Initiation Of Ischemia Research Articles

Effects of adrenaline and vasopressin on cerebral microcirculation at baseline and during global brain ischemia and reperfusion in rabbits

BackgroundDuring cardiopulmonary resuscitation, the brain becomes ischemic. Adrenaline and vasopressin have been recommended for use during cardiopulmonary resuscitation. We aimed to investigate the direct effects of adrenaline and vasopressin on the cerebral microvasculature at baseline and during ischemia and reperfusion in rabbits.MethodsThe closed cranial window method was used to visualize the cerebral microcirculation and changes in the pial arteriole diameter in rabbits. Adrenaline and vasopressin were administered topically on the brain tissue. First, the effects of adrenaline and vasopressin on pial arterioles were evaluated in 7 rabbits that were given 4 different concentrations of adrenaline, and another 7 rabbits that received 4 different concentrations of vasopressin. Second, the effects of adrenaline and vasopressin were determined during the global brain ischemia and reperfusion, which was induced by clamping the brachiocephalic, left common carotid, and left subclavian arteries for 15 min. An additional 21 rabbits were randomly assigned to receive artificial cerebrospinal fluid (aCSF) (n = 7), adrenaline 10–5 mol/L (n = 7), or vasopressin 10–7 mol/L (n = 7). Each drug was continuously infused from 5 min after the initiation of ischemia until 120 min after reperfusion. The pial arteriole diameters were recorded before and during ischemia, and after reperfusion.ResultsAt baseline, adrenaline and vasopressin did not affect the cerebral pial arterioles. During ischemia, vasopressin, but not aCSF and adrenaline constricted the pial vessels. Late in the reperfusion phase, pial diameter became reduced in the vasopressin and aCSF groups whereas pial diameter was higher in the animals treated with adrenaline.ConclusionsAdrenaline and vasopressin did not affect pial arterioles at baseline. During reperfusion, adrenaline may counteract the cerebral vasoconstriction.

Ischemia reperfusion-induced metastasis is resistant to PPARγ agonist pioglitazone in a murine model of colon cancer

Ischemia reperfusion injury (IRI) during liver-metastasis resection for treatment of colon cancer may increase the risk of further metastasis. Peroxisome proliferator-activated receptor-γ (PPARγ) activation has been observed to exert a protective effect against IRI and IRI-induced metastasis of hepatocellular carcinoma. The present study aimed to investigate the effect of the PPARγ agonist pioglitazone on tumor metastasis and liver injury following IRI in a mouse model of colon cancer. Pioglitazone (30 mg/kg weight) was administered orally 1.5 h before and 2 h after the initiation of ischemia and was orally administrated daily to mice from day 0–21. SL4-cancer cells expressing red fluorescent protein (SL4-RFP) (1 × 106) were injected into the spleen. Fifteen minutes after injection, the hepatoduodenal ligament was clamped with a vessel clip, and released 5 min later. Liver, blood and tumor samples were taken from mice in order to determine if inflammation was induced by IRI. The effect of pioglitazone on liver metastasis was assessed. Furthermore, the effect of pioglitazone to control the inflammatory response during IRI progression was examined. Liver metastasis along with MMP-9 activation and the production of inflammatory cytokines were resistant to pioglitazone. Our results indicate that liver metastasis and associated inflammation in mice were resistant to pioglitazone.

Effects of \u03b21-adrenergic receptor blockade on the cerebral microcirculation in the normal state and during global brain ischemia/reperfusion injury in rabbits

BackgroundAlthough recent studies using experimental models of ischemic brain injury indicate that systemically-administered β1-blockers have potential protective effects on the cerebrovascular system, the precise mechanisms remain unclear. In addition to their cardiovascular effects, water-soluble β1-blockers can pass the blood–brain barrier and may exert their vascular action on cerebral microvessels. The aim of this study was to investigate the direct effects of β1-blockade on the cerebral microvasculature both in the normal state and ischemia/reperfusion state using the cranial window method.MethodsThe closed cranial window method was used to visualize the cerebral microcirculation and changes in the pial arteriole diameter in adult male rabbits. In the first experiment, various concentrations of the selective β1-blocker landiolol were administered into the cranial window to evaluate the dose-response. In the second experiment, the effect of β1-blockade on the brain during ischemic/reperfusion injury was investigated. Global brain ischemia/reperfusion was induced by clamping the brachiocephalic, left common carotid, and left subclavian arteries for 15 min. Either landiolol or artificial cerebrospinal fluid was infused 5 min after initiation of ischemia through 120 min after reperfusion. Pial arteriole diameter and hemodynamic and physiological parameters were recorded before ischemia, during ischemia, and 5, 10, 20, 40, 60, 80, 100, and 120 min after reperfusion.ResultsIn the first experiment, topical administration of landiolol at higher concentrations produced slight pial arteriole dilation (10− 8 mol/L: 4.3 ± 3.4%, 10− 6 mol/L: 8.0 ± 5.8%, 10− 4 mol/L: 7.3 ± 4.0%). In the second experiment, the topical administration of landiolol significantly dilated the pial arteriole diameters during ischemia/reperfusion injury (ischemia: 30.6 ± 38.6%, 5 min: 47.3 ± 42.2%, 10 min: 47.8 ± 34.2%, 20 min: 38.0 ± 39.0%). There were no statistical differences in hemodynamic and physiological parameters between the landiolol and control groups.ConclusionsThe blockade of β1-adrenergic receptors induced significant vasodilation of pial arterioles during ischemia/reperfusion injury. By contrast, only a slight dilation of the arterioles was observed in the normal state, indicating that ischemic cerebral microvessels are more susceptible to the vasodilatory effect induced by selective blockade of β1-adrenergic receptors than normal microvessels.

The GTN patch: a simple and effective new approach to cardioprotection?

There remains a significant un-met need to reduce the extent of myocardial injury caused by ischaemia and reperfusion injury in patients experiencing an ST-elevation MI. Although nitric oxide is central to many cardioprotective strategies currently undergoing investigation, cardioprotection from the delivery of nitrates/nitrites has been inconsistently observed. The route of administration appears to be a critical variable. The glyceryl trinitrate (GTN) patch is commonly used as a simple and practical means of delivering nitric oxide to patients with ischaemic heart disease, but whether acute cardioprotection can be achieved by application of a GTN patch has not been investigated before. Here, we use a mouse model to demonstrate that a GTN patch is highly cardioprotective when applied immediately prior to 40 min occlusion of the left anterior coronary artery followed by 2 h reperfusion, reducing infarct size from 54 ± 4% in control mice, to 28 ± 4% (P < 0.001, N = 7). The degree of protection was similar to that achieved with a standard remote ischaemic preconditioning protocol. Furthermore, and of greater potential clinical relevance, a GTN patch was also protective when applied well after the initiation of ischaemia and 15 min prior to reperfusion (28 ± 4 vs 59 ± 4%; P < 0.01, N = 5). Confirmatory experiments verified the expected effect increase in plasma nitrite levels and decrease in blood pressure. The simplicity and rapidity of GTN patch application (easily applied in an ambulance or cardiac catheterization laboratory), and low cost (potentially relevant to low-income countries), make it attractive for further investigation.

Экспериментальное изучение применения церулоплазмина для предотвращения ишемически-реперфузионного повреждения в кожном лоскуте

Цель. Изучить в эксперименте эффективность церулоплазмина, применяемого для полного устранения или минимизации ишемически-реперфузионного повреждения, вызывающего некроз кожного лоскута (КЛ).&#x0D; Материалы и методы. Исследования проведены на 32 белых крысах-самцах линии Sprague Dawley массой тела в среднем 250 г. Животные разделены на 2 группы: группа I (контрольная, n = 8) - интактные животные; группа II (экспериментальная, n = 24). В свою очередь группа II разделена еще на три группы: IIА, IIБ и IIВ. Животным группы IIА церулоплазмин не применяли, ишемию не создавали, но забор КЛ производили. Животным группы IIБ церулоплазмин также не применяли. Этим животным за 30 мин до создания ишемии внутрибрюшинно делали инъекцию сыворотки. Через 30 мин после инъекции на эпигастральные вены и артерии с двух сторон накладывали клипсы на 10 ч. Через 10 ч после появления в лоскуте признаков ишемии клипсы извлекали, артериовенозный кровоток восстанавливался. После этого КЛ проленовыми нитками пришивали к своему месту. Животным группы IIВ за 30 мин до ишемии внутрибрюшинно вводили 60 мг/кг церулоплазмина. После инъекции церулоплазмина на эпигастральные артерии и вены с двух сторон накладывали клипсы, которые по истечении 10 ч извлекали.&#x0D; Результаты. Некротический участок КЛ в группе IIА составил в среднем (1,8 ± 0,6) см, IIБ - (22,4 ± 3,6) см, IIВ - (4,1 ± 1,1) см.

CARDIOPROTECTIVE EFFECT OF INM 176 ON ISCHEMIA REPERFUSION INJURY USING RAT HEART MODEL

Objective: Cardioprotective effect of INM-176 in ischemia reperfusion (I/R) injury in rat heart was evaluated by measuring the extent of release of the enzyme lactate dehydrogenase (LDH) and creatine phosphokinase (CK) and by measuring the myocardial infarction (MI) size.Methods: Five groups of animals were employed. Animals were sacrificed, and heart was mounted over the Langendorff’s apparatus. Group I was employed as control group, Group II was the myocardial ischemic (MI) group, and Group III, IV, and V were pre-treated with different doses of INM- 176 (150, 300, and 600 mg/kg). After this, global ischemia accompanied by reperfusion for 30 and 120 minutes was given, respectively. The effluent was collected from coronary vessel before the initiation of ischemia, immediately, after 5 and 30 minutes of reperfusion. The magnitude of myocardial injury was measured by LDH and CK release in coronary effluent. MI size was also evaluated macroscopically using triphenyl tetrazolium chloride staining.Results: In the present study, INM-176 was found to produce a dose-dependent cardioprotective effect against I/R injury, as observed by a decrease in the level of LDH, CK, and infarct size. Pre-treatment with a dose of 600 mg/kg was found to be more effective as compared to 150 and 300 mg/kg dose of INM-176.Conclusion: INM-176 pre-treatment at a dose of 600 mg/kg provides cardioprotection against I/R-induced myocardial injury.

Evidence of CCR2-independent transmigration of Ly6Chi monocytes into the brain after permanent cerebral ischemia in mice

Previously we showed that INCB3344, a CCR2 antagonist, inhibits transmigration of Ly6Chi monocytes into the brain after ischemia-reperfusion. Here we tested the effect of CCR2 inhibition during permanent cerebral ischemia. Mice were administered either vehicle (dimethyl sulfoxide/carboxymethylcellulose) or INCB3344 (30 or 100mg/kg IP) 1h before middle cerebral artery occlusion and at 2 and 6h after the initiation of ischemia. After 24h, we assessed functional outcome, infarct volume and quantified immune cells in blood and brain. The increase in circulating bone marrow-derived Ly6Chi monocytes, but not the infiltration of those cells into the brain, was blocked by the CCR2 antagonist. INCB3344 had no effect on either neurological deficit or infarct volume. Our data confirm that cerebral ischemia triggers a CCR2-dependent increase in circulating Ly6Chi monocytes, but suggest that in the absence of reperfusion these cells may transmigrate into the ischemic brain in a CCR2-independent manner.

Survival of Transferred Ileum after Ischemia Time Longer than 1 Hour: A Clinical Result Different from Animal Studies

The aim of this study was to identify the ischemic tolerance of the ileum. In microvascular transfer of autologous bowel segments, the most critical factor for survival is ischemic time. In earlier animal studies, the tolerance of ischemic time was shorter for the ileum than for the jejunum, and an ischemic time of <1 hour was suggested for microvascular transfer of the ileum. It was believed that there are more bacteria in the ileum than in the jejunum and therefore autolysis and necrosis will be triggered sooner after the initiation of ischemia. However, in a clinical scenario, the tolerance for ischemic time of the ileum has not yet been clarified. From 1998 to 2011, eight-four cases of microvascular transfer of intestine containing a segment of the ileum were reviewed. Data collected included the ischemia time during surgery, re-exploration, survival, complications, and postoperative functions. Multivariate analysis with exact logistic regression was used to identify the correlation between the ischemic time and necrosis of the transferred segment, as well as other complications. For segmental ileum transfer, the ischemic time >1 hour (but within 2 hours) at room temperature is not a risk factor for flap loss or complications. This contradicts data from animal studies in the literature. Clinically, the ileum segments can tolerate ischemia well within 2 hours. Segmental ileum transfer can be more widely applied in other fields. Care in each step of transfer is mandatory for functional success.

Effect of lidocaine on dynamic changes in cortical reduced nicotinamide adenine dinucleotide fluorescence during transient focal cerebral ischemia in rats

Rats were subjected to 90min of focal ischemia by occluding the left middle cerebral and both common carotid arteries. The dynamic changes in the formation of brain ischemic areas were analyzed by measuring the direct current (DC) potential and reduced nicotinamide adenine dinucleotide (NADH) fluorescence with ultraviolet irradiation. In the lidocaine group (n=10), 30min before ischemia, an intravenous bolus (1.5mg/kg) of lidocaine was administered, followed by a continuous infusion (2mg/kg/h) for 150min. In the control group (n=10), an equivalent amount of saline was administered. Following the initiation of ischemia, an area of high-intensity NADH fluorescence rapidly developed in the middle cerebral artery territory in both groups and the DC potential in this area showed ischemic depolarization. An increase in NADH fluorescence closely correlated with the DC depolarization. The blood flow in the marginal zone of both groups showed a similar decrease. Five minutes after the onset of ischemia, the area of high-intensity NADH fluorescence was significantly smaller in the lidocaine group (67% of the control; P=0.01). This was likely due to the suppression of ischemic depolarization by blockage of voltage-dependent sodium channels with lidocaine. Although lidocaine administration did not attenuate the number of peri-infarct depolarizations during ischemia, the high-intensity area and infarct volume were significantly smaller in the lidocaine group both at the end of ischemia (78% of the control; P=0.046) and 24h later (P=0.02). A logistic regression analysis demonstrated a relationship between the duration of ischemic depolarization and histologic damage and revealed that lidocaine administration did not attenuate neuronal damage when the duration of depolarization was identical. These findings indicate that the mechanism by which lidocaine decreases infarct volume is primarily through a reduction of the brain area undergoing NADH fluorescence increases which closely correlates with depolarization.

A Comparative Study of Neuroprotective Effect of Single and Combined Blockade of AT1 Receptor and PARP-1 in Focal Cerebral Ischaemia in Rat

Cerebral ischaemia results in enhanced expression of type 1 angiotensin receptor and oxidative stress. Free radicals due to oxidative stress lead to excessive DNA damage causing overactivation of poly (ADP-ribose) polymerase-1 resulting in neuronal death. Activation of both type 1 angiotensin receptors and poly (ADP-ribose) polymerase-1 following cerebral ischaemia takes place simultaneously, but until now, no study has explored the effect of combined blockade of both angiotensin type 1 angiotensin receptor and poly (ADP-ribose) polymerase-1 in cerebral ischaemia. Our purpose was to compare the effect of single and combined treatment with angiotensin type 1 angiotensin receptor blocker, candesartan, and the poly (ADP-ribose) polymerase-1 inhibitor, 1, 5 isoquinolinediol, on brain damage and oxidative stress in transient focal cerebral ischaemia in rats. Transient focal cerebral ischaemia was induced in Sprague-Dawley rats by an intraluminal technique for two-hours following 48 h of reperfusion. Candesartan (0·05 mg/kg) was administered just after initiation of ischaemia followed by a repeat administration at 24 h while 1, 5 isoquinolinediol (0·1 mg/kg) was given one-hour after of ischaemia. After 24 h of reperfusion, neurological deficit was evaluated in the different treatment groups. After 48 h of reperfusion, the rats were sacrificed and the brain was isolated. Ischaemic brain damage by 2,3,5 triphenyl tetrazolium chloride staining, oxidative stress markers, and levels of reactive oxygen species were determined biochemically. Single treatment with candesartan and 1, 5 isoquinolinediol significantly reduced neurological deficit, infarct, and oedema volume as compared to ischaemic control and different vehicle groups for each of the drugs. However, treatment with candesartan + 1, 5 isoquinolinediol offered greater reduction in neurological deficit, cerebral infarct volume, and oedema as compared to single-drug treatments. Furthermore, treatment with candesartan + 1, 5 isoquinolinediol significantly decreased oxidative stress as compared to single treatments with each drug. The study suggests that blockade of either type 1 angiotensin receptor or poly (ADP-ribose) polymerase-1 alone provides neuroprotection, but the better result was achieved when both type 1 angiotensin receptor and poly (ADP-ribose) polymerase-1 were blocked together by the combined use of their pharmacological inhibitor in transient cerebral ischaemia in rat.

The silence of the limbs

This editorial refers to ‘Silencing of Int6 gene restores function of the ischaemic hindlimb in a rat model of peripheral arterial disease’ by N. Okamoto et al ., pp. 209–217, this issue. Okamoto et al. 1 report that Int6 levels are reduced by direct administration of silencing small-interfering RNA (siRNA) into the hindlimb (adductor) muscles immediately following ligation of the external iliac artery. Ligation of the external iliac artery is a model of limb ischaemia. Ischaemia, whether in the brain, heart, or other tissue, is a complex metabolic injury. When a blood vessel is occluded, the underperfused area experiences a cascade of events beginning within 1 or 2 min and progresses for months. In tissue undergoing ischaemia, there is a gradient of injury that expands with time through space. The central area, where hypoxia is severe, becomes necrotic. With the initiation of ischaemia, as oxygen levels fall, ATP levels fall in the affected cells, and as minutes go by, ion pumps in the cell membranes stop. Various cell signalling pathways respond and transcription factors are activated and many changes occur in gene …

Sleep Disturbance Impairs Stroke Recovery in the Rat

There is a lack of experimental evidence to support the hypothesis that sleep may modulate stroke outcome as suggested by clinical observations. We have previously shown that sleep disturbance (SDis) over 3 days aggravates brain damage in a rat model of focal cerebral ischemia. The aim of this study is to further investigate effects of SDis on long-term stroke recovery and neuroplasticity as assessed by axonal sprouting, neurogenesis, and angiogenesis. Focal cerebral ischemia was induced by permanent occlusion of the distal branches of middle cerebral artery. Twelve hours after initiation of ischemia, SDis was performed over 3 consecutive days (deprivation of 80% sleep during the 12-h light phase). Weekly assessments on sensorimotor function by the single pellet reaching test (SPR) were performed for 5 weeks after surgery. Axonal sprouting was evaluated by anterograde tracing with biotinylated dextran amine (BDA) and neurogenesis/angiogenesis by bromodeoxyuridine (BrdU) labelling along with cell-type markers. Control groups included ischemia without SDis, sham with SDis, and sham without SDis. Basic sleep research laboratory. Rats subjected to SDis after ischemia showed significantly less recovery of forearm motor skills during the post-stroke period of 5 weeks. This effect was accompanied by a substantial reduction in axonal sprouting, expression of synaptophysin, and the ischemia-stimulated neural and vascular cell proliferation. SDis has detrimental effects on functional and morphological/structural outcomes after stroke, suggesting a role of sleep in the modulation of recovery processes and neuroplasticity.

Effective Pulmonary Artery Perfusion Mode during Cardiopulmonary Bypass

Reducing lung injury during cardiopulmonary bypass (CPB) is important for patients' recovery. The present study was designed to research convenient and effective pulmonary artery perfusion mode during CPB in an animal model. Twelve healthy mongrel dogs were randomly divided into 2 groups: a control group and a perfusion group designed to simulate clinical CPB-induced lung injury. During CPB, pulmonary artery perfusion with modified low-potassium dextran (LPD) solution was performed immediately after the initiation of ischemia and before reperfusion for 3 to 4 minutes each time, with pressure maintained at 15 to 20 mmHg; animals in the control group were not perfused. After pulmonary reperfusion, the changes in pulmonary function and tissue biochemical data were determined. Compared with the control group, lung compliance, oxygenation, and vascular resistance after reperfusion were significantly improved in the perfusion group. The malonaldehyde concentration, neutrophil sequestration ratio, and tissue water content also decreased significantly in the perfusion group. The pulmonary artery perfusion mode used in this experiment could relieve CPB-induced lung injury effectively. Improving cellular tolerance to hypoxia and decreasing inflammatory reaction may be the important mechanisms. Moreover, this mode is convenient and does not interfere with the intended operations, which is promising for clinical use.

Pharmaceutical Modulation of the Renin Angiotensin Aldosterone System for Stroke Prevention: A Review of Experimental and Clinical Evidence

The renin angiotensin aldosterone system (RAAS) has been implicated in the pathogenesis of cerebrovascular diseases. During the past decades, AU: INCLUDE A NUMBER? RAAS has gained attention as an important therapeutic target in cardiovascular medicine. Modulation of the RAAS for primary and secondary stroke prevention seems an appealing strategy. Several experimental studies have showed that pre-treatment with RAAS inhibitors prior to the initiation of ischemia exerts favorable effects on infarct volume, brain edema, and cerebral blood flow in the marginal zone. Furthermore, the activation of angiotensin receptor type 2 has been associated with neuroprotective effects. Accumulating evidence based on recent clinical trials indicate that blockade of RAAS has a potential role in stroke prevention. This review summarizes the pathophysiological aspects of brain RAAS and its constituents, presents experimental data on the neuroprotective actions of RAAS inhibitors and available evidence regarding their effects on stroke risk, and discuss future directions for research.

Connexin 43 mimetic peptide Gap26 confers protection to intact heart against myocardial ischemia injury

Unapposed connexin 43 hemichannels (Cx43Hc) are present on sarcolemma of cardiomyocytes. Whereas Cx43Hc remain closed during physiological conditions, their opening under ischemic stress contributes to irreversible tissue injury and cell death. To date, conventional blockers of connexin channels act unselectively on both gap junction channels and unapposed hemichannels. Here, we test the hypothesis that Gap26, a synthetic structural mimetic peptide deriving from the first extracellular loop of Cx43 and a presumed selective blocker of Cx43Hc, confers resistance to intact rat heart against ischemia injury. Langendorff-perfused intact rat hearts were utilized. Regional ischemia was induced by 40-min occlusion of the left anterior descendent coronary and followed by 180 min of reperfusion. Gap26 was applied either 10 min before or 30 min after the initiation of ischemia. Interestingly, myocardial infarct size was reduced by 48% and 55% in hearts treated with Gap26 before or during ischemia, respectively, compared to untreated hearts. Additionally, myocardial perfusate flow was increased in both groups during reperfusion by 37% and 32%, respectively. Application of Gap26 increased survival of isolated cardiomyocytes after simulated ischemia-reperfusion by nearly twofold compared to untreated cells. On the other hand, superfusion of tsA201 cells transiently expressing Cx43 with Gap26 caused 61% inhibition of Cx43Hc-mediated currents recorded using the patch clamp technique. In summary, we demonstrate for the first time that Cx43 mimetic peptide Gap26 confers protection to intact heart against ischemia-reperfusion injury whether administered before or after the occurrence of ischemia. In addition, we provide unequivocal evidence for the inhibitory effect of Gap26 on genuine Cx43Hc.

The Inter-Dependency of Local Myocardial Metabolism and Epicardial Electrical Activity during Acute Ischemia and Reperfusion

Metabolic changes caused by the lack of adequate coronary flow lead to short and long term disturbances in local activation sequences. Our goal has been to study short term disturbances using parallel fluorescence imaging of epicardial NADH (fNADH) and transmembrane potential (TMP). METHODS: Experiments were conducted using Langendorff-perfused rat hearts while controlling the rate of flow to the left anterior descending coronary artery (LAD). Acute regional ischemia was induced by stopping flow to the LAD, followed by a period of low-flow reperfusion with subsequent full-flow reperfusion. Changes in local epicardial conduction velocities, as well as the incidence and dispersion of epicardial breakthroughs, were analyzed with the corresponding local changes of fNADH. With this approach, conduction velocities and reentrant activity could be correlated with changes in fNADH. RESULTS: Regional ischemia led to a reduction in Purkinje fiber activity within the ischemic zone. Approx 4 minutes after the initiation of ischemia, conduction velocities increased within regions with elevated fNADH. Afterward, conduction velocities in the ischemic zone declined and were lowest in the center, eventually falling to values below 20 cm/sec. Reductions in conduction velocity lagged behind elevations of fNADH, both in time and space. During regional ischemia, occasional breakthroughs occurred, most of them along the boundary between ischemic and normoxic tissue. During low-flow reperfusion, the number of breakthroughs within the ischemic zone increased dramatically as well as the incidence of ventricular fibrillation (compared to ischemia and normal flow conditions). CONCLUSIONS: The inter-dependence of local activation patterns and local myocardial metabolism makes parallel imaging of fNADH and TMP an essential tool for understanding the mechanisms of arrhythmias caused by ischemia and reperfusion.

C-Jun DNAzymes Inhibit Myocardial Inflammation, ROS Generation, Infarct Size, and Improve Cardiac Function After Ischemia-Reperfusion Injury

Coronary reperfusion has been the mainstay therapy for reduced infarct size after a heart attack. However, this intervention also results in myocardial injury by initiating a marked inflammatory reaction, and new treatments are keenly sought. The basic-region leucine zipper protein, c-Jun is poorly expressed in the normal myocardium and is induced within 24 hours after myocardial ischemia-reperfusion injury. Synthetic catalytic DNA molecules (DNAzymes) targeting c-Jun (Dz13) reduce infarct size in the area-at-risk (AAR) regardless of whether it is delivered intramyocardially at the initiation of ischemia or at the time of reperfusion. Dz13 attenuates neutrophil infiltration, c-Jun and ICAM-1 expression in vascular endothelium, cardiomyocyte apoptosis, and the generation of reactive oxygen species in the reperfused myocardium. It inhibits infiltration into the AAR of complement 3 (C3), C3a receptor (C3aR), membrane attack complex-1 (Mac-1), or matrix metalloproteinase-2 (MMP-2) positive inflammatory cells. Dz13 also improves cardiac function without influencing myocardial vascularity or fibrosis. These findings demonstrate the regulatory role of c-Jun in the pathogenesis of myocardial inflammation and infarction following ischemia-reperfusion injury, and inhibition of this process using catalytic DNA.

Tamoxifen as a Powerful Neuroprotectant in Experimental Stroke and Implications for Human Stroke Therapy

Several recent studies from the author's laboratory have shown that tamoxifen, at higher concentrations than used for breast cancer and given i.v., can substantially prevent tissue infarct and behavioral deficits in reversible and permanent rat focal stroke models for up to at least 14 days after initiation of ischemia. Longer times and purely i.p. or oral administration have not yet been tried. Its marked effectiveness may be because it has several neuroprotective modes of action including free radical scavenging and, being highly lipid soluble, readily crosses the blood-brain barrier. Plus, it has a three hour therapeutic window. Thus it meets many of the STAIR criteria and should be a promising candidate for clinical use. However, a number of its positive effects were exhibited by the free radical trapping agent NXY-058, which also functions as a free radical scavenger. In two recently completed clinical trials (SAINT 1 and 2) NXY-058 had marginal positive effects and did not meet treatment criteria, respectively. Differences that may make tamoxifen still desirable and the problem of predicting clinical efficacy from successful animal studies are discussed.

Reperfusion activates metalloproteinases that contribute to neurovascular injury

In this study, we examine the effects of reperfusion on the activation of matrix metalloproteinase (MMP) and assess the relationship between MMP activation during reperfusion and neurovascular injury. Ischemia was produced using suture-induced middle cerebral artery occlusion in rats. The MMP activation was examined with in situ and gel zymography. Injury to cerebral endothelial cells and basal lamina was assessed using endothelial barrier antigen (EBA) and collagen IV immunohistochemistry. Injury to neurons and glial cells was assessed using Cresyl violet staining. These were examined at 3 h after reperfusion (8 h after initiation of ischemia) and compared with permanent ischemia at the same time points to assess the effects of reperfusion. A broad-spectrum MMP inhibitor, AHA ( p-aminobenzoyl-Gly-Pro- d-Leu- d-Ala-hydroxamate, 50 mg/kg intravenously) was administered 30 min before reperfusion to assess the roles of MMPs in activating gelatinolytic enzymes and in reperfusion-induced injury. We found that reperfusion accelerated and potentiated MMP-9 and MMP-2 activation and injury to EBA and collagen IV immunopositive microvasculature and to neurons and glial cells in ischemic cortex and striatum relative to permanent ischemia. Administering AHA 30 min before reperfusion decreased MMP-9 activation and neurovascular injury in ischemic cerebral cortex.

Heparin-Binding EGF-like Growth Factor Decreases Inflammatory Cytokine Expression After Intestinal Ischemia/Reperfusion Injury

Intestinal ischemia/reperfusion (I/R) injury is believed to be the major initiator of the systemic inflammatory response syndrome. As a result of intestinal I/R, the gut becomes a major source of inflammatory cytokine production. We have previously shown that heparin-binding EGF-like growth factor (HB-EGF) is cytoprotective after intestinal I/R and down-regulates pro-inflammatory cytokine production in vitro. We now examine the effects of HB-EGF on pro-inflammatory cytokine expression in vivo. Rats were randomized into three groups: sham-operated, superior mesenteric artery occlusion (SMAO) for 90 min followed by 8 h of reperfusion (I/R), and I/R with intraluminal administration of HB-EGF 25 min after the initiation of ischemia (I/R + HB-EGF). Serum was drawn at 2, 4, 6, and 8 h post reperfusion for determination of cytokine protein levels using a bioplex suspension array system. Additional animals underwent the same ischemic protocol followed by 30 and 60 min of reperfusion with harvesting of ileal mucosa. Ileal pro-inflammatory cytokine gene expression was determined using reverse transcriptase polymerase chain reaction (RT-PCR) with primers specific for TNF-alpha, IL-6, and IL-1beta. HB-EGF decreased TNF-alpha, IL-6, and IL-1beta serum protein levels at 4, 6, and 8 h after intestinal I/R injury. In addition, HB-EGF decreased local intestinal mucosal mRNA expression of TNF-alpha, IL-6, and IL-1beta 30 and 60 min after intestinal injury. We conclude that pro-inflammatory cytokine expression is increased both locally and in the systemic circulation after intestinal I/R and that the administration of HB-EGF significantly reduces intestinal I/R-induced pro-inflammatory cytokine expression in vivo.