelial growth factor-A). Results: Ischemic preconditioning improved postoperative cardiac index and left ventricular stroke work index in both stable and unstable groups on the first postoperative day. Expression of hypoxia-inducible factor 1 alpha regulated genes was not influenced by cardioplegia and reperfusion, ischemic preconditioning or unstable angina. Expression of the NFκB-regulated genes increased after cardioplegia and reperfusion, but this was not influenced by ischemic preconditioning in stable patients. Inducible nitric oxide synthase and tumor necrosis factor expression were reduced after ischemic preconditioning in patients with unstable angina. There were no significant differences in gene expression between stable and unstable patients before cardioplegia and ischemic preconditioning. NFκB translocation at reperfusion was reduced in stable preconditioned and unstable control patients compared to stable controls. Heat shock protein 72 expression increased after preconditioning of unstable patients. Conclusion: Cardiac function was improved by ischemic preconditioning in both stable and unstable patients. Unstable angina per se had no effect. NFκB-regulated genes were influenced by ischemic preconditioning, but hypoxia-inducible genes were not. [Copyright &y& Elsevier]

Abstract: Objective: Corticosteroids and hyperoxia protect the heart against ischemia–reperfusion injury and may attenuate vascular reactivity. We hypothesized that (1) combining these two pretreatments induces an additive cardioprotection, (2) protection depends on activation of survival kinases and/or heat shock proteins, and (3) these interventions would change vascular reactivity into a more relaxed state. Methods: Male rats were randomized (n =10 in each group): 1. control, 2. dexamethasone (3mg/kg) injected 24 and 12h before harvesting the hearts, 3. 60min of hyperoxia (90–95% O2) immediately before harvest, 4. combination of dexamethasone and hyperoxia as in groups 2 and 3. The hearts were Langendorff-perfused and exposed to 30min of global ischemia and reperfused for 120min. Cardiac function was monitored and infarct size determined. Isometric tension to vasoconstrictive and vasodilatory agents was measured in femoral artery rings. Phosphorylation of survival kinases (protein kinase B/AKT, extracellular signal-regulated kinases (ERK1/2), the stress-activated/c-Jun NH2 terminal kinases (SAPK/JNK) and p38 MAPK), adenosine monophosphate dependent kinase (AMPK) and expression of heat shock protein 72 (HSP72) in hearts was evaluated by immunoblotting. Results: Infarct size was attenuated in all pretreated groups versus controls: 29% reduction in the combined group (p <0.01), 23% in hyperoxia group (p <0.05) and 31% in dexamethasone group (p <0.01). There was no significant difference between the treated groups. Combined pretreatment improved postischemic left ventricular end diastolic pressure compared to all other groups (p <0.001 vs controls, p =0.002 vs dexamethasone, p =0.005 vs hyperoxia). Combined pretreatment improved left ventricular developed pressure and coronary flow compared to controls (p <0.001 for both) and hyperoxia (p =0.0047 and p =0.0024, respectively). Combined pretreatment enhanced endothelium-independent relaxation (p =0.0032) compared to controls. Excepting ERK1/2 phosphorylation in the combined group during early reperfusion, there was no increased phosphorylation of the survival kinases AKT, p38, JNK, or AMPK and no increase of HSP72 expression. Conclusion: Combined pretreatment by hyperoxia and dexamethasone improved postischemic heart function, but did not reduce infarct size compared to single pretreatment groups. Except of a possible role of ERK1/2, protection depended neither on survival kinases nor heat shock protein 72. [Copyright &y& Elsevier]

Abstract: Background: Large burns cause systemic inflammation and myocardial depression. We hypothesized that small burns affect cardiac tolerance to ischemia, and that tumor necrosis factor alpha (TNFα) signaling through endothelin-1 (ET) and nuclear factor kappa B (NFκB) are associated. Methods: Mice were randomly assigned to four groups: burn (caused by boiling water on <2% of the body surface area), sham, burn+etanercept (TNFα blocker) treatment and sham+etanercept treatment. Twenty-four hours later, hearts were isolated and subjected to global ischemia followed by reperfusion. Additional hearts and burned skin lesions were sampled to evaluate expression of TNFα (immunoblotting) and endothelin-1 (radioimmunoassay). A NFκB-luciferase reporter mouse was used to evaluate NFκB activation. Results: Baseline cardiac function before ischemia (BI) was only negligibly influenced by burn or etanercept, but was reduced by burn+etanercept. Burn markedly impaired post-ischemic left ventricular function and increased infarct size in comparison with sham-treated mice. Cardiac, but nut cutaneous, expression of TNFα was increased in burned mice, while cardiac NFκB and endothelin-1 were not influenced. TNFα blockade reduced the detrimental effects of burn on cardiac tolerance to ischemia. Conclusions: Small cutaneous burns, that did not influence baseline heart function, impaired the tolerance to ischemia. This effect may be mediated through TNFα, but does not involve signaling through NFκB or endothelin-1. [Copyright &y& Elsevier]

Objective: Adaptation to ischemia by brief episodes of ischemia and reperfusion (preconditioning) of the heart protects the heart against sustained ischemia, where the transcription factor nuclear factor kappa-B (NFκB) appears crucial for the protection. Preconditioning of the heart may even be evoked by brief episodes of ischemia and reperfusion in other organs. The present study investigates a possible role for NFκB and inducible nitric oxide synthase (iNOS) in adaption to ischemia by remote, delayed protection. Methods: Mice (wild-types, or with targeted deletions of the NFκB p105 or the iNOS gene) were subjected to cycles of occlusion and reperfusion of both hind limbs, and 24h later their hearts were isolated and Langendorff-perfused with induced global ischemia and reperfusion. Infarct size was measured. Skeletal muscles from ischemized limbs as well as hearts were also collected for polymerase chain reaction (PCR) and electromobility shift assay (EMSA). Results: Hind limb preconditioning protected left ventricular function and reduced infarct size during reperfusion in wild-type mice. Nuclear translocation of NFκB was detected in both heart and preconditioned skeletal muscle 1–2h after the preconditioning episodes (EMSA); while cardiac mRNA for iNOS gradually increased in a 24-h time course after hind limb preconditioning (real-time PCR). When hind limbs of mice with targeted deletions for the p105 subunit of NFκB or the iNOS gene were preconditioned, no beneficial effect was observed in the heart. Conclusions: Delayed cardioprotection induced by hind limb preconditioning involves signaling through NFκB and iNOS. [Copyright &y& Elsevier]

Objectives: Brief episodes of myocardial ischemia-reperfusion employed during reperfusion after a prolonged ischemic insult may attenuate the total ischemia-reperfusion injury. This phenomenon has been termed ischemic postconditioning. In the present study, we studied the possible effect of postconditioning on persistent reperfusion-induced ventricular fibrillation (VF) in the isolated rat heart model. Methods: Isolated Langendorff-perfused rat hearts (n=46) were subjected to 30 min of regional ischemia and reperfusion. The hearts with persistent VF (n=11) present after 15 min of reperfusion were then randomly assigned into one of the two groups: (1) control hearts (n=6), in which perfusion was continued without intervention; (2) postconditioned hearts (n=5) subjected to 2 min of global ischemia followed by reperfusion. Left ventricular pressures, heart rate, coronary flow, and electrogram were monitored throughout the experiment. Results: Conversion of VF into regular rhythm was observed in all hearts subjected to postconditioning. Regular beating was maintained by all postconditioned hearts during the subsequent reperfusion. None of the hearts in the control group had normal rhythm at the end of the experiment. At the end of reperfusion, the left ventricular developed pressure was lower in beating postconditioned hearts compared to the hearts that did not develop persistent VF. Conclusions: Ischemic postconditioning possesses strong antiarrhythmic effect against persistent reperfusion-induced tachyarrhythmias. Postconditioning may be an interesting, novel adjunct strategy to protect the heart. [Copyright &y& Elsevier]

Short episodes of ischemia and reperfusion in various organs may protect the organ itself, and the heart both as an immediate and a delayed effect. The present study investigates whether a systemic protection of vascular function occurs during adaption to ischemia. Brain ischemia was induced by bilateral ligation of the internal carotid arteries in C57BL6 mice, and 24–36 hours later rings of the thoracic aorta were mounted to study in vitro relaxation and contraction, or proteins were extracted for immunoblotting for endothelial nitric oxide synthase (eNOS) or inducible NOS (iNOS). eNOS decreased, while iNOS increased in the aortic wall after carotid artery ligation. In vitro contraction to increasing concentrations of prostaglandin F2α (PGF2α) was attenuated, while relaxation to acetylcholine (ACh) was enhanced. The latter was abolished by the iNOS-inhibitor aminoguanidine. When brain ischemia was induced in iNOS deficient mice, an increase of aortic eNOS was found 24 hours later. The ischemia-induced attenuated relaxation to PGF2α and enhanced relaxation to ACh were abolished. Aortic rings from mice with severe atherosclerosis (apolipoprotein E and low density lipoprotein receptor double knockout (ApoE/LDLr KO) mice) and spontaneous ischemic events in the heart or brain in vivo were also studied. Spontanous ischemic events in ApoE/LDLr KO animals did not influence iNOS and eNOS in the vessel wall. A reduced contraction to PGF2α was observed, but relaxation to ACh was unchanged. These findings suggest that induced brain ischemia as a model of delayed, remote preconditioning protects vessel reactivity, and this protection is mediated by iNOS. [Copyright &y& Elsevier]