Antioxidant activity of nitro derivative of aspirin against ischemia-reperfusion in hamster cheek pouch microcirculation

Aspirin that has been chemically combined with a nitric oxide (NO) donor (NCX-4016) has been shown to inhibit cyclooxygenase and prostaglandin generation while maintaining the inhibitory effects of aspirin. The possible role of reactive oxygen species (ROS) in the action of NCX-4016 in ischemia-reperfusion (I/R) has not been studied. Furthermore, we were interested in comparing the effects of a conventional NO donor [2,2'-hydroxynitrosohydrazino-bis-etanamine (DETA/NO)] and NCX-4016 at the microvascular level in the hamster cheek pouch visualized by using an intravital fluorescent microscopy technique. Microvascular injury was assessed by measuring diameter change, the perfused capillary length (PCL), and leukocyte adhesion. Animals were treated with NCX-4016 (100 mg/kg or 30 mg.kg(-1).day(-1) for 5 days po) or DETA-NO (0.5 mg/kg). Mean arterial blood pressure increased slightly but significantly after NCX-4016 treatment. During 5- and 15-min reperfusion, lipid peroxides in the systemic blood increased by 72 and 89% vs. baseline, respectively, and were still higher than in basal conditions after 30-min reperfusion in the I/R group. Pretreatment with NCX-4016 maintained ROS at normal levels; increased arteriolar diameter, blood flow, and PCL; and decreased leukocyte adhesion (P < 0.05). DETA-NO decreased ROS during 30-min reperfusion; however, later there was a significant increase during reperfusion. DETA-NO decreased leukocyte adhesion (P < 0.05) but microvascular permeability increased after 30 min of reperfusion. In conclusion, NCX-4016 attenuates oxidative stress and prevents arteriolar constriction during I/R, whereas DETA-NO increases lipid peroxides in the systemic blood and permeability after reperfusion.     

Attenuation of intestinal ischemia/reperfusion injury with sodium nitroprusside: studies on mitochondrial function and lipid changes

Reactive oxygen species have been implicated in cellular injury during ischemia/reperfusion (I/R). Mitochondria are one of the main targets of oxygen free radicals and damage to this organelle leads to cell death. Reports suggest that nitric oxide (NO) may offer protection from damage during I/R. This study has looked at the functional changes and lipid alteration to mitochondria during intestinal I/R and the protection offered by NO. It was observed that I/R of the intestine is associated with functional alterations in the mitochondria as suggested by MTT reduction, respiratory control ratio and mitochondrial swelling. Mitochondrial lipid changes suggestive of activation of phospholipase A(2) and phospholipase D were also seen after (I/R) mediated injury. These changes were prevented by the simultaneous presence of a NO donor in the lumen of the intestine. These studies have suggested that structural and functional alterations of mitochondria are prominent features of I/R injury to the intestine which can be ameliorated by NO.     

Cardioprotective role of endogenous hydrogen peroxide during ischemia-reperfusion injury in canine coronary microcirculation in vivo

We have recently demonstrated that endogenous H2O2 plays an important role in coronary autoregulation in vivo. However, the role of H2O2 during coronary ischemia-reperfusion (I/R) injury remains to be examined. In this study, we examined whether endogenous H2O2 also plays a protective role in coronary I/R injury in dogs in vivo. Canine subepicardial small coronary arteries (>or=100 microm) and arterioles (<100 microm) were continuously observed by an intravital microscope during coronary I/R (90/60 min) under cyclooxygenase blockade (n=50). Coronary vascular responses to endothelium-dependent vasodilators (ACh) were examined before and after I/R under the following seven conditions: control, nitric oxide (NO) synthase (NOS) inhibitor NG-monomethyl-L-arginine (L-NMMA), catalase (a decomposer of H2O2), 8-sulfophenyltheophylline (8-SPT, an adenosine receptor blocker), L-NMMA+catalase, L-NMMA+tetraethylammonium (TEA, an inhibitor of large-conductance Ca2+-sensitive potassium channels), and L-NMMA+catalase+8-SPT. Coronary I/R significantly impaired the coronary vasodilatation to ACh in both sized arteries (both P<0.01); L-NMMA reduced the small arterial vasodilatation (both P<0.01), whereas it increased (P<0.05) the ACh-induced coronary arteriolar vasodilatation associated with fluorescent H2O2 production after I/R. Catalase increased the small arterial vasodilatation (P<0.01) associated with fluorescent NO production and increased endothelial NOS expression, whereas it decreased the arteriolar response after I/R (P<0.01). L-NMMA+catalase, L-NMMA+TEA, or L-NMMA+catalase+8-SPT further decreased the coronary vasodilatation in both sized arteries (both, P<0.01). L-NMMA+catalase, L-NMMA+TEA, and L-NMMA+catalase+8-SPT significantly increased myocardial infarct area compared with the other four groups (control, L-NMMA, catalase, and 8-SPT; all, P<0.01). These results indicate that endogenous H2O2, in cooperation with NO, plays an important cardioprotective role in coronary I/R injury in vivo.     

Electrophoretic mobility of human red blood cells coated with poly(ethylene glycol)

Poly(ethylene glycol), abbreviated as PEG, was covalently attached to the surface of human red blood cells (RBC) and the effects of such coating on the regions near the cell's glycocalyx were explored by means of cell electrophoresis. RBC electrophoretic mobilities were measured, in polymer-free buffers of various ionic strengths, as functions of PEG molecular mass (3.35, 18.5, 35.0, 35.9 kDa), geometry, (linear or 8-arm branched) and polymer/RBC ratio during attachment. The results indicate marked decreases of the mobility (up to 85%) which were affected by polymer molecular mass and geometry. Since PEG is neutral and its covalent attachment only removes positively-charged amino groups on the cell membrane, such decreases of mobility likely reflect structural changes near and within the RBC glycocalyx. Experimental results were analyzed using an extended "hairy sphere" model to consider friction and thickness of the polymer layer. Calculated polymer layer thickness increased with molecular mass for linear PEGs and was less extended for a branched PEG of similar molecular mass. Friction within the polymer layer increased with polymer/RBC ratio and for the linear PEGs was inversely related to molecular mass; friction was greatest for the branched PEG. Our results are consistent with the effects of attached PEGs on RBC aggregation and surface antigenic site masking, and suggest the usefulness of electrophoretic mobility techniques for studies of bound neutral polymers.     

Vascular protection by estrogen in ischemia-reperfusion injury requires endothelial nitric oxide synthase

Estrogen increases nitric oxide (NO) production by inducing the activity of endothelial NO synthase (eNOS) (Simoncini et al. Nature 407: 538, 2000). Ischemia (30 min) and reperfusion (I/R) increased the number of adherent leukocytes and decreased their rolling velocities in mouse cremaster muscle venules with a strong dependence on wall shear rate. Minimum rolling velocity at approximately 5 min after the onset of reperfusion was accompanied by increased P-selectin expression. This preceded the peak in leukocyte adhesion (at 10-15 min). In untreated wild-type mice, I/R caused a decrease of leukocyte rolling velocity from 37 to 26 microm/s and a 2.0-fold increase in leukocyte adhesion. Both were completely abolished by 0.25 mg ip estrogen 1 h before surgery. In eNOS(-/-) mice, the decrease of leukocyte rolling velocity and increase in adhesion were similar but were only marginally improved by estrogen. We conclude that the protective effect of estrogen, as measured by leukocyte rolling and adhesion, is significantly reduced in eNOS(-/-) mice, suggesting that induction of eNOS activity is the major mechanism of vasoprotection by estrogen in this model.     

LOX-1 inhibition in myocardial ischemia-reperfusion injury: modulation of MMP-1 and inflammation

A recently identified lectin-like oxidized low-density lipoprotein receptor (LOX-1) mediates endothelial cell injury and facilitates inflammatory cell adhesion. We studied the role of LOX-1 in myocardial ischemia-reperfusion (I/R) injury. Anesthetized Sprague-Dawley rats were subjected to 60 min of left coronary artery (LCA) ligation, followed by 60 min of reperfusion. Rats were treated with saline, LOX-1 blocking antibody JXT21 (10 mg/kg), or nonspecific anti-goat IgG (10 mg/kg) before I/R. Ten other rats underwent surgery without LCA ligation and served as a sham control group. LOX-1 expression was markedly increased during I/R (P < 0.01 vs. sham control group). Simultaneously, the expression of matrix metalloproteinase-1 (MMP-1) and adhesion molecules (P-selectin, VCAM-1, and ICAM-1) was also increased in the I/R area (P < 0.01 vs. sham control group). There was intense leukocyte accumulation in the I/R area in the saline-treated group. Treatment of rats with the LOX-1 antibody prevented I/R-induced upregulation of LOX-1 and reduced MMP-1 and adhesion molecule expression as well as leukocyte recruitment. LOX-1 antibody, but not nonspecific IgG, also reduced myocardial infarct size (P < 0.01 vs. saline-treated I/R group). To explore the link between LOX-1 and adhesion molecule expression, we measured expression of oxidative stress-sensitive p38 mitogen-activated protein kinase (p38 MAPK). The activity of p38 MAPK was increased during I/R (P < 0.01 vs. sham control), and use of LOX-1 antibody inhibited p38 MAPK activation (P < 0.01). These findings indicate that myocardial I/R upregulates LOX-1 expression, which through p38 MAPK activation increases the expression of MMP-1 and adhesion molecules. Inhibition of LOX-1 exerts an important protective effect against myocardial I/R injury.     

Angiotensin II type 1 receptors and the intestinal microvascular dysfunction induced by ischemia and reperfusion

The acute phase of intestinal ischemia-reperfusion (I/R) injury is mediated by leukocytes and is characterized by oxidative stress and blood cell recruitment. Upregulation of angiotensin II type 1 receptors (AT1-R) has been implicated in the pathogenesis of conditions associated with oxidative stress. The AT1-R-antagonist Losartan (Los) attenuates leukocyte recruitment following I/R. However, the role of AT1-R in intestinal I/R injury and the associated platelet-leukocyte interactions remains unclear. The objective of this study was to define the contribution of AT1-R to I/R-induced blood cell recruitment in intestinal venules. Leukocyte and platelet adhesion were quantified by intravital microscopy in the small bowel of C57Bl/6 [wild-type (WT)] mice exposed to sham operation or 45 min of ischemia and 4 h of reperfusion. A separate WT group received Los for 7 days before gut I/R (WT-I/R + Los). AT1-R bone marrow chimeras that express AT1-R on the vessel wall but not blood cells also underwent I/R. Platelet and leukocyte adhesion as well as AT1-R expression in the gut microvasculature were significantly elevated after I/R. All of these responses were attenuated in the WT-I/R + Los group, compared with untreated I/R mice. A comparable abrogation of I/R-induced blood cell adhesion was noted in AT1-R bone marrow chimeras. I/R-induced platelet adhesion was unaltered in mice overexpressing Cu,Zn-SOD or mice deficient in NAD(P)H oxidase. These data suggest that although gut I/R upregulates endothelial expression of AT1-R, engagement of these angiotensin II receptors on blood cells is more important in eliciting the prothrombogenic and proinflammatory state observed in postischemic gut venules, through a superoxide-independent pathway.     

Reactive oxygen species mediate modification of glycocalyx during ischemia-reperfusion injury

The glycocalyx (Gcx) is a complex and poorly understood structure covering the luminal surface of endothelial cells. It is known to be a determinant of vascular rheology and permeability and may be a key control site for the vascular injuries caused by ischemia-reperfusion (I/R). We used intravital-microscopy to evaluate the effects of I/R injury on two properties of Gcx in mouse cremasteric microvessels: exclusion of macromolecules (anionic-dextrans) and intracapillary distribution of red blood cells (RBC). In this model, the Gcx is rapidly modified by I/R injury with an increase in 70-kDa anionic-dextran penetration without measurable effect on the penetration of 580-kDa anionic-dextran or on RBC exclusion. The effects of I/R injury appear to be mediated by the rapid production of reactive oxygen species (ROS) because they are ameliorated by the addition of exogenous superoxide dismutase-catalase. Intravenous application of allopurinol or heparin also inhibited the effects of I/R injury, and we interpret efficacy of allopurinol as evidence for a role for xanthine-oxidoreductase (XOR) in the response to I/R injury. Heparin, which is hypothesized to displace XOR from a heparin-binding domain in the Gcx, reduced the effects of I/R. The effects of I/R injury were also partially prevented or fully reversed by the intravascular infusion of exogenous hyaluronan. These data demonstrate: 1) the liability of Gcx during I/R injury; 2) the importance of locally produced ROS in the injury to Gcx; and 3) the potential importance of heparin-binding sites in modulating the ROS production. Our findings further highlight the relations between glycosaminoglycans and the pathophysiology of Gcx in vivo.     

Fibronectin-based masking molecule blocks platelet adhesion

Vessel wall extracellular matrix, which underlies the endothelium, is a potent stimulator of platelet adhesion and activation. Exposure of this matrix can result from damage incurred by vascular interventions, such as saphenous vein bypass grafting and angioplasty. Fibrillar collagens are an important component of the thrombogenic extracellular matrix. Herein we describe a means of targeting poly(ethylene glycol) (PEG)-mediated blockade directly to platelet-binding ECM molecules, such as type I collagen, thereby selectively blocking platelet adhesion to vascular matrix. Purified fibronectin (FN), a matrix protein that interacts with fibrillar collagens and platelets, was selectively pegylated to generate a targeted molecular shielding reagent that masked ECM ligands from platelet recognition and adhesion. This approach protects the functions of other vascular proteins, including surface proteins on intact endothelium. To mask the platelet-binding site of FN, PEG-propyl moieties (5000 Da) were covalently appended to lysine residues on the surface of FN, generating FNPEG-5K. To preserve the collagen-binding function of FN, it was pegylated while bound to a gelatin agarose matrix. We demonstrate that FNPEG-5K blocks platelet adhesion to purified type I collagen. Moreover, the same preparation blocks platelet adhesion to vascular wall components, including collagens.     

Intermittent hypoxia modulates nitric oxide-dependent vasodilation and capillary perfusion during ischemia-reperfusion-induced damage

The microvascular function of nitric oxide (NO) during ischemia-reperfusion (I/R) in intermittent hypoxia (IH)-pretreated hamsters was analyzed using 20 mg/kg of the nonselective NO inhibitor N(omega)-nitro-l-arginine methyl ester (l-NAME) and 5 mg/kg of the preferential inducible NO inhibitor S-methylisothiourea sulphate (SMT) injected before I/R. Studies were made in the hamster cheek pouch microcirculation (intravital fluorescence microscopy). IH consisted of 6 min of 8% O(2) breathing followed by 6 min of 21% O(2) for every 8 h for 21 days. Normoxia controls (NCs) were exposed to room air for the same period. The effects were characterized in terms of systemic hemodynamics, diameter, flow, wall shear stress in arterioles, capillary perfusion, and the concentrations of thiobarbituric acid-reactive substances (TBARS) and plasma NO, assessed as nitrite/nitrate (NOx) levels. IH did not change arterial blood pressure and increased hematocrit and shear stress. IH increased NOx and TBARS levels and reduced arterial diameter, blood flow, and capillary perfusion versus the NC. Conversely, TBARS and NOx were lower during I/R in IH-pretreated hamsters, resulting in vasodilation and the increase of capillary perfusion and shear stress. After IH, capillary perfusion was reduced by 24% (2.3%) and enhanced by 115% (1.7%) after I/R (P < 0.05). Both modalities of NO blockade decreased NOx generation and increased TBARS versus IH. l-NAME and SMT induced a significant decrease in arteriolar diameter, blood flow, and capillary perfusion (P < 0.05). l-NAME enhanced TBARS more than SMT and aggravated I/R damage. In conclusion, we demonstrated that preconditioning with IH greatly reduces oxidative stress and stimulates NO-induced vasodilation during I/R injury, thus maintaining capillary perfusion.     

Early recovery of microvascular perfusion induced by t-PA in combination with abciximab or eptifibatide during postischemic reperfusion

Background  

GPIIb/IIIa inhibitors abciximab and eptifibatide have been shown to inhibit platelet aggregation in ischemic heart disease. Our aim was to test the efficacy of abiciximab (Reo Pro) or eptifibatide (Integrilin) alone or in combination with plasminogen activator (t-PA) in an experimental model of ischemia reperfusion (I/R) in hamster cheek pouch microcirculation visualized by fluorescence microscopy. Hamsters were treated with saline, or abiciximab or eptifibatide or these drugs combined with t-PA infused intravenously 10 minutes before ischemia and through reperfusion. We measured the microvessel diameter changes, the arteriolar red blood cell (RBC) velocity, the increase in permeability, the perfused capillary length (PCL), and the platelet and leukocyte adhesion on microvessels.     

Gender specific effect of progesterone on myocardial ischemia/reperfusion injury in rats

The study was designed to investigate the effect of progesterone and its gender based variation on myocardial ischemia/reperfusion (I/R) injury in rats. Adult Sprague Dawley rats were divided into vehicle treated reperfusion injury group male (I/R-M), female (I/R-F), ovariectomised (I/R-OVR) and progesterone treatment (I/R-M+PG, I/R-F+PG, I/R-OVR+PG) groups, respectively. I/R injury was produced by occluding the left descending coronary artery (LCA) for 1 h and followed by re-opening for 1 h. Progesterone (2 mg kg(-1) i.p.) was administered 30 min after induction of ischemia. Hemodynamic parameters (+/-dp/dt, MAP), heart rate, ST-segment elevation and occurrence of ventricular tachycardia (VT) were measured during the I/R period. The myocardial infarct area, oxidative stress markers, activities of myeloperoxidase (MPO) and creatine kinase (CK) were determined after the experiment along with the assessment of the effect on apoptotic activity by using DNA fragmentation analysis. Histological observations were carried out on heart tissue. Treatment with progesterone significantly (P<0.05) reduced infarct area, lipid peroxidation (LPO) level and activity of MPO in females (I/R-F+PG) as compared to ischemic females (I/R-F). Progesterone significantly (P<0.001, P<0.05) inhibited serum CK activity and incidences of VT in female rats. Superoxide dismutase (SOD) activity, reduced glutathione (GSH) levels were significantly elevated (P<0.05) in I/R-F+PG group. Internucleosomal DNA fragmentation was less in I/R-F+PG group when compared to I/R-F group. The ischemic male and ovariectomised (I/R-M and I/R-OVR) counterparts did not show any significant change after progesterone treatment. In conclusion, the cardioprotective effect of progesterone on myocardial I/R injury induced damage is based on gender of the animal. The protective effect could be mediated by attenuation of inflammation and its possible interaction with endogenous estrogen.     

Impact of myocardial structure and function postinfarction on diastolic strain measurements: implications for assessment of myocardial viability

Venular control of arteriolar perfusion has been the focus of several investigations in recent years. This study investigated 1) whether endogenous adenosine helps control venule-dependent arteriolar dilation and 2) whether venular leukocyte adherence limits this response via an oxidant-dependent mechanism in which nitric oxide (NO) levels are decreased. Intravital microscopy was used to assess changes in arteriolar diameters and NO levels in rat mesentery. The average resting diameter of arterioles (27.5 +/- 1.0 microm) paired with venules with minimal leukocyte adherence (2.1 +/- 0.3 per 100-microm length) was significantly larger than that of unpaired arterioles (24.5 +/- 0.8 microm) and arterioles (23.3 +/- 1.3 microm) paired with venules with higher leukocyte adherence (9.0 +/- 0.5 per 100-microm length). Local superfusion of adenosine deaminase (ADA) induced significant decreases in diameter and perivascular NO concentration in arterioles closely paired to venules with minimal leukocyte adherence. However, ADA had little effect on arterioles closely paired to venules with high leukocyte adherence or on unpaired arterioles. To determine whether the attenuated response to ADA for the high-adherence group was oxidant dependent, the responses were also observed in arterioles treated with 10(-4) M Tempol. In the high-adherence group, Tempol fully restored NO levels to those of the low-adherence group; however, the ADA-induced constriction remained attenuated, suggesting a possible role for an oxidant-independent vasoconstrictor released from the inflamed venules. These findings suggest that adenosine- and venule-dependent dilation of paired arterioles may be mediated, in part, by NO and inhibited by venular leukocyte adherence.     

Leukocyte and endothelial cell adhesion molecules in a chronic murine model of myocardial reperfusion injury

Expression of endothelial and leukocyte cell adhesion molecules is a principal determinant of polymorphonuclear neutrophil (PMN) recruitment during inflammation. It has been demonstrated that pharmacological inhibition of these molecules can attenuate PMN influx and subsequent tissue injury. We determined the temporal expression of alpha-granule membrane protein-40 (P-selectin), endothelial leukocyte adhesion molecule 1 (E-selectin), and intercellular cell adhesion molecule 1 (ICAM-1) after coronary artery occlusion and up to 3 days of reperfusion. The expression of all of these cell adhesion molecules peaked around 24 h of reperfusion. We determined the extent to which these molecules contribute to PMN infiltration by utilizing mice deficient (-/-) in P-selectin, E-selectin, ICAM-1, and CD18. Each group underwent 30 min of in vivo, regional, left anterior descending (LAD) coronary artery ischemia and 24 h of reperfusion. PMN accumulation in the ischemic-reperfused (I/R) zone was assessed using histological techniques. Deficiencies of P-selectin, E-selectin, ICAM-1, or CD18 resulted in significant (P < 0.05) attenuation of PMN infiltration into the I/R myocardium (MI/R). In addition, P-selectin, E-selectin, ICAM-1, and CD18 -/- mice exhibited significantly (P < 0.05) smaller areas of necrosis after MI/R compared with wild-type mice. These data demonstrate that MI/R induces coronary vascular expression of P-selectin, E-selectin, and ICAM-1 in mice. Furthermore, genetic deficiency of P-selectin, E-selectin, ICAM-1, or CD18 attenuates PMN sequestration and myocardial injury after in vivo MI/R. We conclude that P-selectin, E-selectin, ICAM-1, and CD18 are involved in the pathogenesis of MI/R injury in mice.     

Ischemia/reperfusion induces the recruitment of leukocytes from whole blood under flow conditions

Ischemia/reperfusion (I/R) occurs in a number of pathological conditions, including myocardial infarction, stroke, and organ transplantation. During the reperfusion phase, leukocytes are recruited into affected tissues, where they can cause tissue damage and organ failure. Various in vitro models have been developed to study the role of adhesion molecules in I/R-mediated leukocyte recruitment. These models traditionally use isolated leukocytes and static conditions and, therefore, may not recapitulate the in vivo situation. We developed two novel in vitro models of I/R-mediated leukocyte recruitment in which leukocyte recruitment was examined using whole blood under shear conditions. Chemical treatments were used to mimic I/R in the first model, while sequential exposure to hypoxia/reoxygenation (H/R) was used to mimic I/R in the second model. We found that leukocytes were recruited from whole blood under shear conditions to endothelial cells treated with chemically induced I/R or H/R. In both models, mRNA for intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1), and E-selectin was upregulated. The role of adhesion molecules in leukocyte recruitment differed slightly between the two models, with E-selectin and VCAM-1 playing approximately equal roles in leukocyte recruitment in the chemically induced I/R model and VCAM-1 being a central mediator of leukocyte recruitment in the H/R model.     

Oxygen radical-dependent expression of CXC chemokines regulate ischemia/reperfusion-induced leukocyte adhesion in the mouse colon

Activation and accumulation of leukocytes constitute a rate-limiting step in ischemia/reperfusion (I/R)-induced tissue injury. The signalling mechanisms, however, that regulate leukocyte rolling and adhesion in the colonic microcirculation are not known. The objective of the study was to define the role of CXC chemokines (MIP-2 and KC) in I/R-induced leukocyte-endothelial cell interactions in the mouse colon. In C57/B16 mice, colonic ischemia was induced by clamping the superior mesenteric artery for 30 min and leukocyte rolling and stationary adhesion were examined in venules after 120 and 240 min of reperfusion. I/R provoked a clear-cut increase in leukocyte rolling and adhesion in colonic venules. Both MIP-2 and KC were upregulated at the gene and protein level in the reperfused colon. Immunoneutralization of MIP-2 and KC by monoclonal antibodies reduced reperfusion-induced firm adhesion of leukocytes by 73% and 75%, respectively. Interestingly, combined inhibition of MIP-2 and KC additionally decreased leukocyte rolling by 79%, but did not further reduce the number of firmly adherent leukocytes. To study the role of oxygen free radicals (OFRs) in the regulation of CXC chemokine expression, additional animals were pretreated with the xanthine-oxidase inhibitor allopurinol. In fact, allopurinol treatment reduced the colonic levels of MIP-2 and KC by 62% and 64%, respectively. This study elucidates important interactions between OFRs and chemokines in the I/R-induced leukocyte response in the mouse colon. Moreover, our data demonstrate that CXC chemokines play a fundamental role in colonic I/R and that functional interference with CXC chemokines may protect against pathological inflammation in the colon.     

Endothelin receptor blockers reduce I/R-induced intestinal mucosal injury: role of blood flow

The aim of the present study was to assess the role of endothelin (ET) in ischemia-reperfusion (I/R)-induced mucosal injury. Mucosal permeability ((51)Cr-EDTA clearance) and tissue myeloperoxidase (MPO) activity were significantly increased after 30 min of ischemia followed by 30 min of reperfusion. The I/R-induced increases in mucosal permeability and polymorphonuclear leukocyte (PMN) infiltration were significantly attenuated by pretreatments with ET(A) (BQ-485) and/or ET(B) (BQ-788) receptor antagonists. Monoclonal antibody (MAb) directed against intercellular adhesion molecule-1 (ICAM-1; MAb 1A29) and superoxide dismutase (SOD) pretreatments significantly attenuated the increased mucosal permeability and PMN infiltration in a similar manner as with ET receptor antagonists. Superior mesenteric artery blood flow was significantly reduced during the reperfusion period. Both ET receptor antagonists caused a significant rise in blood flow compared with an untreated I/R group. In conclusion, our data suggest that ET(A) and/or ET(B) receptors, ICAM-1, and superoxide play an important role in I/R-induced mucosal dysfunction and PMN infiltration. Furthermore, ET is involved in the pathogenesis of post-reperfusion-induced damage and beneficial effects of ET receptor antagonism are related to an improvement of disturbed blood flow during the reperfusion period.     

Oxidative stress and inflammatory response evoked by transient cerebral ischemia/reperfusion: effects of the PPAR-alpha agonist WY14643

This study investigated the effects of the selective peroxisome proliferator-activated receptor-alpha (PPAR-alpha) agonist WY14643 on ischemia/reperfusion (I/R) injury in the rat hippocampus. Transient cerebral ischemia (30 min), followed by 1-24 h reperfusion, significantly increased the generation of reactive oxygen species, nitric oxide (NO), and lipid peroxidation end-products, as well as markedly reducing levels of the endogenous antioxidant glutathione. Reperfusion for 3-6 h led to increased expression of the proteins heme oxygenase-1 (HO-1), cyclooxygenase-2 (COX-2), inducible NO synthase (iNOS), and intercellular adhesion molecule-1 (ICAM-1). Pretreatment with WY14643 suppressed oxidative stress and expression of HO-1, iNOS, and ICAM-1, but had no effect on COX-2. These effects are due to suppression of the activation of p38 mitogen-activated protein kinase and nuclear factor-kappaB. The PPAR-alpha antagonist MK886 abolished the beneficial effects of WY14643. The levels of S100B protein, a marker of cerebral injury used in stroke trials to monitor injury, were high in the hippocampus of rats exposed to I/R, but markedly reduced by WY14643. We propose that WY14643 protects the brain against excessive oxidative stress and inflammation and may thus be useful in treating stroke.     

Nuclear factor-kappaB inhibition by pyrrolidinedithiocarbamate attenuates gastric ischemia-reperfusion injury in rats

Pyrrolidinedithiocarbamate (PDTC) is a potent antioxidant and an inhibitor of nuclear factor-kappaB (NF-kappaB). The present study examined the impact of PDTC preconditioning on gastric protection in response to ischemia-reperfusion (I/R) injury to the rat stomach. Male Wistar rats were recruited and divided into 3 groups (n = 7). One group was subjected to gastric ischemia for 30 min and reperfusion for 1 hour. The second group of rats was preconditioned with PDTC (200 mg/kg body mass i.v.) 15 min prior to ischemia and before reperfusion. The third group of rats was sham-operated and served as the control group. Gastric I/R injury increased serum lactate dehydrogenase level, vascular permeability of gastric mucosa (as indicated by Evans blue dye extravasation) and gastric content of inflammatory cytokine; tumor necrosis factor-alpha (TNF-alpha). Moreover, oxidative stress was increased as indicated by elevated lipid peroxides formation (measured as thiobarbituric acid reactive substances) and depleted reduced glutathione in gastric tissues. NF-kappaB translocation was also detected by electrophoretic mobility shift assay. Microscopically, gastric tissues subjected to I/R injury showed ulceration, hemorrhages, and neutrophil infiltration. Immunohistochemical studies of gastric sections revealed increased expression of p53 and Bcl-2 proteins. PDTC pretreatment reduced Evans blue extravasation, serum lactate dehydrogenase levels, gastric TNF-alpha levels, and thiobarbituric acid reactive substances content, and increased gastric glutathione content. Moreover, PDTC pretreatment abolished p53 expression and inhibited NF-kappaB translocation. Finally, histopathological changes were nearly restored by PDTC pretreatment. These results clearly demonstrate that NF-kappaB activation and pro-apoptotic protein p53 induction are involved in gastric I/R injury. PDTC protects against gastric I/R injury by an antioxidant, NF-kappaB inhibition, and by reduction of pro-apoptotic protein p53 expression, which seems to be downstream to NF-kappaB, thus promoting cell survival.