Reperfusion Damage Research Articles

Theophylline as an add-on to thrombolytic therapy in acute ischaemic stroke (TEA-Stroke): A randomized, double-blinded, placebo-controlled, two-centre phase II study.

Early reperfusion of brain tissue at risk of injury (penumbra salvage) is crucial in treating acute ischaemic stroke. Neuroprotective agents may extend the time window for the reperfusion. The vasoactive agent theophylline redistributes the perfusion to ischaemic brain tissue and thus reduces brain damage, brain tissue oedema and mortality in animal stroke models. Furthermore, treatment with theophylline has been shown to result in considerable and rapid clinical improvement, albeit only temporary, in some stroke patients.We hypothesize that treatment with theophylline will improve the collateral supply in acute ischaemic brain tissue and thus facilitate reperfusion despite proximal vessel occlusion. The primary study objective is to evaluate whether theophylline is safe and efficient in acute ischaemic stroke patients as an add-on to thrombolytic therapy. The TEA-Stroke Trial is a two-centre, proof of concept phase II clinical study with a randomized, double-blinded, placebo-controlled design. One hundred and twenty patients with acute ischaemic stroke and significant perfusion-diffusion mismatch, as determined by magnetic resonance imaging, are randomized 1:1 to either theophylline or placebo as an add-on to standard thrombolytic therapy. The dual primary outcome measures include penumbra salvage (penumbral tissue not developing into infarcted tissue) and clinical improvement at the 24-h follow-up. Results from studies of theophylline in stroke animal models, clinical case series and randomized clinical trials are controversial. A Cochrane analysis from 2004 concluded that there was not enough evidence to assess whether theophylline is safe and improves outcomes in patients with acute ischaemic stroke. The TEA-Stroke Trial will clarify whether theophylline as an add-on to standard thrombolytic therapy improves penumbra salvage with a reduced risk of reperfusion damage, reduced final infarct size, and improved clinical outcome.

Activation of the Na+/H+ exchanger in isolated cardiomyocytes through β-Raf dependent pathways. Role of Thr653 of the cytosolic tail

The mammalian Na+/H+ exchanger isoform 1 (NHE1) is a ubiquitous plasma membrane protein that is a key regulator of intracellular pH in isolated cardiomyocytes. A 500 amino acid membrane domain removes protons and is regulated by a 315 amino acid cytosolic domain. In the myocardium, aberrant regulation of NHE1 contributes to ischemia reperfusion damage and to heart hypertrophy. We examined mechanisms of regulation of NHE1 in the myocardium by endothelin and β-Raf. Endothelin stimulated NHE1 activity and activated Erk-dependent pathways. Inhibition of β-Raf reduced NHE1 activity and Erk-pathway activation. We demonstrated that myocardial β-Raf binds to the C-terminal 182 amino acids of the NHE1 protein and that β-Raf is associated with NHE1 in intact cardiomyocytes. NHE1 was phosphorylated in vivo and the protein kinase inhibitor sorafenib reduced NHE1 phosphorylation levels. Immunoprecipitates of β-Raf from cardiomyocytes phosphorylated the C-terminal 182 amino acids of NHE1 and mass spectrometry analysis showed that amino acid Thr653 was phosphorylated. Mutation of this amino acid to Ala resulted in defective activity while mutation to Asp restored the activity. The results demonstrate that Thr653 is an important regulatory amino acid of NHE1 that is activated through β-Raf dependent pathways by phosphorylation either directly or indirectly by β-Raf, and this affects NHE1 activity.

Role of sphingomyelinase in mitochondrial ceramide accumulation during reperfusion

Ceramide accumulation in mitochondria has been associated with reperfusion damage, but the underlying mechanisms are not clearly elucidated. In this work we investigate the role of sphingomyelinases in mitochondrial ceramide accumulation, its effect on reactive oxygen species production, as well as on mitochondrial function by using the sphingomyelinase inhibitor, tricyclodecan-9-yl-xanthogenate (D609). Correlation between neutral sphingomyelinase (nSMase) activity and changes in ceramide content were performed in whole tissue and in isolated mitochondria from reperfused hearts. Overall results demonstrated that D609 treatment attenuates cardiac dysfuncion, mitochondrial injury and oxidative stress. Ceramide was accumulated in mitochondria, but not in the microsomal fraction of the ischemic-reperfused (I/R) group. In close association, the activity of nSMase increased, whereas glutathione (GSH) levels diminished in mitochondria after reperfusion. On the other hand, reduction of ceramide levels in mitochondria from I/R+D609 hearts correlated with diminished nSMase activity, coupling of oxidative phosphorylation and with mitochondrial integrity maintenance. These results suggest that mitochondrial nSMase activity contributes to compartmentation and further accumulation of ceramide in mitochondria, deregulating their function during reperfusion.

The estimation of selen content in blood serum and the diet correction in patients with acute Q-wave myocardial infarction

The use of selenium-containing specialized food products in prophylactic and therapeutic diets is based on hypothesis that selenium as antioxidant can potentially decrease the risk of oxidative stress induced diseases. There is a hypothesis, which is strongly supported during last 10-15 years that the reactive oxygen species are generated during ischemic myocardium reperfusion and cardiomyocyte damage. In addition selenium as an important component of glutationperoxidaze, it promotes protection against ischemic damage, improvement of functional regeneration and reduction of morphological changes in cardiomyocytes. We established a low serum selenium level in patients with acute and subacute stage of Q-wave myocardial infarction (75.5±1.8 mcg/L). New dietary product - jam made of seaweed with dried apricot enriched with selenium - was included in patient diet. We observed 72 persons (40-75 years old) who were randomized in a control group (30 patients) on standard treatment and a main group (42 patients) that received the dietary product (80 mcg Se) in addition to the standard treatment. Critical serum selenium level was found initially in 16 main group patients (38%), in 2 patients (7%) 2 weeks later and in none of patients in a month of dietary jam treatment. Selen concentration in blood serum increased from basal value 78.3±3.1 mcg/L after 2 weeks of treatment by 16% and after one month of treatment by 20% (p <0.05). In the control group the serum selenium level remained on critical and selenium-deficient levels during a month on standard treatment without dietary compensation. No adverse events were revealed in patients of main group, the investigated product was well tolerated.

Neuroprotective effects of Withania coagulans root extract on CA1 hippocampus following cerebral ischemia in rats.

Oxygen free radicals may be implicated in the pathogenesis of ischemia reperfusion damage. The beneficial effects of antioxidant nutrients, as well as complex plant extracts, on cerebral ischemia-reperfusion injuries are well known. This study was conducted to determine the effects of the hydro-alcoholic root extract of Withania coagulans on CA1 hippocampus oxidative damages following global cerebral ischemia/reperfusion in rat. Male Wistar rats were randomly divided in five groups: control, sham operated, Ischemia/ Reperfiusion (IR), and Withania Coagulans Extract (WCE) 500 and 1000mg/kg + I/R groups. Ischemia was induced by ligation of bilateral common carotid arteries for 30 min after 30 days of WCE administration. Three days after, the animals were sacrificed, their brains were fixed for histological analysis (NISSL and TUNEL staining) and some samples were prepared for measurement of malondialdehyde (MDA) level and superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activity in hippocampus. WCE showed neuroprotective activity by significant decrease in MDA level and increase in the SOD, CAT and GPx activity in pretreated groups as compared to I/R groups (p<0.001). The number of intact neurons was increased while the number of TUNEL positive neurons in CA1 hippocampal region in pretreated groups were decreased as compared to I/R group (p<0.001). WCE showed potent neuroprotective activity against oxidative stress-induced injuries caused by global cerebral ischemia/ reperfusion in rats probably by radical scavenging and antioxidant activities.

Comparisons of the effects of the sevoflurane and propofol on acute ischemia reperfusion and DNA damages in rabbits

Background and objectivesThe aim of this study was to compare the effects of sevoflurane and propofol anesthesia on oxidative DNA damage that occurs in low-extremity ischemia and is caused by tourniquet application. MethodsFourteen New Zealand rabbits were randomly allocated into two equal groups. Group S (n=7) received sevoflurane (2.5–4 percent) inhalation and Group P (n=7) received a propofol infusion (1–2mg·kg−1·min−1), after which a pneumatic tourniquet was placed on the right lower extremity. Blood samples were collected prior to tourniquet placement (baseline), 120min after ischemia, 15min after ischemia and 120minutes (min) after ischemia. Malondialdehyde (MDA) levels were analyzed to determine lipid peroxidation, and single cell gel electrophoresis (SCGE) was used to determine DNA damage. ResultsAt 15min after ischemia, the MDA levels in Group P (8.15±2.61μM) were higher than baseline (6.26±3.19μM, p=0.026) and Group S (4.98±0.77μM, p=0.01). DNA damage was similar in both groups, although DNA damage was higher than baseline (tail moment 0.63±0.27, tail intensity 3.76±1.26) in Group P at the 15th minute of reperfusion (tail moment 1.05±0.45, p=0.06; tail intensity 5.33±1.56, p=0.01). The increase in tail moment and tail intensity returned to normal levels in both groups 2hours after the termination of ischemia. ConclusionGiven that oxidative stress and genotoxic effect disappear in the late stages of reperfusion, we conclude that neither sevoflurane nor propofol can be considered superior to the other in anesthesia practices for extremity surgeries involving the use of a tourniquet.

Hydroxysafflor Yellow A Protects Neurons From Excitotoxic Death through Inhibition of NMDARs.

Excessive glutamate release causes overactivation of N-methyl d-aspartate receptors (NMDARs), leading to excitatory neuronal damage in cerebral ischemia. Hydroxysafflor yellow A (HSYA), a compound extracted from Carthamus tinctorius L., has been reported to exert a neuroprotective effect in many pathological conditions, including brain ischemia. However, the underlying mechanism of HSYA's effect on neurons remains elusive. In the present study, we conducted experiments using patch-clamp recording of mouse hippocampal slices. In addition, we performed Ca2+ imaging, Western blots, as well as mitochondrial-targeted circularly permuted yellow fluorescent protein transfection into cultured hippocampal neurons in order to decipher the physiological mechanism underlying HSYA's neuroprotective effect.Through the electrophysiology experiments, we found that HSYA inhibited NMDAR-mediated excitatory postsynaptic currents without affecting α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor and γ-aminobutyric acid A-type receptor-mediated currents. This inhibitory effect of HSYA on NMDARs was concentration dependent. HSYA did not show any preferential inhibition of either N-methyl d-aspartate receptor subtype 2A- or N-methyl d-aspartate receptor subtype 2B- subunit-containing NMDARs. Additionally, HSYA exhibits a facilitatory effect on paired NMDAR-mediated excitatory postsynaptic currents. Furthermore, HSYA reduced the magnitude of NMDAR-mediated membrane depolarization currents evoked by oxygen-glucose deprivation, and suppressed oxygen-glucose deprivation–induced and NMDAR-dependent ischemic long-term potentiation, which is believed to cause severe reperfusion damage after ischemia. Through the molecular biology experiments, we found that HSYA inhibited the NMDA-induced and NMDAR-mediated intracellular Ca2+ concentration increase in hippocampal cultures, reduced apoptotic and necrotic cell deaths, and prevented mitochondrial damage. Together, our data demonstrate for the first time that HSYA protects hippocampal neurons from excitotoxic damage through the inhibition of NMDARs. This novel finding indicates that HSYA may be a promising pharmacological candidate for the treatment of brain ischemia.

Pathophysiological aspects and management workflow of coronary microvascular obstruction in ST-segment elevation myocardial infarction

Restoring blood flow to ischemic but viable myocardium and reducing infarct size constitute the goals of reperfusion therapy with fibrinolytic drugs or primary percutaneous intervention in patients with ST-segment elevation myocardial infarction (STEMI). However, in a sizable proportion of patients, this intervention gains to reopen the obstructed epicardial coronary artery but does not achieve myocardial reperfusion because of coronary microvascular obstruction phenomenon (CMVO). In the last years, consistent evidence has pointed out how CMVO has a negative impact on outcome in patients with acute STEMI. Of note, most of the trials in this setting, mainly targeting reperfusion damage, have failed to show beneficial effects. In this article we provide a revision of mechanisms, diagnosis and prognosis of CMVO in acute STEMI, also pointing out the need of an integrated approach in order to prevent and treat CMVO in the different time windows of the acute event.

Controversies on the role of mitochondria in coronary reperfusion damage as potential therapeutic target

Reperfusion of an occluded coronary artery is the most effective intervention in reducing cardiac injury which occurs as a consequence of an acute myocardial infarction. However, reperfusion per se may exacerbate the extent of myocardial injury which started at the onset of ischemia, thus leading to the so-called "reperfusion injury". The published studies in the last few years provide new hopes on the mitochondrion and its role in ischemia/reperfusion injury, through the modulation of the mitochondrial permeability pore. After very encouraging experimental data, two clinical trials are presented, CIRCUS and CYCLE, which should help addressing the still outstanding question: may an intervention targeted to decrease the permeability of the mitochondrial pore during reperfusion reduce infarct size and improve clinical outcomes of patients with myocardial infarction undergoing primary percutaneous coronary intervention?

Scaffold Optimisation of Tetravalent Antagonists of the Mannose Binding Lectin.

Antagonists of mannose binding lectin (MBL) have shown a protective role against brain reperfusion damage after acute ischemic stroke. Here we describe the design and streamlined synthesis of glycomimetic MBL antagonists based on a new tetravalent dendron scaffold. The dendron was developed by optimisation of a known polyester structure previously demonstrated to be very efficient for ligand presentation to MBL. Replacement of a labile succinyl ester bond with a more robust amide functionality, use of a longer and more hydrophilic linker, fast modular synthesis and orthogonal functionalisation at the focal point are the main features of the new scaffold. The glycoconjugate constructs become stable to silica gel chromatography and to water solutions at physiological pH, while preserving water solubility and activity in an SPR assay against the murine MBL-C isoform. Higher-order constructs were easily assembled, as demonstrated by the synthesis of a 16-valent dendrimer, which leads to two orders of magnitude increase in activity over the tetravalent version against MBL-C.

Interferon-β Modulates Inflammatory Response in Cerebral Ischemia.

BackgroundStroke is a leading cause of death in the world. In >80% of strokes, the initial acute phase of ischemic injury is due to the occlusion of a blood vessel resulting in severe focal hypoperfusion, excitotoxicity, and oxidative damage. Interferon‐β (IFNβ), a cytokine with immunomodulatory properties, was approved by the US Food and Drug Administration for the treatment of relapsing‐remitting multiple sclerosis for more than a decade. Its anti‐inflammatory properties and well‐characterized safety profile suggest that IFNβ has therapeutic potential for the treatment of ischemic stroke.Methods and ResultsWe investigated the therapeutic effect of IFNβ in the mouse model of transient middle cerebral artery occlusion/reperfusion. We found that IFNβ not only reduced infarct size in ischemic brains but also lessened neurological deficits in ischemic stroke animals. Further, multiple molecular mechanisms by which IFNβ modulates ischemic brain inflammation were identified. IFNβ reduced central nervous system infiltration of monocytes/macrophages, neutrophils, CD4+ T cells, and γδ T cells; inhibited the production of inflammatory mediators; suppressed the expression of adhesion molecules on brain endothelial cells; and repressed microglia activation in the ischemic brain.ConclusionsOur results demonstrate that IFNβ exerts a protective effect against ischemic stroke through its anti‐inflammatory properties and suggest that IFNβ is a potential therapeutic agent, targeting the reperfusion damage subsequent to the treatment with tissue plasminogen activator.

Induction of Hypometabolism as a Novel Strategy to Minimize Ischemia Reperfusion Injury in Renal Reconstruction Surgeries

Purpose: Hibernation is an altered physiological state which is marked by a characteristic lowering of body temperature and extreme metabolic rate depression. During this period, an animal undergoing hibernation may have its heart rate and blood flow decrease to 1/30 and its oxygen consumption to 1/100 of its respective euthermic levels. Even though these physiological parameters mimic conditions of ischemia their organs particularly their brain, heart, liver and kidneys do not show any detrimental effects of severely reduced blood flow when the animal comes out of hibernation. We wished to explore the benefits of inducing such a hypometabolic state in a mouse ischemiareperfusion injury model. Methods: We investigated the pre-conditioning effect of administering 5’-Adenosyl Monophosphate (5’-AMP) to mice to create a state of hypometabolism akin to hibernation. This hibernation-like state is used to study its protective effect on a subsequent renal ischemic episode. Results: Our results strongly suggest that an induction of a hypometabolic state greatly attenuates the ischemic and reperfusion damage to the kidneys, as studied by several specific parameters. Conclusions: We conclude that reprogramming of the kidney metabolism to a hypometabolic state before ischemia may protect the organ from oxidative damage and reduce ischemia/reperfusion injury. This concept may also be applicable to improve clinical outcomes in renal reconstruction as well as transplantation surgeries.

Antiarrhythmic Activity of Taurepar during Ischemic and Reperfusion Damage to Myocardium.

The antiarrhythmic effect of taurepar, an N-phenylalkyl derivative of taurine, was examined in experiments on rats subjected to acute myocardial ischemia/reperfusion leading to arrhythmia development. During acute ischemia, taurepar (25 mg/kg) completely prevented early postocclusion arrhythmias including extrasystoles, ventricular tachycardia, and ventricular fibrillation. During postischemic reperfusion, taurepar (25 mg/kg) did not prevent extrasystoles and ventricular tachycardia, but precluded the development of ventricular fibrillation and the death of animals. The antiarrhythmic potency of taurepar surpassed that of lidocaine during acute myocardial ischemia and that of propranolol during ischemia/reperfusion injury. The results suggest that taurepar is a promising antiarrhythmic drug with high antifibrillation activity.

Protective Effect of Platelet Rich Plasma on Experimental Ischemia/Reperfusion Injury in Rat Ovary

Background/Aims: Ovarian torsion is a common cause of local ischemic damage, reduced follicular activity and infertility. Platelet-rich plasma (PRP) contains growth factors with demonstrated cytoprotective properties; so we evaluated PRP efficacy in a rat ischemia/reperfusion (I/R) model. Methods: Sixty adult female Sprague-Dawley albino rats were randomly assigned to 6 groups of 8 animals each: Sham, Ischemia, I/R, Sham + PRP, I + PRP and I/R + PRP; and the remaining 12 used to prepare PRP. Ischemia groups were subjected to bilateral adnexal torsion for 3 h, while I/R and I/R + PRP groups received subsequent detorsion for 3 h. Intraperitoneal PRP was administered 30 min prior to ischemia (Ischemia + PRP) or reperfusion (I/R + PRP). Results: Total oxidant status (TOS), oxidative stress index (OSI) and total ovarian histopathological scores were higher in Ischemia and I/R groups than in the Sham group (p < 0.05). PRP decreased mean TOS, OSI and histopathological scores in I + PRP and I/R + PRP groups compared to the corresponding Ischemia and I/R groups (p < 0.001). There was a strong correlation between total histopathological score and OSI (r = 0.877, p < 0.001). Peritoneal vascular endothelial growth factor was significantly higher in PRP-treated groups than corresponding untreated groups (p < 0.05). Conclusion: PRP is effective for the prevention of ischemia and reperfusion damage in rat ovary.

Induction of substantial myocardial regeneration by an active fraction of the Chinese herb Rosa laevigata Michx.

BackgroundThe replacement of lost cardiac tissues by regenerated myocardium would be a therapeutic ideal for myocardial infarction. The objective of this study was therefore to evaluate the ability of an active fraction that was isolated from Rosa laevigata Michx in therapeutic cardiomyogenesis in a myocardial infarction rat model.MethodsThe myocardial infarction animal model was induced by the permanent ligation of the left anterior descending coronary artery in rats. The active fraction, which improves the survival rate and prevents ischemic reperfusion damage, was used to test the therapeutic effect of this fraction on myocardial infarction.ResultsThe oral administration of the active fraction for 4 weeks could progressively restore the decreased cardiac function due to myocardial infarction. The significantly improved cardiac function was probably attributed to the active fraction-induced myocardial regeneration, which replaced the lost cardiac tissues in the myocardial infarction animals.ConclusionsThe property of this active fraction appears to be entirely novel and may provide a potential therapeutic alternative for myocardial infarction.

Reply to Letter to the Editor

Previous studies show that chronic pyelonephritis and end stage renal disease may follow acute pyelonephritis in children and adolescents when improperly or inadequately treated. Our study shows that there is a significant decrease in renal function following untreated acute bacterial pyelonephritis due to nephron loss. The acute inflammatory response is responsible for much of the renal damage, although damage from renal ischemia is an additional significant factor. The present study used a combination of an antibiotic and a xanthine oxidase inhibitor (allopurinol) as compared to antibiotic therapy alone begun 72 hours after infection. Both were successful in eradicating the infection rapidly, but did not entirely prevent renal damage. Treatment prior to 72 hours thus is important. It appears that the combined treatment, designed to eradicate the bacteria as well as reduce the post-ischemic reperfusion damage and the phagocytic burst of phagocytosis is ideal, as this combined treatment was effective in preventing almost all renal damage and loss of renal function.

Inhibition of the nitric oxide/cyclic guanosine monophosphate pathway limited the cardioprotective effect of post-conditioning in hearts with apical myocardial infarction

Reperfusion damage involves opening of the mitochondrial permeability transition pore (mPTP) and loss of ATP synthesis. Several cardioprotective pathways are activated by ischemic or pharmacological post-conditioning (PC). The mechanisms that are activated by PC in no co-morbidity murine models include: activation of rescue kinases, oxidative stress reduction, glycolytic flux regulation and preservation of ATP synthesis. However, relatively scarce efforts have been made to define whether the efficacy of PC signaling is blunted by risk factors or systemic diseases associated with ischemic heart pathology. Experimental evidence has shown that the nitric oxide (NO)/cyclic guanosine monophosphate (cGMP) signaling is a main mechanism activated by PC in hearts without pathological history. In this work we evaluated the participation of the NO pathway, through downstream kinase activation and inhibition of mPTP in hearts with previous infarct.Myocardial infarction was induced with a single dose of isoproterenol (85mg/kg i.p.) to male Wistar rats. After 24h, the hearts were mounted into the Langendorff system and subjected to 30min of ischemia and 60min of reperfusion. PC consisted of 5 cycles of 30s of reperfusion/30s of ischemia, then the hearts were reperfused with or without inhibitors of the NO/cGMP pathway.PC activates the NO/cGMP pathway, as increased cGMP and NO levels were detected in isoproterenol-treated hearts. The cardioprotective effect of PC was abolished with both l-NAME (inhibitor of constitutive NO synthase) and ODQ (inhibitor of soluble guanylate cyclase), whereas the NO donor (DETA-NO) restored cardioprotection even in the presence of l-NAME or ODQ. We also found that mitochondrial structure and function was preserved in PC hearts.We conclude that PC exerts cardioprotection in hearts with previous infarct by maintaining mitochondrial structure and function through NO-dependent pathway.

Carbonic Anhydrase Protects Fatty Liver Grafts against Ischemic Reperfusion Damage.

Carbonic anhydrases (CAs) are ubiquitous metalloenzymes that catalyze the reversible hydration of carbon dioxide to bicarbonate and a proton. CAs are involved in numerous physiological and pathological processes, including acid-base homeostasis, electrolyte balance, oxygen delivery to tissues and nitric oxide generation. Given that these processes are found to be dysregulated during ischemia reperfusion injury (IRI), and taking into account the high vulnerability of steatotic livers to preservation injury, we hypothesized a new role for CA as a pharmacological agent able to protect against ischemic damage. Two different aspects of the role of CA II in fatty liver grafts preservation were evaluated: 1) the effect of its addition to Institut Georges Lopez (IGL-1) storage solution after cold ischemia; 2) and after 24h of cold storage followed by two hours of normothermic ex-vivo perfusion. In all cases, liver injury, CA II protein concentration, CA II mRNA levels and CA II activity were determined. In case of the ex-vivo perfusion, we further assessed liver function (bile production, bromosulfophthalein clearance) and Western blot analysis of phosphorylated adenosine monophosphate activated protein kinase (AMPK), mitogen activated protein kinases family (MAPKs) and endoplasmic reticulum stress (ERS) parameters (GRP78, PERK, IRE, eIF2α and ATF6). We found that CA II was downregulated after cold ischemia. The addition of bovine CA II to IGL-1 preservation solution efficiently protected steatotic liver against cold IRI. In the case of reperfusion, CA II protection was associated with better function, AMPK activation and the prevention of ERS and MAPKs activation. Interestingly, CA II supplementation was not associated with enhanced CO2 hydration. The results suggest that CA II modulation may be a promising target for fatty liver graft preservation.

Effect of metformin pretreatment on myocardial injury during coronary artery bypass surgery in patients without diabetes (MetCAB): a double-blind, randomised controlled trial

During coronary artery bypass graft (CABG) surgery, ischaemia and reperfusion damage myocardial tissue, and increased postoperative plasma troponin concentration is associated with a worse outcome. We investigated whether metformin pretreatment limits cardiac injury, assessed by troponin concentrations, during CABG surgery in patients without diabetes. We did a placebo-controlled, double-blind, single-centre study in an academic hospital in Nijmegen (Netherlands) in adult patients without diabetes undergoing an elective on-pump CABG procedure. We randomly assigned patients (1:1) in blocks of ten via a computer-generated randomisation sequence to either metformin hydrochloride (500 mg three times per day) or placebo (three times per day) for 3 days before surgery. The last dose was given roughly 3 h before surgery. Patients, investigators, trial staff, and the statistician were all masked to treatment allocation. The primary endpoint was the plasma concentration of high-sensitive troponin I at 6, 12, and 24 h postreperfusion after surgery, analysed in the per-protocol population with a mixed-model analysis using all these timepoints. Secondary endpoints included the occurrence of clinically relevant arrhythmias within 24 hours after reperfusion, the need for inotropic support, time to detubation, duration of stay in the intensive-care unit, and postoperative use of insulin. This study is registered with ClinicalTrials.gov, number NCT01438723. Between Nov 8, 2011, and Nov 22, 2013, we randomly assigned 111 patients to treatment (57 to metformin and 54 to placebo). Five patients dropped out from the metformin group, and six from the placebo group. 52 patients in the metformin group and 48 patients in the placebo group were included in the per-protocol analysis. Geometric mean high-sensitivity troponin I increased from 0 μg/L to 3·67 μg/L (95% CI 3·06-4·41) with metformin and to 3·32 μg/L (2·75-4·01) with placebo at 6 h after reperfusion; 2·84 μg/L (2·37-3·41) and 2·45 μg/L (2·02-2·96), respectively, at 12 h; and to 1·77 μg/L (1·47-2·12) and 1·60 μg/L (1·32-1·94) at 24 h. The concentrations did not differ significantly between the groups (difference 12·3% for all timepoints [95% CI -12·4 to 44·1] p=0·35). Occurrence of arrhythmias did not differ between groups (three [5·8%] of 52 patients who received metformin vs three [6·3%] of 48 patients who received placebo; p=1·00). There was no difference between groups in the need for inotropic support, time to detubation, duration of stay in the intensive-care unit, or postoperative use of insulin. No patients died within 30 days after surgery. Occurrence of gastrointestinal discomfort (mostly diarrhoea) was significantly higher with metformin than with placebo (11 [21·2%] of 52 vs two [4·2%] of 48 patients; p=0·01). Short-term metformin pretreatment, although safe, does not seem to be an effective strategy to reduce periprocedural myocardial injury in patients without diabetes undergoing CABG surgery. Netherlands Organisation for Health Research and Development and Netherlands Heart Foundation.

TEMPOL has limited protective effects on renal oxygenation and hemodynamics but reduces kidney damage and inflammation in a rat model of renal ischemia/reperfusion by aortic clamping

Renal ischemia-reperfusion (I/R) is a common clinical complication in critically ill patients that is associated with considerable morbidity and mortality. Renal I/R is a major cause of acute kidney injury (AKI) resulting from I/R-induced oxidative stress, sterile inflammation, and microcirculatory perfusion defects, which can be ameliorated with the superoxide scavenger TEMPOL. The most common cause of AKI in the clinical setting is aortic surgery with suprarenal aortic clamping. The protective effect of TEMPOL in aortic clamping-induced renal I/R has not been studied before. To evaluate the protective effects of TEMPOL on oxidative stress, inflammation, tissue injury, and renal hemodynamics and oxygenation in a clinically representative rat model of I/R using aortic cross-clamping. Animals (N = 24) were either sham-operated or subjected to ischemia (30 min) and 90-min reperfusion, with or without TEMPOL treatment (15 min before ischemia and during entire reperfusion phase, 200 μmol/kg/h). Systemic and renal hemodynamics, renal oxygenation, and blood gas values were determined at 15 min and 90 min of reperfusion. At 90-min reperfusion, iNOS, inflammation (IL-6, MPO), oxidative stress (MDA), and tissue damage (NGAL, L-FABP) were determined in tissue biopsies. TEMPOL administration at a cumulative dose of 400 μmol/kg conferred a protective effect on AKI in terms of reducing renal damage, inflammation, and iNOS activation. With respect to renal hemodynamics and oxygenation, TEMPOL only reduced renal vascular resistance to near-baseline levels at both reperfusion time points and partially ameliorated the I/R-induced drop microvascular partial tension of oxygen at 90 min reperfusion. Also, TEMPOL alleviated the I/R-induced metabolic acidosis. However, TEMPOL exerted no restorative effect in terms of the severely reduced mean arterial pressure, renal blood flow, and renal oxygen delivery and consumption. The renal oxygen extraction ratio remained unchanged during the 90-min reperfusion phase. Kidneys in all groups were anuric throughout the experiment. This clinically representative renal I/R model, which entails both renal I/R and hind limb I/R as opposed to the standardly used renal I/R model that employs renal artery clamping, resulted in relatively moderate direct AKI. The damage was exacerbated by the perturbed systemic hemodynamics and metabolic acidosis as a result of the hind limb I/R. TEMPOL partially intervened in the factors that led to AKI as well as renal microvascular partial tension of oxygen and metabolic acidosis. However, more effective interventions should be devised for the mean arterial pressure drop (i.e., anuria) associated with aortic clamping and for restoring other critical renal hemodynamic and oxygenation parameters in order to improve post-I/R renal function. TEMPOL is a promising compound that has been shown to protect kidneys from I/R damage, which is relevant in kidney transplantation, pancreas transplantation, and aortic aneurysm repair in kidney transplant patients. This study suggests that intervening with TEMPOL is not sufficient to ensure optimal clinical outcome in patients that have undergone aortic clamping and that more effective interventions should be investigated.