Decrease Reperfusion Injury Research Articles

Abstract 12689: Electron Transport Chain Inhibition to Decrease Reperfusion Injury in Transplanted Donation After Circulatory Death Rat Hearts

Introduction: Intrinsic to the donation after circulatory death (DCD) donation is ischemia and reperfusion injury, primarily mediated through release of reactive oxygen species (ROS) from the damaged mitochondria. Amobarbital (AMO) is a transient, reversible inhibitor of complex I known to decrease ROS generation. We studied whether AMO treatment during reperfusion can improve function in transplanted DCD hearts. Methods: Sprague Dawley rats, were assigned to four heart donor groups- DCD or DCD+AMO and control beating-heart donor (CBD) or CBD+AMO (n=6-8 each). Donor rats were anesthetized using ketamine/xylazine, intubated and connected to a ventilator. The carotid access was used to deliver heparin (1000 U/kg) and vecuronium bromide (4 mg/kg) to paralyze diaphragmatic muscles. The DCD process was initiated by disconnecting the ventilator. Donor hearts were procured following 25 minutes of in-vivo ischemia in the DCD groups, while CBD hearts were procured without terminating the ventilation. At procurement, after clamping the aortic arch, all donor hearts received 10 ml of University of Wisconsin cardioplegia solution delivered through the carotid cannula. The CBD+AMO and DCD+AMO groups received AMO (2 mM), dissolved in the cardioplegia. Heterotopic heart transplantation was performed in recipient rat’s abdomen. After 14 days the transplanted heart function was measured with a balloon tip catheter. Results: Compared to CBD hearts, DCD hearts had significantly lower developed pressure, the rate pressure product (developed pressure times heart rate), and negative dP/dt indicating decreased contractile and impaired diastolic function (Figure). AMO treatment of the DCD hearts resulted in significantly improved cardiac function compared to the untreated DCD hearts. Conclusions: A single treatment with AMO during the heart procurement improves the transplanted DCD heart function which was comparable to the control CBD hearts.

The cardioprotective effect of intralipid in decreasing the ischemic insults during off-pump coronary artery revascularization

BackgroundOff pump coronary artery revascularization (OPCAB) surgeries have benefits over the conventional on pump cardiac surgery, because it avoids the trauma caused by cardiopulmonary bypass (CPB) and minimize aortic manipulation. However, some disadvantages of OPCAB include the concern of ineffective coronary revascularization. Some drugs have shown the ability to protect the myocardium in different studies, by different methods. The usage of intralipid has been shown to make a better functional recovery of the cardiac muscles and help to decrease the myocardial infarct size, it shortens the action potential time, which show polyunsaturated fatty acids diets mechanism as an antiarrhythmic drug, and are associated with low incidence of coronary artery disease.MethodsWe divided patients into two groups according to the randomization envelopes: intralipid group (group A) received 1.5 ml/kg intralipid 20% through central venous line after sternotomy over 1 h and during infusion, blood pressure, heart rate, and temperature were monitored all through the infusion time. Control group (group B) received normal saline 0.9% in the same volume over the same duration.ResultsThis study showed that infusion of 1.5 ml/kg intralipid after sternotomy in off pump coronary artery revascularization given as preconditioning agent improve the myocardial ischemia reperfusion injury, decrease the need for high doses of nor adrenaline infusion after revascularization, earlier normalization in troponin levels starting 24 h after surgery and higher values of cardiac index were measured in ICU using PICCO.ConclusionsThis study showed the benefits of infusion of 1.5 ml/kg of intralipid after sternotomy, in preconditioning during OPCABG. Preconditioning with intralipid proved to decrease reperfusion injury in myocardium expressed by improvement in cardiac functions (EF and cardiac index) and normalization of specific cardiac marker (cardiac troponin I).

A Novel Oxygen Carrier (M101) Attenuates Ischemia-Reperfusion Injuries during Static Cold Storage in Steatotic Livers.

The combined impact of an increasing demand for liver transplantation and a growing incidence of nonalcoholic liver disease has provided the impetus for the development of innovative strategies to preserve steatotic livers. A natural oxygen carrier, HEMO2life®, which contains M101 that is extracted from a marine invertebrate, has been used for static cold storage (SCS) and has shown superior results in organ preservation. A total of 36 livers were procured from obese Zucker rats and randomly divided into three groups, i.e., control, SCS-24H and SCS-24H + M101 (M101 at 1 g/L), mimicking the gold standard of organ preservation. Ex situ machine perfusion for 2 h was used to evaluate the quality of the livers. Perfusates were sampled for functional assessment, biochemical analysis and subsequent biopsies were performed for assessment of ischemia-reperfusion markers. Transaminases, GDH and lactate levels at the end of reperfusion were significantly lower in the group preserved with M101 (p < 0.05). Protection from reactive oxygen species (low MDA and higher production of NO2-NO3) and less inflammation (HMGB1) were also observed in this group (p < 0.05). Bcl-1 and caspase-3 were higher in the SCS-24H group (p < 0.05) and presented more histological damage than those preserved with HEMO2life®. These data demonstrate, for the first time, that the addition of HEMO2life® to the preservation solution significantly protects steatotic livers during SCS by decreasing reperfusion injury and improving graft function.

Cardiomyocyte Protection by Hibernating Brown Bear Serum: Toward the Identification of New Protective Molecules Against Myocardial Infarction.

Ischemic heart disease remains one of the leading causes of death worldwide. Despite intensive research on the treatment of acute myocardial infarction, no effective therapy has shown clinical success. Therefore, novel therapeutic strategies are required to protect the heart from reperfusion injury. Interestingly, despite physical inactivity during hibernation, brown bears (Ursus arctos) cope with cardiovascular physiological conditions that would be detrimental to humans. We hypothesized that bear serum might contain circulating factors that could provide protection against cell injury. In this study, we sought to determine whether addition of bear serum might improve cardiomyocyte survival following hypoxia–reoxygenation. Isolated mouse cardiomyocytes underwent 45 min of hypoxia followed by reoxygenation. At the onset of reoxygenation, cells received fetal bovine serum (FBS; positive control), summer (SBS) or winter bear serum (WBS), or adult serums of other species, as indicated. After 2 h of reoxygenation, propidium iodide staining was used to evaluate cell viability by flow cytometry. Whereas, 0.5% SBS tended to decrease reperfusion injury, 0.5% WBS significantly reduced cell death, averaging 74.04 ± 7.06% vs. 79.20 ± 6.53% in the FBS group. This cardioprotective effect was lost at 0.1%, became toxic above 5%, and was specific to the bear. Our results showed that bear serum exerts a therapeutic effect with an efficacy threshold, an optimal dose, and a toxic effect on cardiomyocyte viability after hypoxia–reoxygenation. Therefore, the bear serum may be a potential source for identifying new therapeutic molecules to fight against myocardial reperfusion injury and cell death in general.

RBC Storage Lesion Studies in Humans and Experimental Models of Shock.

The finding of toxicity in a meta-analysis of observational clinical studies of transfused longer stored red blood cells (RBC) and ethical issues surrounding aging blood for human studies prompted us to develop an experimental model of RBC transfusion. Transfusing older RBCs during canine pneumonia increased mortality rates. Toxicity was associated with in vivo hemolysis with release of cell-free hemoglobin (CFH) and iron. CFH can scavenge nitric oxide, causing vasoconstriction and endothelial injury. Iron, an essential bacterial nutrient, can worsen infections. This toxicity was seen at commonly transfused blood volumes (2 units) and was altered by the severity of pneumonia. Washing longer-stored RBCs mitigated these detrimental effects, but washing fresh RBCs actually increased them. In contrast to septic shock, transfused longer stored RBCs proved beneficial in hemorrhagic shock by decreasing reperfusion injury. Intravenous iron was equivalent in toxicity to transfusion of longer stored RBCs and both should be avoided during infection. Storage of longer-stored RBCs at 2 °C instead of higher standard temperatures (4-6 °C) minimized the release of CFH and iron. Haptoglobin, a plasma protein that binds CFH and increases its clearance, minimizes the toxic effects of longer-stored RBCs during infection and is a biologically plausible novel approach to treat septic shock.

Trimetazidine in the Prevention of Tissue Ischemic Conditions.

Trimetazidine (TMZ) is a metabolic agent with significant anti-ischemic properties. By inhibiting the terminal enzyme in the β-oxidation pathway, it shifts the energy substrate metabolism, enhancing glucose metabolism. Thus, it maintains the required energy production with less oxygen consumption, an effect necessary in cases of myocardi. Trimetazidine was recently reaccredited as add-on therapy for symptomatic treatment in patients with stable angina, not adequately controlled or intolerant to first-line therapy. Trimetazidine was included in the European Society of Cardioloy 2013 guidelines for the management of stable coronary artery disease. Although TMZ has been used in cardiology for >40 years, only a few studies have assessed its effects in patients with acute ischemic conditions. This review summarizes the current literature regarding the addition of TMZ in patients with acute ischemic conditions (acute myocardial infarction, ST-segment elevation myocardial infarction, non-ST-segment elevation myocardial infarction, percutaneous coronary intervention, and coronary artery bypass grafting). There is growing evidence from recent studies that the addition of TMZ in patients with such conditions is beneficial in terms of myocardial damage and major cardiac events as well as decreasing reperfusion injury and contrast-induced nephropathy.

Soluble epoxide hydrolase inhibition decreases reperfusion injury after focal cerebral ischemia

Epoxyeicosatrienoic acids (EETs) are produced by cytochrome P450 epoxygenases from arachidonic acid, and their rapid metabolism is mainly through soluble epoxide hydrolase (sEH). EETs exert vasodilatory, anti-inflammatory, anti-apoptotic, and pro-angiogenic effects. Administration of sEH inhibitors before or at the onset of stroke is protective, but the effects of post-treatment at reperfusion, when inflammation is augmented, has not been as well studied. We tested the hypothesis that 1-Trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl)urea (TPPU), a potent and highly selective sEH inhibitor, suppresses inflammation and protects the brain when administered at reperfusion. Vehicle or 1 mg/kg TPPU was administered at reperfusion after 90 minutes of focal ischemia and again 24 hours later. Protein expression and activity of sEH increased after reperfusion and activity was decreased by TPPU administration. TPPU decreased infarct volume by 50%, reduced neurologic deficits and improved performance on sensorimotor tasks. Furthermore, TPPU significantly lowered the mRNA expression of interleukin-1beta by 3.5-fold and tumor necrosis factor-alpha by 2.2-fold, increased transforming growth factor-beta mRNA by 1.8-fold, and augmented immunostaining of vascular endothelial growth factor in peri-infarct cortex. Thus, inhibition of sEH at reperfusion significantly reduces infarction and improves sensorimotor function, possibly by suppressing early proinflammatory cytokines and promoting reparative cytokines and growth factors.

Hemodynamic, Biochemical, and Ventilatory Parameters are Independently Associated with Outcome after Cardiac Arrest.

Hypotension, hyperglycemia, dysoxia, and dyscarbia may contribute to reperfusion injury, and each is independently associated with poor outcome (PO) after cardiac arrest. We investigated whether the combined effects of these physiological derangements are associated with cardiac arrest outcomes. This institutional review board-approved retrospective cohort study included consecutive resuscitated cardiac arrest patients that received targeted temperature management at Maine Medical Center from 2013 to 2015. We abstracted demographics, intra-arrest factors, and physiological parameters. The primary outcome was dichotomized cerebral performance category (CPC 1-2 vs 3-5) at hospital discharge. After comparing demographics, clinical factors, and persistent post-arrest physiological derangements in patients with good and PO, we constructed a logistic regression model comprised of clinical and demographic factors separately associated with severity, and physiology variables, attempting to evaluate the independent effects of persistent physiological derangements on outcome. Sixty-eight of 222 (31%) patients had CPC 1-2 (good outcome [GO]) at discharge. In bivariate analysis, factors associated with PO included increased time from collapse to resuscitation, non-shockable rhythm, and age-combined Charlson comorbidity index. In multivariate analysis, each persistent physiological derangement incrementally decreased the likelihood of GO [OR GO per derangement 0.71 (interquartile range [IQR] 0.51-0.99), p=0.042, area under the curve (AUC) for final model 0.769]. Uncorrected physiological derangements in the first 24h after cardiac arrest are independently associated with PO. Although causality cannot be established, these findings support preclinical models suggesting that aggressive normalization of physiology after resuscitation may be a reasonable strategy to decrease reperfusion injury.

Hypothermic oxygenated perfusion (HOPE) for fatty liver grafts in rats and humans

Pretreatment of marginal organs by perfusion is a promising opportunity to make more organs available for transplantation. Protection of human donation after cardiac death (DCD) livers by a novel machine perfusion technique, hypothermic oxygenated perfusion (HOPE), was recently established. Herein, we tested whether HOPE is also useful for fatty liver grafts, using a rodent transplant model. Rats were fed over three weeks with a special methionine-choline-deficient diet (MCDD) to induce severe hepatic macrosteatosis (≥60%). Afterwards, livers were transplanted with either minimal or 12h cold storage. Additional liver grafts were treated after 12h cold storage with 1h HOPE before transplantation. Graft injury after orthotopic liver transplantation (OLT) was assessed in terms of oxidative stress, damage-associated molecular patterns release, toll-like receptor-4 activation, cytokine release, endothelial activation, and the development of necrosis and fibrosis. Implantation of cold stored macrosteatotic liver grafts induced massive reperfusion injury after OLT, compared to controls (non-fatty livers). HOPE treatment after cold storage failed to change the degree of steatosis itself, but markedly decreased reperfusion injury after OLT, as detected by less oxidative stress, less nuclear injury, less Kupffer- and endothelial cell activation, as well as less fibrosis within one week after OLT. Protective effects were lost in the absence of oxygen in the HOPE perfusate. HOPE after cold storage of fatty livers prevents significant reperfusion injury and improves graft function, comparable to the effects of HOPE in DCD livers and DCD kidneys. HOPE treatment is easy and may become a universal concept to further expand the donor pool. An increasing number of donor livers contain fat. It is important to harness marginal livers, which may contain fat, as the stock of donor livers is limited. Hypothermic oxygenated perfusion (HOPE) prevents reperfusion injury and improves liver graft function. HOPE offers a simple and low-cost option for treating liver grafts in transplant centers, even after cold storage, instead of transporting machines to the place of procurement. HOPE could be used globally to expand the donor pool.

Acid-base optimization during hypothermia.

Cardiac arrest (CA) often results in hemodynamic and metabolic compromise with associated poor prognosis. Therapeutic hypothermia (TH) has become the standard of care for CA survivors, decreasing reperfusion injury and intercellular acid-base disturbances, with improved neurologic outcomes. These benefits are realized despite a mild acidosis that can potentially occur during TH. By contrast, the severity of acidosis after return of spontaneous circulation (ROSC) must be monitored carefully and managed appropriately. Bicarbonate should be used only in case of severe acidosis and as a continuous infusion. The blood gas samples are usually warmed to 37°C before analysis; hence, it is worth noting that the blood gas values are temperature dependent. Therefore, a calculated correction for values may be necessary.

Systematic Review and Meta-Analysis of the Efficacy of Sphingosine-1-Phosphate (S1P) Receptor Agonist FTY720 (Fingolimod) in Animal Models of Stroke

ABSTRACTBackground and Purpose: FTY720 (fingolimod) is a known sphingosine-1-phosphate (S1P) receptor agonist, which has been used in clinical trials for treating multiple sclerosis, renal transplantation, and decreasing reperfusion injury in heart, liver, and kidney. Most of these clinical trials have showed a positive effect. Especially, the trials of MS showed a reduction of relapse rate in FTY720-treated patients. Now, some animal experiments indicated that FTY720 could be a new compound available treatment for stroke patients by exerting neuroprotection via S1P1 mediated antiapoptotic mechanisms. Whether it could be effective in animals is unclear, so we conducted a systematic review to make it clear. Methods: We conducted a systematic review and meta-analysis of the efficacy of FTY720 in animal models of focal cerebral ischemia by electronic and manual searches of the literature. Data on study quality, FTY720 dose, time of administration, and outcome measured as infarct volume or functional deficit were extracted. Data from all studies were analyzed by means of a standardized mean difference meta-analysis. Results: Of the 19 identified studies, 9 were included. Among all the included studies, 178 animals were calculated for infarct size and 194 animals were assessed of neurological deficits. The methodological quality of the studies ranged from 2 to 10 according to a published 11-item quality scale. Of the nine studies selected, only one reported a negative result of FTY720. The result indicated that FTY720 reduced the infarct volume (SMD = −1.31, 95% CI −1.99 to −0.63) and improve the functional outcome (SMD = −1.61,95% CI −2.17 to −1.05). Conclusions: The data we included supporting FTY720 was a candidate drug for stroke, but it should be considered with caution. More good quality experimental studies should be performed to evaluate the safety of FTY720 in the future. Whether FTY720 is effective in aged animals that mimicked human with comorbidities like diabetes and hypertension should also be deliberated.

Abstract 185: Whole Body Periodic Acceleration (pGz) in Mice Increases Antioxidant Enzymes in the Heart

BACKGROUND: Mild to moderate exercise produces cardio and neuroprotection. Exercise increases pulsatile shear stress that provide several cytoprotective and signaling substances from muscle and endothelium. Some beneficial effects of exercise on cytoprotection are derived from increased antioxidant capacity. pGz is a passive exercise strategy that moves the body in a sinusoidal head to foot motion, adding pulses to the circulation thereby increasing pulsatile, cyclical shear stress to the endothelium. pGz activates the PI3/Akt eNOS pathway and increases release of eNO and its enzyme (eNOS), and other cytoprotective signaling and substances, and decreases reperfusion injury. Optimum frequency of pGz has been established in our laboratory for various animal species and humans. We tested the hypothesis that repeated exposure to pGz in mice would increase antioxidant enzymes, and determined its time course. METHODS: Acclimated awake mice (n=48) where exposed to shear stress at f ƒ=480 cpm, Gz ± 3mt/sec 2 (pGz) 1 hr daily sessions of 1, 2, and 4 weeks of pGz. Myocardial tissue was harvested prior to pGz (BL) and 24 hrs after the last pGz treatment for proteins via Western Blot analyses: Glutathioneperoxidase-1(GPX1), Superoxide Dismutase -1 (Cu-Zn SOD), Superoxide Dismutase -2 (mitochondrial or Mn-SOD) and Catalase (Cat) , Protein content was normalized to GAPDH. RESULTS:One to four weeks of pGz increased GPX1, Cat and SOD1 from BL levels, whereas SOD2 remained unchanged. 4 weeks of pGz increased GPX1, Cat and SOD1, by 110%, 80% and 240% of basal enzyme levels. (Data= mean ± SEM, Figure † p&lt; 0.01 BL vs. 1,2,4 wks) CONCLUSIONS: One hour daily pGz increases cardiac antioxidant enzymes and may serve as a method to augment endogenous antioxidant capacity, and decrease reperfusion injury.

Myocardial Preservation: Controlled Reperfusion

Reperfusion injury after reestablishing coronary flow by releasing the aortic cross clamp after cardiac surgery with cardioplegic arrest causes myocardial damage and even death. Attenuation of this reperfusion response by controlling the biochemical and physical environment can avoid morbidity and mortality. Use of a warm reperfusate with addition of drugs that are known to decrease reperfusion injury with manipulation of coronary vascular resistance and the physical parameters of the reperfusion environment helps the heart to reestablish coronary perfusion while decreasing the harm produced by the period of ischemia that occurs during cardiac surgery with intermittent cardioplegic arrest.

The combination therapy of cilostazol and ischemic postconditioning protects against ischemia-reperfusion injury in non-diabetic and diabetic rat hearts with long-term atorvastatin treatment

ObjectiveThis study aims to investigate if the combination therapy of cilostazol and ischemic postconditioning (Ipost) could protect the non-diabetic and diabetic rat hearts against reperfusion injury after long-term atorvastatin treatment.MethodsNon-diabetic...

The combination therapy of cilostazol and ischemic postconditioning protects against ischemia-reperfusion injury in non-diabetic and diabetic rat hearts with long-term atorvastatin treatment

ObjectiveThis study is to investigate if the combination therapy of cilostazol and ischemic postconditioning (Ipost) could protect the non-diabetic and diabetic rat hearts against reperfusion injury after long-term atorvastatin treatment.MethodsNon-diabetic...

Glucagon-like Peptide-1 (GLP-1), Immediately Prior to Reperfusion, Decreases Neutrophil Activation and Reduces Myocardial Infarct Size in Rodents

Glucagon-like peptide-1 (GLP-1) is an incretin that has glucoregulatory effects as well as protective effects in a variety of tissues, including the heart. We hypothesized that GLP-1 may have a direct effect on neutrophils (PMNs) after myocardial ischemia, to ameliorate reperfusion injury. Deeply anesthetized Sprague-Dawley rats underwent 30 min of left coronary artery occlusion followed by 120 min of reperfusion. Immediately prior to reperfusion, rats were treated with either GLP-1 (human rGLP-1, 30 pM/kg/min) or PBS as placebo. GLP-1 significantly decreased myocardial infarct size [73.2±11.7% INF/AAR in PBS (n=4) vs. 15.7 ±5.52% INF/AAR in GLP-1-treated animals (n=5), p<0.05], PMN activation in blood in vivo (fMLP-stimulated CD11b surface expression: PBS 2.78±1.14 vs. GLP-1 1.7±0.21, TFI, p<0.05), and accumulation in myocardium (PBS: 6.52±0.31 vs. GLP-1: 4.78±0.90, n=4-6 animals/group, p<0.05). In addition, we found that GLP-1 mitigated PMN CD11b surface expression in whole rat blood in vitro, an effect that was abolished by GLP-1 receptor blockade (PBS 6.52±0.31 vs. GLP-1 4.78±0.90, TFI, p<0.05). These findings suggest that one mechanism by which GLP-1 decreases reperfusion injury may be the attenuation of PMN-mediated reperfusion injury.

Fingolimod provides long‐term protection in rodent models of cerebral ischemia

The sphingosine-1-phosphate (S1P) receptor agonist fingolimod (FTY720), that has shown efficacy in advanced multiple sclerosis clinical trials, decreases reperfusion injury in heart, liver, and kidney. We therefore tested the therapeutic effects of fingolimod in several rodent models of focal cerebral ischemia. To assess the translational significance of these findings, we asked whether fingolimod improved long-term behavioral outcomes, whether delayed treatment was still effective, and whether neuroprotection can be obtained in a second species. We used rodent models of middle cerebral artery occlusion and cell-culture models of neurotoxicity and inflammation to examine the therapeutic potential and mechanisms of neuroprotection by fingolimod. In a transient mouse model, fingolimod reduced infarct size, neurological deficit, edema, and the number of dying cells in the core and periinfarct area. Neuroprotection was accompanied by decreased inflammation, as fingolimod-treated mice had fewer activated neutrophils, microglia/macrophages, and intercellular adhesion molecule-1 (ICAM-1)-positive blood vessels. Fingolimod-treated mice showed a smaller infarct and performed better in behavioral tests up to 15 days after ischemia. Reduced infarct was observed in a permanent model even when mice were treated 4 hours after ischemic onset. Fingolimod also decreased infarct size in a rat model of focal ischemia. Fingolimod did not protect primary neurons against glutamate excitotoxicity or hydrogen peroxide, but decreased ICAM-1 expression in brain endothelial cells stimulated by tumor necrosis factor alpha. These findings suggest that anti-inflammatory mechanisms, and possibly vasculoprotection, rather than direct effects on neurons, underlie the beneficial effects of fingolimod after stroke. S1P receptors are a highly promising target in stroke treatment.

Urinary trypsin inhibitor: miraculous medicine in many surgical situations?

Urinary trypsin inhibitor: miraculous medicine in many surgical situations?

Dipropionylcysteine Ethyl Ester Compensates for Loss of Citric Acid Cycle Intermediates During Post Ischemia Reperfusion in the Pig Heart

During reperfusion, following myocardial ischemia, uncompensated loss of citric acid cycle (CAC) intermediates may impair CAC flux and energy transduction. Propionate has an anaplerotic effect when converted to the CAC intermediate succinyl-CoA, and may improve contractile recovery during reperfusion. Antioxidant therapy with N-acetylcysteine decreases reperfusion injury. To synergize the antioxidant effects of cysteine with the anaplerotic effects of propionate, we synthesized a novel bi-functional compound, N,S-dipropionyl cysteine ethyl ester (DPNCE) and tested its anaplerotic and anti-oxidative capacity in anesthetized pigs. Ischemia was induced by a 70% reduction in left anterior descending coronary artery flow for one hour, followed by 1 h of reperfusion. After 30 min of ischemia and throughout reperfusion animals were treated with saline or intravenous DPNCE (1.5 mg x kg(-1) x min(-1), n = 8/group). Arterial concentrations and myocardial propionate, cysteine, free fatty acids, glucose and lactate uptakes, cardiac mechanical functions, myocardial content of CAC intermediates and oxidative stress were assessed. Ischemia resulted in reduction in myocardial tissue concentration of CAC intermediates. DPNCE treatment elevated arterial propionate and cysteine concentrations and myocardial propionate uptake, and increased myocardial concentrations of citrate, succinate, fumarate, and malate compared to saline treated animals. DPNCE treatment did not affect blood pressure or myocardial contractile function, but increased arterial free fatty acid concentration and myocardial fatty acid uptake. Arterial cysteine concentration was elevated by DPNCE, but there was negligible myocardial cysteine uptake, and no change in markers of oxidative stress. DPNCE elevated arterial cysteine and propionate, and increased myocardial concentration of CAC intermediates, but did not affect mechanical function or oxidative stress.

Nonanticoagulant heparin reduces myocyte Na+and Ca2+loading during simulated ischemia and decreases reperfusion injury

Heparin desulfated at the 2-O and 3-O positions (ODSH) decreases canine myocardial reperfusion injury. We hypothesized that this occurs from effects on ion channels rather than solely from anti-inflammatory activities, as previously proposed. We studied closed-chest pigs with balloon left anterior descending coronary artery occlusion (75-min) and reperfusion (3-h). ODSH effects on [Na(+)](i) (Na Green) and [Ca(2+)](i) (Fluo-3) were measured by flow cytometry in rabbit ventricular myocytes after 45-min of simulated ischemia [metabolic inhibition with 2 mM cyanide, 0 glucose, 37 degrees C, pacing at 0.5 Hz; i.e., pacing-metabolic inhibition (PMI)]. Na(+)/Ca(2+) exchange (NCX) activity and Na(+) channel function were assessed by voltage clamping. ODSH (15 mg/kg) 5 min before reperfusion significantly decreased myocardial necrosis, but neutrophil influx into reperfused myocardium was not consistently reduced. ODSH (100 microg/ml) reduced [Na(+)](i) and [Ca(2+)](i) during PMI. The NCX inhibitor KB-R7943 (10 microM) or the late Na(+) current (I(Na-L)) inhibitor ranolazine (10 microM) reduced [Ca(2+)](i) during PMI and prevented effects of ODSH on Ca(2+) loading. ODSH also reduced the increase in Na(+) loading in paced myocytes caused by 10 nM sea anemone toxin II, a selective activator of I(Na-L). ODSH directly stimulated NCX and reduced I(Na-L). These results suggest that in the intact heart ODSH reduces Na(+) influx during early reperfusion, when I(Na-L) is activated by a burst of reactive oxygen production. This reduces Na(+) overload and thus Ca(2+) influx via NCX. Stimulation of Ca(2+) extrusion via NCX later after reperfusion may also reduce myocyte Ca(2+) loading and decrease infarct size.