cyclic-gmp and Reperfusion-Injury

Ischemia/reperfusion (I/R) occurs in renal artery stenosis, partial nephrectomy and most commonly during kidney transplantation. It brings serious consequences such as DGF (Delayed Graft Function) or organ dysfunction leading to renal failure and ultimate death. There is no effective therapy to handle the consequences of Renal Ischemia/Reperfusion (I/R) injury. Cyclic nucleotides, cAMP and cGMP are the important second messengers that stimulate intracellular signal transduction for cell survival in response to growth factors and peptide hormones in normal tissues and in kidneys plays significant role that involves vascular tone regulation, inflammation and proliferation of parenchymal cells. Renal ischemia and subsequent reperfusion injury stimulate signal transduction pathways involved in oxidative stress, inflammation, alteration in renal blood flow leading to necrosis and apoptosis of renal cell.. An extensive literature review of various search engines like PubMed, Medline, Bentham, Scopus, and EMBASE (Elsevier) databases was carried out. To understand the functioning of Phosphodiesterases (PDEs) and its pharmacological modulation in Renal Ischemia-Reperfusion Injury.. Current therapeutic options may not be enough to treat renal I/R injury in group of patients and therefore, the current review has discussed the general characteristics and physiology of PDEs and preclinical-studies defining the relationship between PDEs expression in renal injury due to I/R and its outcome on renal function.. The role of PDE inhibitors in renal I/R injury and the clinical status of drugs for various renal diseases have been summarized in this review.

Topics: Cyclic AMP; Cyclic GMP; Humans; Kidney; Kidney Diseases; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Reperfusion Injury; Signal Transduction

Limiting the injurious effects of myocardial ischemia-reperfusion is a desirable therapeutic target, which has been investigated extensively over the last three decades. Here we provide an up to date review of the literature documenting the experimental and clinical research demonstrating the effects of manipulating cGMP for the therapeutic targeting of the injurious effects of ischemic heart disease. Augmentation of the cyclic nucleotide cGMP plays a crucial role in many cardioprotective signaling pathways. There is an extensive body of literature which supports pharmacological targeting of cGMP or upstream activators in models of ischemia-reperfusion to limit injury. NO donors have long been utilised to manipulate cGMP, and more recently non-NO synthase derived NOx species have been investigated, resulting in their evaluation in clinical trials for the treatment of ischemic heart disease. Encouraging results demonstrate that natriuretic peptides are worthy candidates in manipulating cGMP and its downstream effectors to afford cytoprotection. Synthetic ligands have been designed which co-activate natriuretic peptide receptors to improve targeting this pathway. Advances have been made in targeting the soluble guanylyl cyclase which catalyzes the production of cGMP independently of the endogenous ligand NO using NO-independent stimulators and activators of sGC. These novel compounds show promise as a new class of drugs that target this signaling cascade specifically under pathological conditions when endogenous NO production may be compromised. Regulating the degradation of cGMP via phosphodiesterase inhibition also shows therapeutic potential. It is clear that production and regulation of cGMP is complex, indeed its spatial production and cellular distribution are only just emerging.

Topics: Cyclic GMP; Guanylate Cyclase; Heme Oxygenase (Decyclizing); Humans; Natriuretic Peptides; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitrites; Phosphodiesterase Inhibitors; Receptors, Adrenergic, beta-3; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Signal Transduction; Soluble Guanylyl Cyclase

Natriuretic peptides elicit vasodilation, increased sodium excretion and concomitant diuresis, and counteract the RAAS. In the heart itself, natriuretic peptides may also act anti-inflammatory and antifibrotic. This has led to the pursuit of natriuretic peptides and chemically modified peptides as adjunctive therapy in myocardial ischaemia. However, natriuretic peptide infusion may also influence the endogenous natriuretic peptide response and lipid accumulation. We hypothesised that a) natriuretic peptide infusion (BNP and CD-NP) is cardiomyocyte protective, b) affects the endogenous response, and c) facilitate cardiac lipid accumulation. We examined these effects in a minimally invasive porcine model of regional cardiac ischaemia and reperfusion. The studies were supplemented by a 48-hour porcine model of ischemia and reperfusion as well as an in vitro study of BNP administered in a HL-1 cell model of "ischaemia/reperfusion". Infarct size was determined by TTC staining, plasma troponin T release, and total RNA integrity in cardiac tissue samples. The endogenous response was assessed by a processing-independent proANP immunoassay and mRNA quantitation. Lipids in plasma and myocardial tissue were determined by TLC. The studies show that natriuretic peptides decrease cardiomyocyte damage, possibly partly through indirect mechanisms. Furthermore, BNP infusion completely inverts the endogenous response, whereas CD-NP infusion does not. Finally, both natriuretic peptides increase plasma free fatty acids, which is associated with an increased cardiac lipid accumulation in non-ischaemic myocardium. In conclusion, the studies suggest that natriuretic peptides are beneficial in terms of reduced cardiac injury. In addition, the endogenous natriuretic peptide response is inverted. The results advocate for pursuing natriuretic peptide treatment in ischaemia/reperfusion damage. However, the metabolic consequences in a cardiac tissue challenged by ischaemia should be pursued before testing the peptides in patients.

Topics: Animals; Blood Pressure; Caspase 1; Cholesterol; Cyclic GMP; Diuresis; Fatty Acids, Nonesterified; Female; Glycerol; Mice; Myocardial Infarction; Myocardial Ischemia; Myocytes, Cardiac; Natriuretic Peptide, Brain; Natriuretic Peptide, C-Type; Natriuretic Peptides; Reperfusion Injury; RNA, Messenger; Swine; Triglycerides; Troponin T; Tumor Cells, Cultured; Vascular Endothelial Growth Factor A

Topics: Animals; Apoptosis; Biological Availability; Cyclic GMP; Guanylate Cyclase; Humans; Nitric Oxide; Reperfusion Injury; Thrombosis

Topics: Adrenomedullin; Animals; Apoptosis; Cyclic GMP; DNA-Binding Proteins; Endothelial Cells; Gene Expression Regulation; Humans; Hypoxia-Inducible Factor 1; Hypoxia-Inducible Factor 1, alpha Subunit; Neovascularization, Physiologic; Nitric Oxide; Nuclear Proteins; Peptides; Phosphatidylinositol 3-Kinases; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Reperfusion Injury; RNA, Messenger; Transcription Factors; Vasodilation

Current methods of lung preservation allow for effective, expeditious transplantation as a treatment for end-stage pulmonary disease. However, the utilization of hypothermia, hyperkalemia, and pulmonary artery distension as a single rapid flush for perfusion is less than ideal. All these interventions result in increased pulmonary vascular resistance and suboptimal preservation of lung function. The ability to preserve lungs for longer time intervals and with less risk of tissue injury would provide significant advantages. There would be a greater likelihood that rare size or blood types could find matches by enlarging the area of organ distribution. Optimal preservation would also improve the perioperative outcomes in regard to primary graft failure and subsequently reduce the later complication of chronic rejection and graft lung dysfunction. Finally, through a better understanding of the mechanisms of lung injury during preservation and by developing means to limit the injury, it would be possible to utilize organs from donors that at this time would not be considered optimal. This would increase the donor pool without compromising the recipient's outcome.

Topics: Animals; Antioxidants; Cold Temperature; Cyclic AMP; Cyclic GMP; Free Radical Scavengers; History, 20th Century; Humans; Lung; Lung Injury; Lung Transplantation; Organ Preservation; Platelet Activating Factor; Reperfusion Injury; Respiration, Artificial; Solutions; Vasodilator Agents

After graft reperfusion in orthotopic liver transplantation (OLT), ischemia reperfusion syndrome (IRS) is characterized by persistent hypotension with a low systemic vascular resistance. Methylene blue (MB) has been used as a vasopressor in sepsis and acute liver failure. We investigated the effect of MB on IRS during OLT. Thirty-six patients undergoing elective OLT were randomized to receive either a bolus of MB 1.5 mg/kg before graft reperfusion, or normal saline (placebo). We recorded hemodynamic variables, postoperative liver function tests, and time to hospital discharge. Blood samples were analyzed for arterial lactate concentration, cyclic 3',5'-monophosphate, and plasma nitrite/nitrate concentrations. The MB group had higher mean arterial pressure (P = 0.035), higher cardiac index (P = 0.04), and less epinephrine requirement (P = 0.02). There was no difference in systemic vascular resistance or central venous pressure. Serum lactate levels were lower at 1 h after reperfusion in MB patients, suggesting better tissue perfusion (P = 0.03). In the presence of MB, there was a reduction in cyclic 3',5'-monophosphate (P < 0.001), but not plasma nitrites. Postoperative liver function tests and time to hospital discharge were the same in both groups. MB attenuated the hemodynamic changes of IRS in OLT acting via guanylate cyclase inhibition.. Methylene blue attenuates the hemodynamic changes of the ischemia reperfusion syndrome in liver transplantation, and this effect involves guanylate cyclase inhibition.

Topics: Blood Pressure; Cardiac Output; Central Venous Pressure; Cyclic GMP; Enzyme Inhibitors; Female; Guanylate Cyclase; Hemodynamics; Humans; Lactic Acid; Liver Cirrhosis; Liver Transplantation; Male; Methylene Blue; Middle Aged; Nitrites; Reperfusion; Reperfusion Injury; Vascular Resistance; Vasoconstrictor Agents

Ischemic stroke is one of the leading causes of mortality worldwide. The available treatments are not effective. Phosphodiesterase 9A (PDE9A) is an intracellular cyclic guanosine monophosphate (cGMP) hydrolase considered to be a promising therapeutic target for brain diseases. This study explored neuroprotective effects and the underlying mechanism of LW33, a novel PDE9A inhibitor, on ischemic stroke in vitro and in vivo.. A middle cerebral artery occlusion (MCAO) model was established in adult male Sprague-Dawley rats and an oxygen-glucose deprivation/reoxygenation (OGD/R) model was established in human SH-SY5Y cells to mimic ischemia-reperfusion injury in vitro.. LW33 increased cell viability, reduced lactate dehydrogenase activity, and OGD/R-induced apoptosis of SH-SY5Y cells. The protective effects of LW33 against stroke occurred in the recovery phase. LW33 administration significantly reduced cerebral infarction volume in MCAO rats, without causing significant deformation or necrosis of neurons in the cortex. LW33 also improved learning and cognitive dysfunction and reduced other pathological changes in MCAO rats in the recovery period. Moreover, LW33 stimulated the cGMP/PKG/CREB pathway and up-regulated the expression of the apoptosis-related proteins, and this effect was reversed by KT5823 treatment.. LW33 inhibited cell apoptosis and promoted neuronal repair to alleviate OGD/R and MCAO induced pathological alterations via the cGMP/PKG/CREB pathway, indicating that LW33 may be a promising therapeutic target for ischemic stroke.

Topics: Animals; Apoptosis; Apoptosis Regulatory Proteins; Brain Ischemia; Cyclic GMP; Glucose; Infarction, Middle Cerebral Artery; Ischemic Stroke; Male; Neuroprotective Agents; Oxygen; Phosphodiesterase Inhibitors; Phosphoric Diester Hydrolases; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction

Ischaemia-reperfusion injury impairs the nitric oxide/soluble guanylate cyclase/cyclic guanosine monophosphate (cGMP) signalling pathway and leads to vascular dysfunction. We assessed the hypothesis that the soluble guanylate cyclase activator cinaciguat would protect the vascular graft against ischaemia-reperfusion injury.. In the treatment groups, rats (n = 8/group) were pretreated with either intravenous saline or intravenous cinaciguat (10 mg/kg) 2 h before an aortic transplant. Aortic grafts were stored for 2 h in saline and transplanted into the abdominal aorta of the recipients. Two hours after the transplant, the grafts were harvested and mounted in an organ bath. Vascular function of the grafts was investigated in the organ bath. Terminal deoxynucleotidyl transferase dUTP nick end labelling, cluster of differentiation 31, caspase-3, endothelial nitric oxide synthase, cGMP, nitrotyrosine and vascular cell adhesion molecule 1 immunochemical reactions were also investigated.. Pretreatment with cinaciguat significantly improved endothelium-dependent maximal relaxation 2 h after reperfusion compared with the saline group (maximal relaxation control: 96.5 ± 1%, saline: 40.4 ± 3% vs cinaciguat: 54.7 ± 2%; P < 0.05). Pretreatment with cinaciguat significantly reduced DNA fragmentation and nitro-oxidative stress; decreased the caspase-3 and vascular cell adhesion molecule 1 scores; and increased endothelial nitric oxide synthase, cGMP and cluster of differentiation 31 scores.. Our results demonstrated that enhancement of cGMP signalling by pharmacological activation of the soluble guanylate cyclase activator cinaciguat might represent a beneficial therapy for treating endothelial dysfunction of arterial bypass graft during cardiac surgery.

Topics: Animals; Cyclic GMP; Endothelium, Vascular; Nitric Oxide; Rats; Reperfusion Injury; Soluble Guanylyl Cyclase; Vascular Grafting

The nitric oxide/cyclic guanosine monophosphate (NO/cGMP) pathway has a significative influence in hemodynamic changes that occur in transplants. Classically, the ischemia-reperfusion syndrome (IRS) is characterized by hypotension and low vascular resistance, when cGMP and nitric oxide (NO) are increased, contributing to oxidative stress, within an inflammatory context. These mechanisms occur in several types of transplants, such as liver, heart, lungs, kidney, which are a therapeutic choice in several clinical conditions when conventional treatments failed. It is well known the significant relation between graft dysfunction or rejection and ischemia-reperfusion injury that is linked to inflammatory response and NO/cGMP pathway activation. This review aims to study the NO/cGMP pathway in solid organ transplants. Finally, we inquire whether physicians do not underestimate the NO/cGMP pathway.

Topics: Cyclic GMP; Hemodynamics; Humans; Hypotension; Liver; Lung; Methylene Blue; Nitric Oxide; Reperfusion; Reperfusion Injury; Signal Transduction; Transplants; Vascular Resistance

Nicorandil is a hybrid between nitrates and K. Nicorandil attenuated IR injury in isolated rat lungs. This protective effect appears to involve its activation as K

Topics: Animals; Capillary Permeability; Cyclic GMP; KATP Channels; Lung; Lung Injury; Male; Membrane Transport Modulators; Nicorandil; Perfusion; Pulmonary Circulation; Pulmonary Edema; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Soluble Guanylyl Cyclase; Vascular Resistance

Prohibitin (PHB) is a critical protein involved in many cellular activities. In brain, PHB resides in mitochondria, where it forms a large protein complex with PHB2 in the inner TFmembrane, which serves as a scaffolding platform for proteins involved in mitochondrial structural and functional integrity. PHB overexpression at moderate levels provides neuroprotection in experimental brain injury models. In addition, PHB expression is involved in ischemic preconditioning, as its expression is enhanced in preconditioning paradigms. However, the mechanisms of PHB functional regulation are still unknown. Observations that nitric oxide (NO) plays a key role in ischemia preconditioning compelled us to postulate that the neuroprotective effect of PHB could be regulated by NO. Here, we test this hypothesis in a neuronal model of ischemia-reperfusion injury and show that NO and PHB are mutually required for neuronal resilience against oxygen and glucose deprivation stress. Further, we demonstrate that NO post-translationally modifies PHB through protein S-nitrosylation and regulates PHB neuroprotective function, in a nitric oxide synthase-dependent manner. These results uncover the mechanisms of a previously unrecognized form of molecular regulation of PHB that underlies its neuroprotective function.

Topics: Animals; Cell Death; Cells, Cultured; Cyclic GMP; Enzyme Inhibitors; Mice; Neurons; Neuroprotection; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Prohibitins; Reperfusion Injury; Repressor Proteins; Signal Transduction

Carbon monoxide (CO) releasing molecule (CORM)-3, a water-soluble CORM, has protective effects against inflammatory and ischemia/reperfusion injury. We determined the effect of CORM-3 against neuronal pyroptosis in a model of hemorrhagic shock and resuscitation (HSR) in rats via mitochondrial regulation.. Rats were treated with CORM-3 (4 mg/kg) in vitro after HSR. We measured cortical CO content 3-24 h after HSR; assessed neuronal pyroptosis, mitochondrial morphology, ROS production, and mitochondrial membrane potential at 12 h after HSR; and evaluated brain magnetic resonance imaging at 24 h after HSR and learning ability 30 days after HSR. We also measured soluble guanylate-cyclase (sGC)-cyclic guanosine monophosphate (cGMP) signaling pathway activity using a blocker of sGC, NS2028, and. Among rats that underwent HSR, CORM-3-treated rats had more CO in the cortical tissue than sham- and iCORM-3-treated rats. CORM-3-treated rats had significantly less neuronal pyroptosis in the cortical tissue; higher sGC activity and cGMP content; lower ROS production; better mitochondrial morphology, function, and membrane potential; and enhanced learning/memory ability than HSR-treated rats. However, these neuroprotective effects of CORM-3 were partially inhibited by NS2028.. CORM-3 may alleviate neuronal pyroptosis and improve neurological recovery in HSR through mitochondrial regulation mediated by the sGC-cGMP pathway. Thus, CO administration could be a promising therapeutic strategy for hemorrhagic shock.

Topics: Animals; Carbon Monoxide; Cyclic GMP; Disease Models, Animal; Humans; Mitochondria; Neurons; Neuroprotective Agents; Organometallic Compounds; Pyroptosis; Rats; Reperfusion Injury; Resuscitation; Shock, Hemorrhagic; Signal Transduction; Soluble Guanylyl Cyclase

Ischaemia reperfusion (IR) injury occurs during vascular graft harvesting and implantation during vascular/cardiac surgery. Elevated intracellular cyclic guanosine monophosphate (cGMP) levels contribute to an effective endothelial protection in different pathophysiological conditions. The hypothesis that the phosphodiesterase-5 inhibitor vardenafil would protect vascular grafts against IR injury by upregulating the nitric oxide-cGMP pathway in the vessel wall of the bypass graft was investigated.. Lewis rats (n = 6-7/group) were divided into Group 1, control; Group 2, donor rats received intravenous saline; Group 3, received intravenous vardenafil (30 μg/kg) 2 h before explantation. Whereas aortic arches of Group 1 were immediately mounted in an organ bath, aortic segments of Groups 2 and 3 were stored for 2 h in saline and transplanted into the abdominal aorta of the recipient. Two hours after transplantation, the implanted grafts were harvested. Endothelium dependent and independent vasorelaxations were investigated. TUNEL, CD-31, ICAM-1, VCAM-1, α-SMA, nitrotyrosine, dihydroethidium and cGMP immunochemistry were also performed.. The results support the view that impairment of intracellular cGMP signalling plays a role in the pathogenesis of the endothelial dysfunction of an arterial graft after bypass surgery, which can effectively be prevented by vardenafil. Its clinical use as preconditioning drug could be a novel approach in vascular/cardiac surgery.

Topics: Actins; Animals; Aorta, Thoracic; Cold Ischemia; Cyclic GMP; Cytoprotection; DNA Damage; Intercellular Adhesion Molecule-1; Male; Nitrosative Stress; Phosphodiesterase 5 Inhibitors; Platelet Endothelial Cell Adhesion Molecule-1; Rats, Inbred Lew; Reperfusion Injury; Signal Transduction; Tissue and Organ Harvesting; Tyrosine; Vardenafil Dihydrochloride; Vascular Cell Adhesion Molecule-1; Vascular System Injuries; Vasodilator Agents; Warm Ischemia

The current study is focusing on the role of brain natriuretic peptide (BNP), a substrate of dipeptidyl peptidase-4 (DPP-4) enzyme, and its signaling survival pathway in the cardioprotective mechanism of sitagliptin, a DPP-4 inhibitor.. Pretreatment with sitagliptin provided significant protection against I/R injury as manifested by decreasing, percentage of infarct size, suppressing the elevated ST segment, reducing the increased cardiac enzymes, as well as DPP-4 activity and elevating both heart rate (HR) and left ventricular developed pressure (LVDP). However, the addition of either blocker to sitagliptin regimen reversed partly its cardioprotective effects. Although I/R increased BNP content, it unexpectedly decreased that of cGMP; nevertheless, sitagliptin elevated both parameters, an effect that was not affected by the use of the two blockers. On the molecular level, sitagliptin decreased caspase-3 activity and downregulated the mRNA levels of BNP, Bax, and Cyp D, while upregulated that of Bcl2. The use of either KT-5823 or 5-HD with sitagliptin hindered its effect on the molecular markers tested.. The results of the present study suggest that the cardioprotective effect of sitagliptin is mediated partly, but not solely, through the BNP/cGMP/PKG survival signaling pathway.

Topics: Animals; bcl-2-Associated X Protein; Carbazoles; Cardiotonic Agents; Caspase 3; Cyclic GMP; Cyclophilins; Decanoic Acids; Dipeptidyl Peptidase 4; Hemodynamics; Hydroxy Acids; Male; Natriuretic Peptide, Brain; Peptidyl-Prolyl Isomerase F; Proto-Oncogene Proteins c-bcl-2; Rats; Reperfusion Injury; Signal Transduction; Sitagliptin Phosphate

Total knee arthroplasty (TKA) is the most common and cost-effective treatment for older adults with long-standing osteoarthritis. During TKA, muscle cells suffer from prolonged oxygen deficiency, which leads to altered cell metabolism that reduces the energy demand and maintains cell homeostasis before blood flow is restored. This study focused on the role of the lncRNA muscleblind-like 1 antisense RNA 1 (MBNL1-AS1) in protecting sevoflurane-pretreated mice against ischemia-reperfusion (I/R) injury after TKA, as well as the elucidation of the potential associated mechanism. Identification of differentially expressed lncRNAs was performed using the microarray dataset GSE21164, which was extracted from the GEO database. Target genes of the lncRNA were determined using Multi-Experiment Matrix (MEM), a dual-luciferase reporter gene assay, and KEGG enrichment analyses. The results showed that MBNL1-AS1 was overexpressed in skeletal muscle cells in mice, while KCNMA1, which was enriched in the cGMP-PKG signaling pathway, was negatively regulated by MBNL1-AS1. Furthermore, I/R mice displayed serious inflammatory reactions. Down-regulation of MBNL1-AS1 increased the expression of KCNMA1, PKGII, VASP, VEGF, Bcl-2, Cyclin D1, Cyclin D3, and Cdc 42 but decreased the expression of Bax, cleaved caspase-3, and cleaved PARP. Furthermore, upon MBNL1-AS1 upregulation, the rate of cell apoptosis increased while the rate of cell proliferation decreased. Our data suggested that down-regulated lncRNA MBNL1-AS1 might promote the proliferation and inhibit the apoptosis of skeletal muscle cells by upregulating KCNMA1 expression via activation of the cGMP-PKG signaling pathway, thus protecting sevoflurane-pretreated mice against I/R injury after TKA.

Topics: Animals; Apoptosis; Arthroplasty, Replacement, Knee; Cell Cycle; Cell Proliferation; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Down-Regulation; Interleukin-1beta; L-Lactate Dehydrogenase; Large-Conductance Calcium-Activated Potassium Channel alpha Subunits; Male; Mice; Muscle, Skeletal; Reperfusion Injury; RNA, Long Noncoding; Sevoflurane; Signal Transduction; Tumor Necrosis Factor-alpha

To explore the protective effect of curcumin on renal ischemia-reperfusion injury (RIRI) in rats, and its influence on nephridial tissue's NO and cGMP levels as well as downstream signaling pathway, to elucidate the possible mechanism of curcumin on RIRI.. 36 Sprague Dawley rats (SD rats) were randomly divided into Sham group, Model group, curcumin (CUR +) Model group, 12 rats per group. They were all given RIRI model preparation by unilateral artery occlusion method. All groups' β2-MG in urine in 24h, serum Cr and BUN were compared, and UAER were calculated. Nitric oxide synthase (NOS), cGMP-dependent protein kinase (PKG), Caspase-3 expression were all determined by western blot. Nitric oxide (NO), NOS and cGMP levels were also examined by using ELISA. All groups' nephridial histomorphology and kidney tubules score were evaluated and compared.. β2-MG and UAER in urine, serum Cr and BUN, in renal tissue were all elevated in Model of RIRI, indicating the success of animal model of RIRI establishment, and above index in CUR + Model group were all lower than those in Model group. Furthermore, iNOS, NO, cGMP, PKG and Caspase-3 in renal tissue were all increased in Model of RIRI, indicating the NO signaling pathway was activated, which is one of the pathogenesis of RIRI, and above index in CUR + Model group were all lower than those in Model group, suggesting that inactivation of iNOS/NO/cGMP/PKG signaling pathway is one of the reasons that explain the protective effect of curcumin in RIRI.. The activation of iNOS/NO/cGMP/PKG signaling pathway and the consequent promoted apoptosis of renal tubules are significantly involved in the pathogenesis of development of RIRI, and curcumin treatment could protect renal tubules against RIRI, at least partially, by suppressing the activated iNOS/NO/cGMP/PKG signaling pathway.

Topics: Animals; beta 2-Microglobulin; Blood Urea Nitrogen; Caspase 3; Cell Line; Creatinine; Curcumin; Cyclic GMP; Disease Models, Animal; Humans; Kidney Tubules; Male; Nitric Oxide; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Phosphoglycerate Kinase; Protective Agents; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction

Scutellarin (SCU), a flavonoid from a traditional Chinese medicinal plant. Our previous study has demonstrated that SCU relaxes mouse aortic arteries mainly in an endothelium-depend-ent manner. In the present study, we investigated the vasoprotective effects of SCU against HR-induced endothelial dysfunction (ED) in isolated rat CA and the possible mechanisms involving cyclic guanosine monophosphate (cGMP) dependent protein kinase (PKG). The isolated endothelium-intact and endothelium-denuded rat CA rings were treated with HR injury. Evaluation of endothelium-dependent and -independent vasodilation relaxation of the CA rings were performed using wire myography and the protein expressions were assayed by Western blotting. SCU (10-1 000 μmol·L(-1)) could relax the endothelium-intact CA rings but not endothelium-denuded ones. In the intact CA rings, the PKG inhibitor, Rp-8-Br-cGMPS (PKGI-rp, 4 μmol·L(-1)), significantly blocked SCU (10-1 000 μmol·L(-1))-induced relaxation. The NO synthase (NOS) inhibitor, NO-nitro-L-arginine methylester (L-NAME, 100 μmol·L(-1)), did not significantly change the effects of SCU (10-1 000 μmol·L(-1)). HR treatment significantly impaired ACh-induced relaxation, which was reversed by pre-incubation with SCU (500 μmol·L(-1)), while HR treatment did not altered NTG-induced vasodilation. PKGI-rp (4 μmol·L(-1)) blocked the protective effects of SCU in HR-treated CA rings. Additionally, HR treatment reduced phosphorylated vasodilator-stimulated phosphoprotein (p-VASP, phosphorylated product of PKG), which was reversed by SCU pre-incubation, suggesting that SCU activated PKG phosphorylation against HR injury. SCU induces CA vasodilation in an endothelium-dependent manner to and repairs HR-induced impairment via activation of PKG signaling pathway.

Topics: Animals; Apigenin; Cell Adhesion Molecules; Cell Hypoxia; Coronary Vessels; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Glucuronates; Microfilament Proteins; NG-Nitroarginine Methyl Ester; Phosphoproteins; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Thionucleotides; Vasodilation

Phosphodiesterase-5 inhibitors prevent the breakdown of cyclic guanosine 3',5'-monophosphate (cGMP) and therefore may be useful in reducing the detrimental effects of ischemia-reperfusion (I/R) injury. The aim of this study was to assess the effects of the phosphodiesterase-5 inhibitor sildenafil, on I/R injury in a porcine model of donation after circulatory death kidney transplantation.. Kidneys were subjected to 20 min warm ischemia followed by 2 or 18 hr of cold storage (n=6 kidneys per group). After preservation kidneys were reperfused on an ex vivo perfusion system for 3 hr with an oxygenated blood based solution. Kidneys were treated with 1.4 mg/kg sildenafil infused 10 min before and for 20 min after reperfusion (n=6 kidneys per group). Renal function and injury markers were measured throughout reperfusion.. Prolonged cold ischemia (CI) significantly reduced levels of cGMP (2 hr 3.5 [1.5-5.7] vs. 18 hr 1.2 [0.3-2.8] pmol/mL; P=0.010). The administration of sildenafil significantly increased the levels (P=0.047, 0.064). Sildenafil improved the renal blood flow for the first 30 min in the 2-hr group (sildenafil, 81.8 [43.8-101.9] vs. control 40.2 [6.4-76.9] mL/min/100 g; P=0.026) and up to 60 min in the 18-hr group (sildenafil, 67.4 [38.0-87.0] vs. control 36.2 [30.5-50.0] mL/min/100 g; P=0.009) during reperfusion. Renal function was significantly impaired after 18-hr CI (P=0.0.26), and treatment with sildenafil did not improve renal function in the 2-hr (P=0.384) or 18-hr CI (P=0.099) groups.. Sildenafil had a vasodilatory action and increased levels of cGMP but did not affect recovery of renal function or protect against I/R injury.

Topics: Animals; Area Under Curve; Cold Ischemia; Creatine; Cyclic GMP; Endothelin-1; Kidney; Kidney Transplantation; Nitric Oxide; Phosphodiesterase 5 Inhibitors; Piperazines; Purines; Renal Circulation; Reperfusion Injury; Sildenafil Citrate; Sulfones; Swine; Time Factors; Warm Ischemia

Although atrial natriuretic peptide has been shown to attenuate ischemia-reperfusion (IR)-induced kidney injury, the effect of natriuretic peptide receptor (NPR)-B activation on IR-induced acute kidney injury is not well documented. The purpose of the present study was to identify the effect of C-type natriuretic peptide (CNP), a selective activator of NPR-B, on the IR-induced acute kidney injury and its mechanisms involved.. Unilaterally nephrectomized rats were insulted by IR in their remnant kidney, and they were randomly divided into three groups: sham, vehicle+IR, and CNP+IR groups. CNP (0.2μg/kg/min) was administered intravenously at the start of a 45-min renal ischemia for 2h. Rats were then killed 24h after I/R, and the blood and tissue samples were collected to assess renal function, histology, TUNEL assay, and Western blot analysis of kidney Bax and Bcl-2 expressions.. The levels of blood urea nitrogen and serum creatinine were significantly increased in rats after IR compared with vehicle-treated rats. IR elevated apoptosis, Bcl-2/Bax ratio, TUNEL positivity, oxidative stress parameters, malondialdehyde concentration, and superoxide dismutase activity. IR also induced epithelial desquamation of the proximal tubules and glomerular shrinkage. CNP significantly attenuated the IR-induced increase in BUN and serum creatinine. Furthermore, CNP restored the suppressed renal cyclic guanosine 3' 5'-monophosphate levels caused by IR insult.. Study findings suggest that CNP could ameliorate IR-induced acute kidney injury through inhibition of apoptotic and oxidative stress pathways, possibly through NPR-B-cGMP signaling.

Topics: Acute Kidney Injury; Animals; Apoptosis; bcl-2-Associated X Protein; Blood Urea Nitrogen; Blotting, Western; Creatinine; Cyclic GMP; In Situ Nick-End Labeling; Male; Malondialdehyde; Natriuretic Peptide, C-Type; Oxidative Stress; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Wistar; Receptors, Atrial Natriuretic Factor; Reperfusion Injury; Signal Transduction; Superoxide Dismutase

Carbon monoxide (CO) is an accepted cytoprotective molecule. The extent and mechanisms of protection in neuronal systems have not been well studied. We hypothesized that delivery of CO via a novel releasing molecule (CORM) would impart neuroprotection in vivo against ischemia-reperfusion injury (IRI)-induced apoptosis of retinal ganglion cells (RGC) and in vitro of neuronal SH-SY5Y-cells via activation of soluble guanylate-cyclase (sGC).. To mimic ischemic respiratory arrest, SH-SY5Y-cells were incubated with rotenone (100 nmol/L, 4 h) ± CORM ALF186 (10-100 µmol/L) or inactivated ALF186 lacking the potential of releasing CO. Apoptosis and reactive oxygen species (ROS) production were analyzed using flow-cytometry (Annexin V, mitochondrial membrane potential, CM-H2DCFDA) and Western blot (Caspase-3). The impact of ALF186± respiratory arrest on cell signaling was assessed by measuring expression of nitric oxide synthase (NOS) and soluble guanylate-cyclase (sGC) and by analyzing cellular cGMP levels. The effect of ALF186 (10 mg/kg iv) on retinal IRI in Sprague-Dawley rats was assessed by measuring densities of fluorogold-labeled RGC after IRI and by analysis of apoptosis-related genes in retinal tissue.. ALF186 but not inactivated ALF186 inhibited rotenone-induced apoptosis (Annexin V positive cells: 25 ± 2% rotenone vs. 14 ± 1% ALF186+rotenone, p<0.001; relative mitochondrial membrane potential: 17 ± 4% rotenone vs. 55 ± 3% ALF186+rotenone, p<0.05). ALF186 increased cellular cGMP levels (33±5 nmol/L vs. 23±3 nmol/L; p<0.05) and sGC expression. sGC-inhibition attenuated ALF186-mediated protection (relative mitochondrial membrane potential: 55±3% ALF186+rotenone vs. 20 ± 1% ODQ + ALF186+rotenone, p<0.05). ALF186 protected RGC in vivo (IRI 1255 ± 327 RGC/mm(2) vs. ALF186 + IRI 2036 ± 83; p<0.05) while sGC inhibition abolished the protective effects of ALF186 (ALF186 + IRI 2036 ± 83 RGC/mm(2) vs. NS-2028 + ALF186 + IRI 1263 ± 170, p<0.05).. The CORM ALF186 inhibits IRI-induced neuronal cell death via activation of sGC and may be a useful treatment option for acute ischemic insults to the retina and the brain.

Topics: Animals; Apoptosis; Brain Ischemia; Carbon Monoxide; Cell Line, Tumor; Coordination Complexes; Cyclic GMP; Dose-Response Relationship, Drug; Female; Gene Expression Regulation, Enzymologic; Guanylate Cyclase; Humans; Male; Molybdenum; NADPH Oxidases; Neurons; Neuroprotective Agents; Organometallic Compounds; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Retinal Ganglion Cells; Rotenone; Signal Transduction; Soluble Guanylyl Cyclase

Based upon the well known protective effect of intracellular cyclic guanosine monophosphate (cGMP) accumulation, we tested the hypothesis that storage solution enriched with optimal concentration of the phosphodiestherase-5 inhibitor vardenafil could provide better protection of vascular grafts against reperfusion injury after long-term cold ischaemic storage.. Isolated thoracic aorta obtained from rats underwent 24-h cold ischaemic preservation in physiological saline or vardenafil (10(-11) M)-supplemented saline solution. Reperfusion injury was simulated by hypochlorite (200 μM) exposure for 30 minutes. Endothelium-dependent vasorelaxation was assessed, and histopathological and molecular-biological examination of the aortic tissue were performed.. Compared with the control group, the saline group showed significantly attenuated endothelium-dependent maximal relaxation (Rmax) to acetylcholine after hypoxia-reoxygenation, which was significantly improved by vardenafil supplementation (Rmax control: 98 ± 1%; saline: 48 ± 6%; vardenafil: 75 ± 4%; p < .05). Vardenafil treatment significantly reduced DNA strand breaks (control: 10.6 ± 6.2%; saline: 72.5 ± 4.0%; vardenafil: 14.2 ± 5.2%; p < .05) and increased cGMP score in the aortic wall (control: 8.2 ± 0.6; saline: 4.5 ± 0.3; vardenafil: 6.7 ± 0.6; p < .05).. Our results support the view that impairment of intracellular cGMP signalling plays a role in the pathogenesis of the endothelial dysfunction induced by cold storage warm reperfusion, which can be effectively reversed by pharmacological phosphodiesterase-5 inhibition.

Topics: Animals; Aorta, Thoracic; Apoptosis; Cold Ischemia; Cyclic GMP; Cytoprotection; DNA Damage; Dose-Response Relationship, Drug; Endothelium, Vascular; Gene Expression Regulation; Imidazoles; Male; Organ Preservation; Organ Preservation Solutions; Phosphodiesterase 5 Inhibitors; Piperazines; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sulfones; Time Factors; Triazines; Vardenafil Dihydrochloride; Vascular Grafting; Vascular System Injuries; Vasoconstriction; Vasoconstrictor Agents; Vasodilation; Vasodilator Agents

Previously, we showed that transient inhibition of TGF- β1 resulted in correction of key aspects of diabetes-induced CD34(+) cell dysfunction. In this report, we examine the effect of transient inhibition of plasminogen activator inhibitor-1 (PAI-1), a major gene target of TGF-β1 activation. Using gene array studies, we examined CD34(+) cells isolated from a cohort of longstanding diabetic individuals, free of microvascular complications despite suboptimal glycemic control, and found that the cells exhibited reduced transcripts of both TGF-β1 and PAI-1 compared to age, sex, and degree of glycemic control-matched diabetic individuals with microvascular complications. CD34(+) cells from diabetic subjects with microvascular complications consistently exhibited higher PAI-1 mRNA than age-matched non-diabetic controls. TGF- β1 phosphorodiamidate morpholino oligo (PMO) reduced PAI-1 mRNA in diabetic (p<0.01) and non-diabetic (p=0.05) CD34(+) cells. To reduce PAI-1 in human CD34(+) cells, we utilized PAI-1 siRNA, lentivirus expressing PAI-1 shRNA or PAI-1 PMO. We found that inhibition of PAI-1 promoted CD34(+) cell proliferation and migration in vitro, likely through increased PI3(K) activity and increased cGMP production. Using a retinal ischemia reperfusion injury model in mice, we observed that recruitment of diabetic CD34(+) cells to injured acellular retinal capillaries was greater after PAI-1-PMO treatment compared with control PMO-treated cells. Targeting PAI-1 offers a promising therapeutic strategy for restoring vascular reparative function in defective diabetic progenitors.

Topics: Adult; Animals; Antigens, CD34; Cell Movement; Cell Proliferation; Cells, Cultured; Cohort Studies; Cyclic GMP; Diabetes Mellitus; Diabetic Angiopathies; Humans; Leukocytes, Mononuclear; Mice; Middle Aged; Oligonucleotide Array Sequence Analysis; Phosphatidylinositol 3-Kinases; Plasminogen Activator Inhibitor 1; Reperfusion Injury; Retinal Vessels; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Transcriptome; Transforming Growth Factor beta1

To evaluate the effect of vardenafil on renal function after renal ischemia-reperfusion (IR) injury (IRI) in a rat model.. Seventy-one Wistar rats were divided into 7 groups including (1) a vehicle-treated group, (2) a vehicle pretreated-IR group, (3-6) vardenafil pretreated-IR groups in doses of 0.02, 0.2, 2 and 20 μg/kg, respectively, (7) a group of IR followed by treatment with 2 μg/kg of vardenafil. Vardenafil or vehicle solution was administered one hour before unilateral nephrectomy and the induction of 45 min of ischemia on the contralateral kidney by clamping of renal pedicle. Four hours of reperfusion were allowed after renal ischemia. Studied parameters were serum creatinine, fractional excretion of sodium (FENa), and histological evaluation of renal specimens. In addition, renal tissue cGMP levels, ERK1/2 phosphorylation as well as renal function by renal scintigraphy were also evaluated.. Administration of vardenafil before the induction of ischemia resulted in a significant reduction in creatinine and FENa levels as well as in less histological lesions observed in treated kidneys in comparison with the vehicle-treated group. The underlying mechanism of cytoprotection was cGMP depended and involved the phosphorylation of ERK proteins. Renal scintigraphy confirmed that PDE5 inhibition attenuates renal IRI.. Vardenafil attenuates renal IRI. Based on similar results from relevant studies on other PDE-5 inhibitors in renal and cardiac IRI, it can be assumed that all PDE-5 inhibitors share a common mechanism of cytoprotection.

Topics: Animals; Cyclic GMP; Imidazoles; Ischemic Preconditioning; Kidney; Male; MAP Kinase Signaling System; Models, Animal; Phosphodiesterase 5 Inhibitors; Piperazines; Rats; Rats, Wistar; Reperfusion Injury; Sulfones; Time Factors; Treatment Outcome; Triazines; Vardenafil Dihydrochloride

Nitric oxide (NO) seems to play an important role during renal ischemia/reperfusion (I/R) injury. We investigated whether rutin inhibits inducible nitric oxide synthase (iNOS) and reduces 3-nitrotyrosine (3-NT) formation in the kidneys of rats during I/R.. Wistar albino rats were nephrectomized unilaterally and, 2 weeks later, subjected to 45 minutes of left renal pedicle occlusion followed by 3 hours of reperfusion. We intraperitoneally administered L-N6-(1-iminoethyl)lysine (L-NIL; 3 mg/kg) for 30 minutes or rutin (1 g/kg) for 60 minutes before I/R. After reperfusion, kidney samples were taken for immunohistochemical analysis of iNOS and 3-NT. We measured plasma nitrite/nitrate and cyclic guanosine monophosphate (cGMP) to evaluate NO levels.. Ischemia/reperfusion caused plasma cGMP to increase significantly. Similarly, plasma nitrite/nitrate was elevated in the I/R group compared with the control group. Histochemical staining was positive for iNOS and 3-NT in the I/R group. Pretreatment with L-NIL or rutin significantly mitigated the elevation of plasma cGMP and nitrite/nitrate. These changes in biochemical parameters were also associated with changes in immunohistochemical appearance. Pretreatment with L-NIL or rutin significantly decreased the incidence and severity of iNOS and 3-NT formation in the kidney tissues.. Our findings suggest that high activity of iNOS causes renal I/R injury, and that rutin exerts protective effects, probably by inhibiting iNOS.

Topics: Animals; Biomarkers; Cyclic GMP; Enzyme Inhibitors; Immunohistochemistry; Infusions, Parenteral; Kidney; Lysine; Male; Nephrectomy; Nitrates; Nitric Oxide Synthase Type II; Nitrites; Protective Agents; Rats; Rats, Wistar; Reperfusion Injury; Rutin; Tyrosine

We sought to determine whether ventilation of lungs after death in non-heart-beating donors with carbon monoxide during warm ischemia and ex vivo lung perfusion and after transplant would reduce ischemia-reperfusion injury and improve lung function.. One hour after death, Sprague-Dawley rats were ventilated for another hour with 60% oxygen (control group) or 500 ppm carbon monoxide in 60% oxygen (CO-vent group; n=6/group). Then, lungs were flushed with 20 mL cold Perfadex, stored cold for 1 hour, then warmed to 37 °C in an ex vivo lung perfusion circuit perfused with Steen solution. At 37 °C, lungs were ventilated for 15 minutes with alveolar gas with or without 500 ppm carbon monoxide, then perfusion-cooled to 20 °C, flushed with cold Perfadex and stored cold for 2 hours. The left lung was transplanted using a modified cuff technique. Recipients were ventilated with 60% oxygen with or without carbon monoxide. One hour after transplant, we measured blood gases from the left pulmonary vein and aorta, and wet-to-dry ratio of both lungs. The RNA and protein extracted from graft lungs underwent real-time polymerase chain reaction and Western blotting, and measurement of cyclic guanosine monophosphate by enzyme-linked immunosorbent assay.. Carbon monoxide ventilation begun 1 hour after death reduced wet/dry ratio after ex vivo lung perfusion. After transplantation, the carbon monoxide-ventilation group had better oxygenation; higher levels of tissue cyclic guanosine monophosphate, heme oxidase-1 expression, and p38 phosphorylation; reduced c-Jun N-terminal kinase phosphorylation; and reduced expression of interleukin-6 and interleukin-1β messenger RNA.. Administration of carbon monoxide to the deceased donor and non-heart-beating donor lungs reduces ischemia-reperfusion injury in rat lungs transplanted from non-heart-beating donors. Therapy to the deceased donor via the airway may improve post-transplant lung function.

Topics: Animals; Blotting, Western; Carbon Monoxide; Cyclic GMP; Enzyme-Linked Immunosorbent Assay; Extracellular Signal-Regulated MAP Kinases; Heme Oxygenase (Decyclizing); I-kappa B Kinase; Interleukin-6; JNK Mitogen-Activated Protein Kinases; Lung; Lung Injury; Lung Transplantation; Male; p38 Mitogen-Activated Protein Kinases; Perfusion; Phosphorylation; Rats; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; Reperfusion Injury; Respiration, Artificial; Reverse Transcriptase Polymerase Chain Reaction; Time Factors; Tissue and Organ Harvesting; Warm Ischemia

To explore the protective effect of the phosphodiesterase-5 inhibitor, sildenafil, on lung ischemia-reperfusion injury, 30 rats were randomly divided into 3 groups of 10: a sham-operated group A, a lung ischemia-reperfusion injury group B, and a sildenafil preconditioned group C. A 0.1% sildenafil solution was administrated orally 2 h before establishing an in-vivo lung ischemia-reperfusion model in group C; 0.9% normal saline solution was used in the controls. The lung wet-to-dry ratio, malondialdehyde content, myeloperoxidase and nitric oxide synthase activity in groups B and C were significant higher than those in group A, while the partial pressure of oxygen in arterial blood and cyclic guanosine-3',5'-monophosphate content in groups B and C were significant lower than those in group A. Compared to group B, lung wet/dry ratio, malondialdehyde content, myeloperoxidase and nitric oxide synthase activity in group C were significantly lower, while arterial O(2) and cyclic guanosine-3',5'-monophosphate content in group C were significantly higher. The expected histological and cytological changes were significantly alleviated in group C. Oral preconditioning with sildenafil prevented rat lung ischemia-reperfusion injury and improved pulmonary function. The mechanisms of this effect might be prevention of cyclic guanosine monophosphate degradation and inhibition of nitric oxide synthase activity.

Topics: Alveolar Epithelial Cells; Animals; Cyclic GMP; Endothelium, Vascular; Lung Diseases; Male; Malondialdehyde; Nitric Oxide Synthase; Peroxidase; Phosphodiesterase 5 Inhibitors; Piperazines; Purines; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sildenafil Citrate; Sulfones

Remote ischemic preconditioning (RIPC) protects against liver ischemia reperfusion (IR) injury. An essential circulating mediator of this protection is nitric oxide (NO) induced by lower limb RIPC. One of the mechanisms through which NO generally acts is the soluble guanylyl cyclase-cyclic GMP (sGC-cGMP) pathway. The present study aimed to assess the role of hepatic sGC-cGMP in lower limb RIPC-induced protection against liver IR injury.. Mice were allocated to 4 groups: 1.Sham; 2.IR: 40 min of lobar hepatic ischemia and 2 hr reperfusion; 3.RIPC+IR: 6 cycles of 4x4 min IR of the lower limb followed by IR group procedure; (4) 1H-[1,2,4]oxadiazole[4,3-a]quinoxalin-1-one (ODQ)+RIPC+IR: ODQ (sGC inhibitor) was administered followed by RIPC+IR group procedure. Hepatic microcirculatory blood flow (MBF) was measured throughout the experiment. Plasma transaminases, hepatic histopathological and transmission electron microscopy studies were performed at the end of the experiment. Hepatic cGMP levels were measured in groups 1-3 in addition to an RIPC alone group.. Compared to liver IR alone, RIPC+IR increased hepatic MBF during liver reperfusion (P<0.05), and reduced plasma transaminases (P<0.05) and ultrastructural markers of injury. In contrast compared to RIPC+IR, ODQ+RIPC+IR decreased hepatic MBF (P<0.05) and ultrastructural markers of injury. However, plasma transaminases were not significantly different in the ODQ+RIPC+IR compared to the RIPC+IR group. Hepatic cGMP levels were significantly elevated in the RIPC compared to sham group.. The hepatic sGC-cGMP pathway is required for mediating the protective effects of lower limb RIPC on hepatic MBF in liver IR injury.

Topics: Animals; Cyclic GMP; Disease Models, Animal; Follow-Up Studies; Guanylate Cyclase; Ischemic Preconditioning; Liver; Liver Circulation; Liver Diseases; Mice; Microcirculation; Microscopy, Electron, Transmission; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Soluble Guanylyl Cyclase

To evaluate immunological protection of nitric oxide (NO) in hepatopulmonary syndrome and probable mechanisms of ischemia-reperfusion (IR) injury in rat liver transplantation.. Sixty-six healthy male Wistar rats were randomly divided into three groups (11 donor/recipient pairs). In group II, organ preservation solution was lactated Ringer's solution with heparin 10, 000/μL at 4 °C. In groups I and III, the preservation solution added, respectively, L-arginine or N(G)-L-arginine methyl ester (L-NAME) (1 mmol/L) based on group II, and recipients were injected with L-arginine or L-NAME (50 mg/kg) in the anhepatic phase. Grafted livers in each group were stored for 6 h and implanted into recipients. Five rats were used for observation of postoperative survival in each group. The other six rats in each group were used to obtain tissue samples, and executed at 3 h and 24 h after transplantation. The levels of alanine aminotransferase (ALT), tumor necrosis factor (TNF)-α and NO metabolites (NOx) were detected, and expression of NO synthase, TNF-α and intercellular adhesion molecule 1 (ICAM-1) was examined by triphosphopyridine nucleotide diaphorase histochemical and immunohistochemical staining.. By supplementing L-arginine to strengthen the NO pathway, a high survival rate was achieved and hepatic function was improved. One-week survival rate of grafted liver recipients in group I was significantly increased (28.8 ± 36.6 d vs 4 ± 1.7 d, P < 0.01) as compared with groups II and III. Serum levels of ALT in group I were 2-7 times less than those in groups II and III (P < 0.01). The cyclic guanosine monophosphate (cGMP) levels in liver tissue and NOx in group I were 3-4 times higher than those of group II after 3 h and 24 h reperfusion, while in group III, they were significantly reduced as compared with those in group II (P < 0.01). The levels of TNF-α in group I were significantly lower than in group II after 3 h and 24 h reperfusion (P < 0.01), while being significantly higher in group III than group II (P < 0.01). Histopathology revealed more severe tissue damage in graft liver and lung tissues, and a more severe inflammatory response of the recipient after using NO synthase inhibitor, while the pathological damage to grafted liver and the recipient's lung tissues was significantly reduced in group I after 3 h and 24 h reperfusion. A small amount of constitutive NO synthase (cNOS) was expressed in liver endothelial cells after 6 h cold storage, but there was no expression of inducible NO synthase (iNOS). Expression of cNOS was particularly significant in vascular endothelial cells and liver cells at 3 h and 24 h after reperfusion in group II, but expression of iNOS and ICAM-1 was low in group I. There was diffuse strong expression of ICAM-1 and TNF-α in group III at 3 h after reperfusion.. The NO/cGMP pathway may be critical in successful organ transplantation, especially in treating hepatopulmonary syndrome during cold IR injury in rat orthotopic liver transplantation.

Topics: Alanine Transaminase; Animals; Arginine; Cyclic GMP; Disease Models, Animal; Enzyme Inhibitors; Hepatorenal Syndrome; Immunohistochemistry; Intercellular Adhesion Molecule-1; Liver; Liver Transplantation; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Wistar; Reperfusion Injury; Time Factors; Tumor Necrosis Factor-alpha

The vasodegenerative phase of diabetic retinopathy is likely caused by endothelial dysfunction and reduced endothelial repair. Migration of endothelial progenitor cells (EPCs) into areas of vascular injury is critical to vascular repair. This key function, often defective in diabetes, is largely mediated by nitric oxide (NO), which is known to be inactivated by superoxide produced by NADPH oxidase. The authors tested the hypothesis that either increasing eNOS expression or inhibiting NADPH oxidase would restore the reparative function in diabetic EPCs.. Peripheral blood was obtained from healthy (n = 27) and diabetic (n = 31) persons, and CD34(+) cells were isolated. Expression and activation of eNOS and NADPH oxidase and intracellular levels of NO, superoxide, and peroxynitrite were evaluated. cGMP production and migration to SDF-1α were also determined. Reparative function was evaluated in a mouse model of retinal ischemia-reperfusion injury.. Diabetic EPCs demonstrate reduced eNOS expression and decreased NO bioavailability and migration in response to SDF-1α. Increasing eNOS expression in diabetic cells by AVE3085 resulted in increased peroxynitrite levels and, therefore, did not enhance NO-mediated functions in vitro and in vivo. Expression of Nox2, NADPH oxidase activity, and superoxide levels were higher in diabetic than in nondiabetic EPCs. Pretreatment with apocynin or gp91ds-tat increased NO bioavailability without increasing eNOS activity in response to SDF-1α. Ex vivo NADPH oxidase inhibition in diabetic cells restored migratory function in vitro and enhanced their homing to ischemic retinal vasculature in vivo.. The NADPH oxidase system is a promising target for correcting vasoreparative dysfunction in diabetic EPCs.

Topics: Acetophenones; Adult; Animals; Antigens, CD34; Benzodioxoles; Chemokine CXCL12; Cyclic GMP; Diabetic Retinopathy; Disease Models, Animal; Endothelium, Vascular; Enzyme Inhibitors; Female; Glycoproteins; Humans; Indans; Male; Mice; Middle Aged; NADPH Oxidases; Nitric Oxide; Nitric Oxide Synthase Type III; Peroxynitrous Acid; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; Superoxides; Young Adult

To investigate the interactions between inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in response to ischaemia-reperfusion (I/R) of rabbit bladder.. Rabbit bladders were exposed to 2 h of ischaemia by bilaterally clamping the major arteries entering the bladder and then a subsequent 36 h of reperfusion (I/R) with or without intraperitoneal administration of a selective iNOS inhibitor n-(3-(amynomethyl)benzyl)acetamidine (1400W) or a selective COX-2 inhibitor NS-398 given 1 h before killing. The bladder tissues were processed for isometric tension experiments, enzymatic NOS activitiy, tissue contents of nitrite/nitrate (NO(X) ), cyclic guanosine monophosphate (cGMP) and COX activity determined by prostaglandin E(2) (PGE(2) ) production.. iNOS and constitutive NOS (cNOS) activities, NO(X) and PGE(2) contents in the bladder tissues at 36 h after reperfusion were significantly higher than those in the sham group with no significant increase in cGMP. Treatment with 1400W abrogated the increases in iNOS activity and NO(X) as well as PGE(2) without changing cNOS activity. In the tension experiments, a NOS substrate, l-arginine, induced detrusor contraction only in the I/R group, which was inhibited by 1400W or NS-398 but not by a selective soluble guanylate cyclase inhibitor 1H-[1,2,4] oxadiazole[4,3-a]quinoxalin-1-one (ODQ). 8-Br-cGMP induced detrusor relaxation in the sham and I/R groups. Also, l-arginine increased NO(X) and PGE(2) in the bladder tissues only in the I/R group, which were inhibited by pretreatment with 1400W. While, l-arginine increased cGMP contents in the I/R group and this increase was suppressed by ODQ but not by 1400W.. These results show that NO derived from an up-regulation of iNOS after I/R increases COX-2-derived PG via a cGMP-independent mechanism. NO-mediated activation of COX-2 may be an important mechanism for the modulation of bladder function after I/R injury.

Topics: Animals; Cyclic GMP; Cyclooxygenase 2; Male; Nitric Oxide; Nitric Oxide Synthase Type II; Rabbits; Reperfusion Injury; Up-Regulation; Urinary Bladder

Post-stroke depression (PSD) is one of the psychiatric complications after stroke. Present study was conducted to elucidate the protective effect of sertraline and possible involvement of nitric oxide mechanism against transient global ischemia induced behavioral despair. Bilateral common carotid artery occlusion was given twice for 5 min at 10 min interval followed by 96 h reperfusion. Ischemia reperfusion significantly increased immobility period and decreased resistance to lateral push as compared to sham-operated group. Ischemia reperfusion caused significant oxidative damage and mitochondrial enzyme complex (I-III) dysfunction as compared to sham group. Sertraline (5 and 10mg/kg) treatment significantly reduced immobility period, increased resistance to lateral push, attenuated oxidative damage and restored mitochondrial enzyme complex activities as compared to ischemia group. L-Arginine (100mg/kg) or sildenafil (5mg/kg) pretreatment with sertraline (5mg/kg) significantly reversed the protective effect of sertraline. However, L-NAME (10mg/kg) or 7NI (10mg/kg) pretreatment with sertraline (5mg/kg) significantly potentiated their protective effect which were significant as compared to their effect alone. The present study shows that nitric oxide modulation is involved in the protective effect of sertraline.

Topics: Adult; Animals; Antidepressive Agents; Behavior, Animal; Cell Respiration; Cyclic GMP; Depression; Disease Models, Animal; Enzyme Inhibitors; Humans; Ischemic Attack, Transient; Mice; Motor Activity; Neuroprotective Agents; Neuropsychological Tests; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Donors; Oxidative Stress; Random Allocation; Reperfusion Injury; Sertraline

Nitroglycerin (NTG) has been reported to possess preconditioning-like (PCL) protections on heart and other tissues. Our previous studies showed that NTG has acute PCL effects on rat small intestine. The present studies were designed to study whether NTG has delayed PCL protection on rat small intestine and to explore its mechanism(s).. The intestine lesions were evaluated by histologic examination and serum lactate dehydrogenase (LDH) measurement. The effects of nitric oxide (NO), cGMP, and alpha-calcitonin gene-related peptide (CGRP) synthesis on the effects of NTG were analyzed.. Pretreatment with NTG (0.12 mg/kg i.v.) 24 h before ischemia-reperfusion (I/R) of super mesenteric artery significantly reduced histologic lesions and serum LDH with elevated blood levels of NO and CGRP. Inhibition of guanylate cyclase by methylene blue (30 mg/kg i.p.) or specific depletion of transmitters in capsaicin-sensitive sensory nerve by capsaicin (50 mg/kg s.c.) abrogated the protection conferred by NTG. Reverse-transcription polymerase chain reaction analysis showed that NTG upregulates the expression of alpha-CGRP messenger RNA (mRNA), but not beta-CGRP mRNA in lumbar dorsal root ganglia.. In conclusion, NTG prevents rat small intestine from I/R injury by delayed PCL effects 24 h after administration. The protective effects are mediated by NO-cGMP pathway and alpha-CGRP upregulation.

Topics: Animals; Calcitonin Gene-Related Peptide; Capsaicin; Cyclic GMP; Guanylate Cyclase; Intestine, Small; Ischemic Preconditioning; Male; Nitric Oxide; Nitroglycerin; Rats; Rats, Wistar; Reperfusion Injury; Vasodilator Agents

Development of intracellular calcium overload is an important pathophysiological factor in myocardial ischemia/reperfusion or anoxia/reoxygenation injury. Recent studies have shown that Sodium Ferulate (SF) stimulates nitric oxide (NO) production and exerts a cardioprotective effect in the ischemia-reperfused heart. However, it has not been determined whether the cardioprotection of SF is associated with suppression of Ca(2+) overload via NO/cyclic GMP (cGMP)/cGMP-dependent protein kinase (PKG) pathway. In this work, after cardiomyocytes were incubated with 100, 200, 400, or 800 microM SF for 3 h, anoxia/reoxygenation injury was induced and intracellular Ca(2+) concentration, NO synthase (NOS) activity, guanylate cyclase activity, NO, and cGMP formation were measured appropriately. The results showed that treatment with SF concentration-dependently inhibited calcium overload induced by anoxia/reoxygenation. We also demonstrated that SF (100-800 microM) concentration dependently enhanced NO and cGMP formation through increasing NOS activity and guanylate cyclase activity in the cardiomyocytes. On the contrary, inhibition of calcium overload by SF was markedly attenuated by addition of an NOS inhibitor, an NO scavenger, an soluble guanylate cyclase inhibitor, and a PKG inhibitor: N(G)-nitro-l-arginine methyl ester (L-NAME, 100 microM), 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazole-1-oxyl-3-oxide (c-PTIO, 1.0 microM), 1H-[1, 2, 4] oxadiazolo [4, 3-alpha] quinoxalin-1-one (ODQ, 20 microM) and KT5823 (0.2 microM), respectively. Our findings indicate that SF significantly attenuates anoxia/reoxygenation-induced Ca(2+) overload and improves cell survival in cultured cardiomyocytes through NO/cGMP/PKG signal pathway.

Topics: Animals; Animals, Newborn; Calcium; Coumaric Acids; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Gene Expression Regulation; Guanylate Cyclase; Hypoxia; Myocytes, Cardiac; Nitric Oxide; Nitric Oxide Synthase; Oxygen; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction; Solubility

Ischemia-reperfusion injury remains a devastating complication of lung transplantation. Phosphodiesterase inhibitors have been shown to precondition tissues against ischemia-reperfusion injury. Little is known, however, about the utility of phosphodiesterase inhibition in reperfusion injury after lung transplantation. We evaluated the long-acting phosphodiesterase-5 inhibitor, tadalafil, in an ex vivo lung transplant model.. New Zealand White rabbits (4 kg), were given oral tadalafil (n = 11) 24 hours before lung harvest and compared with rabbits given oral vehicle alone (n = 11). Lungs were recovered with Perfadex solution (Vitrolife, Kungsbacka, Sweden) and cold stored for 18 hours. After storage, lung blocks were reperfused with donor rabbit blood in an ex vivo apparatus. Pulmonary artery pressures were recorded with serial arterial and venous blood gas sampling and animals served as their own controls. Phosphodiesterase-5 and protein kinase G tissue activity assays confirmed drug effects. Luminol chemiluminescence assay was used to measure reactive oxygen species and levels of endothelial and inducible nitric oxide synthase were measured.. Extended cold storage, followed by reperfusion produced a consistent reproducible decrease in oxygenation and increase in pulmonary pressure. Tadalafil-treated animals exhibited greater Pao(2) throughout the course of reperfusion (P = .001) Mean pulmonary artery pressure was lower in tadalafil-treated animals (22 vs 40 mm Hg; P = .04). Phosphodiesterase-5 activity was decreased (143 +/- 8 vs 205 +/- 32 mP; P < .001) with protein kinase G activity increased (25 +/- 12 vs 12 +/- 2.4 fU/microg; P = .01) in the experimental group confirming that oral pretreatment resulted in active phosphodiesterase inhibition in the lung tissue. Reactive oxygen species (as measured by luminol activity) were decreased in tadalafil-treated animals (7.8 +/- 1.5 vs 10.2 +/- 1.2 relative light units; P = .003).. Our experimental model demonstrates that oral donor pretreatment with a long-acting phosphodiesterase inhibitor is an effective strategy for improving pulmonary performance after reperfusion. Importantly, phosphodiesterase enzymes and their downstream effectors may play a critical role in reperfusion injury after lung transplantation.

Topics: Administration, Oral; Analysis of Variance; Animals; Carbolines; Cyclic GMP; Delayed-Action Preparations; Disease Models, Animal; Graft Rejection; Graft Survival; Immunohistochemistry; Lung Transplantation; Phosphodiesterase Inhibitors; Preoperative Care; Probability; Rabbits; Random Allocation; Reactive Oxygen Species; Reference Values; Reperfusion Injury; Tadalafil; Tissue and Organ Harvesting

Sildenafil was the first selective inhibitor of phosphodiesterase-5 (PDE5) to be widely used for treating erectile dysfunction. Many recent studies have investigated the cardioprotective role of sildenafil in animal models. We evaluated the protective effects of sildenafil in experimental renal ischemia-reperfusion (IR) injury in two studies. In study 1, male Sprague-Dawley rats were divided into four groups: sham, sildenafil-treated sham, vehicle-treated IR, and sildenafil-treated IR groups. In study 2, we divided the rats into two groups: sildenafil-treated IR rats and PD98059 (ERK inhibitor)+sildenafil-treated IR rats. Functional parameters of the kidney were evaluated at the molecular and structural levels. Blood urea nitrogen (BUN) and serum creatinine levels were lower in sildenafil-treated IR rats than in vehicle-treated IR rats. The expression of inducible (iNOS) and endothelial nitric oxide synthase (eNOS) proteins in sildenafil-treated IR rats was significantly higher than in vehicle-treated IR rats. Pretreatment with sildenafil in IR rats increased ERK phosphorylation and reduced the renal Bax/Bcl-2 ratio, renal caspase-3 activity, and terminal dUTP nick end-labeling-positive apoptotic cells. In contrast, PD98059 treatment increased BUN and serum creatinine levels and attenuated the sildenafil-induced expression of pERK, iNOS, eNOS, and Bcl-2. PD98059 also increased caspase-3 activity but did not decrease the sildenafil-induced accumulation of cGMP. In conclusion, this study suggests that sildenafil has antiapoptotic effects in experimental IR renal injury via ERK phosphorylation, induction of iNOS and eNOS production, and a decrease in the Bax/Bcl-2 ratio.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Blood Urea Nitrogen; Caspase 3; Creatinine; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Drug Administration Schedule; Extracellular Signal-Regulated MAP Kinases; Flavonoids; Kidney Diseases; Male; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Phosphodiesterase 5 Inhibitors; Phosphodiesterase Inhibitors; Phosphorylation; Piperazines; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-bcl-2; Purines; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sildenafil Citrate; Sulfones; Time Factors

Lungs from non-heart-beating donors (NHBDs) would enhance the donor pool. Ex vivo perfusion and ventilation of NHBD lungs allows functional assessment and treatment. Ventilation of rat NHBD lungs with nitric oxide (NO) during ischemia, ex vivo perfusion and after transplant reduced ischemia-reperfusion injury (IRI) and improved lung function posttransplant. One hour after death, Sprague-Dawley rats were ventilated for another hour with either 60% O2 or 60% O2/40 ppm NO. Lungs were then flushed with 20-mL cold Perfadex, stored cold for 1 h, perfused in an ex vivo circuit with Steen solution and warmed to 37 degrees C, ventilated 15 min, perfusion-cooled to 20 degrees C, then flushed with cold Perfadex and stored cold. The left lung was transplanted and ventilated separately. Recipients were sacrificed after 1 h. NO-ventilation was associated with significantly reduced wet:dry weight ratio in the ex vivo circuit, better oxygenation, reduced pulmonary vascular resistance, increased lung tissue levels of cGMP, maintained endothelial NOS eNOS, and reduced increases in tumor necrosis factor alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS). NO-ventilation had no effect on MAP kinases or NF-kappaB activation. NO administration to NHBDs before and after lung retrieval may improve function of lungs from NHBDs.

Topics: Animals; Cyclic GMP; Lung; Lung Transplantation; Mitogen-Activated Protein Kinases; Myocardial Contraction; Nitric Oxide; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Organ Preservation; Pulmonary Edema; Rats; Reperfusion Injury; Tissue Donors; Tumor Necrosis Factor-alpha

Nitric oxide has prosurvival effects that can limit ischemia-reperfusion injuries. However, the matrix glycoprotein thrombospondin-1 is induced following ischemia-reperfusion injury and limits nitric oxide signaling by engaging its cell surface receptor CD47. In this article, the authors examine whether postinjury blocking of this inhibitory signal can protect from ischemia-reperfusion injury in a rat flap model.. A total of 40 tissue flaps were created in rats based solely on the deep inferior epigastric vessels. Microvascular clamps were used to create 45 minutes of ischemia time to the flaps. The flaps were then treated using a monoclonal antibody to CD47 or an isotype-matched control immunoglobulin G1 5 or 30 minutes after clamp removal. Twenty-four or 72 hours postoperatively, the necrotic area of the flap was determined, and serum, deep inferior epigastric vessels, and flaps were harvested for analysis from five rats in each respective group.. Treatment with a CD47 antibody 5 minutes after reperfusion significantly reduces flap necrosis compared with immunoglobulin G1 control (9 percent versus 43 percent; p < 0.01). The protective effect is even more dramatic when treatment is delayed until 30 minutes after reperfusion (10 percent versus 88 percent for control; p < 0.01). Markers of neutrophil and endothelial cell activation along with total leukocytes are reduced in CD47 antibody-treated flaps, as are tissue malondialdehyde levels. Levels of cyclic guanosine monophosphate are elevated 72 hours postoperatively in the CD47 antibody-treated deep inferior epigastric vessels versus the control flaps.. Therapies targeting the thrombospondin-1 receptor CD47 offer potential for increasing tissue survival in ischemia-reperfusion injuries. The ability to protect when given after ischemia-reperfusion injury enables a broader clinical applicability.

Topics: Analysis of Variance; Animals; Antibodies, Monoclonal; CD47 Antigen; Constriction; Cyclic GMP; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Immunohistochemistry; Interferon-gamma; Lipid Peroxidation; Male; Malondialdehyde; Nitric Oxide; Probability; Random Allocation; Rats; Rats, Inbred F344; Reperfusion Injury; Signal Transduction; Surgical Flaps

Recently, the infarct reducing and cardioprotective effects of phosphodiesterase-5-inhibitors were described. In this study, we investigated these effects on ischemia/reperfusion injury in a rat model of heart transplantation. Three groups were assigned for our study: a vardenafil preconditioning group, an ischemic control, and a nonischemic control. Hemodynamic parameters were significantly increased in the vardenafil group (Pmax: 82+/-4 vs. 110+/-12 vs. 127+/-13 mm Hg; dP/dtmax: 1740+/-116 vs. 3197+/-599 vs. 4397+/-602 mm Hg/sec; ischemic control vs. vardenafil vs. nonischemic control; P<0.05 vs. ischemic control). Furthermore, we recorded increased ATP levels and significantly less apoptosis in the treatment group after terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (apoptosis index: 27.23%+/-1.54% vs. 16.77%+/-1.42% vs. 18.86%+/-1.07%; ischemic control vs. vardenafil vs. nonischemic control; P<0.05 vs. ischemic control). Our current results support the concept that the cGMP-PKG-pathway plays an important role in ischemia/reperfusion injury. We could show that up-regulating this pathway has a preconditioning-like effect and can effectively reduce ischemia/reperfusion injury.

Topics: Animals; Aorta, Abdominal; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Heart Transplantation; Hemodynamics; Imidazoles; Male; Myocardial Contraction; Phosphodiesterase Inhibitors; Piperazines; Postoperative Complications; Rats; Rats, Inbred Lew; Reperfusion Injury; Sulfones; Systole; Transplantation, Heterotopic; Transplantation, Isogeneic; Triazines; Vardenafil Dihydrochloride; Vena Cava, Inferior; Ventricular Function, Left

We investigated whether alpha-lipoic acid (alpha-LA), an antioxidant, attenuates the ischemia-reperfusion (I/R)-induced dysregulation of these transporters. Both renal pedicles of male Sprague-Dawley rats were clamped for 40 min. alpha-LA (80 mg/kg) was administered intraperitoneally before and immediately after induction of ischemia. After 2 days, the expression of aquaporins (AQPs), sodium transporters, and nitric oxide synthases (NOS) was determined in the kidney by immunoblotting and immunohistochemistry. The expression of endothelin-1 (ET-1) mRNA was determined by real-time PCR. Activities of adenylyl cyclase and guanylyl cyclase were measured by stimulated generation of cAMP and cGMP, respectively. The expression of AQP1-3 as well as that of the alpha(1)-subunit of Na-K-ATPase, type 3 Na/H exchanger, Na-K-2Cl cotransporter, and Na-Cl cotransporter was markedly decreased in response to I/R. The expression of type VI adenylyl cyclase was decreased in I/R-injured rats, which was counteracted by the treatment of alpha-LA. AVP-stimulated cAMP generation was blunted in I/R rats and was then ameliorated by alpha-LA treatment. alpha-LA treatment attenuated the downregulation of AQPs and sodium transporters. The expression of endothelial NOS was decreased in I/R rats, which was prevented by alpha-LA. The cGMP generation in response to sodium nitroprusside was blunted in I/R rats, which was also significantly prevented by alpha-LA. The mRNA expression of ET-1 was increased, which was recovered to the control level by alpha-LA treatment. In conclusion, alpha-LA treatment prevents I/R-induced dysregulation of AQPs and sodium transporters in the kidney, possibly through preserving normal activities of local AVP/cAMP, nitric oxide/cGMP, and ET systems.

Topics: Adenylyl Cyclases; Animals; Antioxidants; Aquaporins; Cyclic GMP; Endothelin-1; Guanylate Cyclase; Kidney; Male; Models, Animal; Nitric Oxide; Rats; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Sodium-Hydrogen Exchanger 3; Sodium-Hydrogen Exchangers; Sodium-Potassium-Chloride Symporters; Sodium-Potassium-Exchanging ATPase; Soluble Guanylyl Cyclase; Solute Carrier Family 12, Member 1; Thioctic Acid

Ischemia-reperfusion (IR) causes human lung injury in association with the release of atrial and brain natriuretic peptides (ANP and BNP), but the role of ANP/BNP in IR lung injury is unknown. ANP and BNP bind to natriuretic peptide receptor-A (NPR-A) generating cGMP and to NPR-C, a clearance receptor that can decrease intracellular cAMP. To determine the role of NPR-A signaling in IR lung injury, we administered the NPR-A blocker anantin in an in vivo SWR mouse preparation of unilateral lung IR. With uninterrupted ventilation, the left pulmonary artery was occluded for 30 min and then reperfused for 60 or 150 min. Anantin administration decreased IR-induced Evans blue dye extravasation and wet weight in the reperfused left lung, suggesting an injurious role for NPR-A signaling in lung IR. In isolated mouse lungs, exogenous ANP (2.5 nM) added to the perfusate significantly increased the filtration coefficient sevenfold only if lungs were subjected to IR. This effect of ANP was also blocked by anantin. Unilateral in vivo IR increased endogenous plasma ANP, lung cGMP concentration, and lung protein kinase G (PKG(I)) activation. Anantin enhanced plasma ANP concentrations and attenuated the increase in cGMP and PKG(I) activation but had no effect on lung cAMP. These data suggest that lung IR triggered ANP release and altered endothelial signaling so that NPR-A activation caused increased pulmonary endothelial permeability.

Topics: Animals; Atrial Natriuretic Factor; Cell Adhesion Molecules; Cyclic AMP; Cyclic GMP; Guanylate Cyclase; Lung; Male; Mice; Mice, Inbred Strains; Microfilament Proteins; Perfusion; Phosphoproteins; Receptors, Atrial Natriuretic Factor; Reperfusion Injury; Respiratory Distress Syndrome; Signal Transduction

Ischemic preconditioning (IPC) constitutes an endogenous protective mechanism in which one or more brief periods of myocardial ischemia and reperfusion render the myocardium resistant to a subsequent more-sustained ischemic insult. Pharmacological preconditioning represents an ideal alternative of IPC. We now describe the design and synthesis of indole, quinoline, and purine systems with an attached pharmacophoric nitrate ester group. The indole and quinoline derivatives 4 and 5 possess structural features of the nitrate containing K(ATP) channel openers. Purine analogues 11 and 12, substituted at the position 6 by a piperidine moiety and at position 9 by an alkyl nitrate, could combine the effects of the nitrate containing K(ATP) channel openers and those of adenosine. Compound 13 bears the nicotinamide moiety of nicorandil instead of nitrate ester. Compounds 4, 5, and 11 reduced infarction and the levels of malondialdehyde (MDA) at reperfusion in anesthetized rabbits. Compounds 12 and 13 did not significantly reduce the infarct size. Analogues 4 and 5 increased cGMP and MDA during ischemia, while combined analogue 4 and mitoK(ATP) blocker 5-hydroxydecanoic acid (5-HD) abrogated this benefit suggesting an action through mitoK(ATP) channel opening. Treatment with derivative 11 combined with 5-HD as well as treatment with 11 and adenosine receptor blocker 8-(p-sulfophenyl)theophylline (SPT) did not abrogate cardioprotection. Compound 11 is a lead molecule for the synthesis of novel analogues possessing a dual mode of action through cGMP-mitoK(ATP) channel opening-free radicals and through adenosine receptors.

Topics: Animals; Cyclic GMP; Drug Design; Esters; Heterocyclic Compounds; Ischemic Preconditioning; Male; Malondialdehyde; Molecular Structure; Nitrates; Rabbits; Reperfusion Injury; Structure-Activity Relationship

Since the generation of nitric oxide (NO) is an essential step in the trigger phase of ischemic preconditioning, short-term inhalation of NO before ischemia should ameliorate ischemia/reperfusion (I/R) injury of the lung. We tested this hypothesis in high oxygen (>99%) ventilated rats in order to additionally evaluate compatibility of NO and exposure to hyperoxia. Male adult Sprague-Dawley rats inhaled NO (15 ppm, 10 min) before the left lung hilum was clamped for 1 h, and the reperfusion phase was observed for 4 h (NO group). Animals in the I/R group underwent the same treatment, but without NO inhalation. A third group without I/R served as time-matched controls. Animals in the I/R group showed severe I/R injury in terms of arterial pO2 (apO2), which was reduced to 22% of surgical controls (SCs) at time point 30 min reperfusion, and increased endothelial permeability (Evans blue procedure). The pretreatment with NO attenuated these effects. The pO2 after 4 h reperfusion was still 3.0-fold higher in the NO group compared to I/R. In contrast, the I/R- and hyperoxia-induced invasion of leukocytes, as determined by measuring myeloperoxidase (MPO) activity, was not affected by NO. These data were correlated with the activity of major cellular signaling pathways by measuring the phosphorylation at activating and inhibitory sites of extracellular-signal regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38, protein kinase B (AKT), and glycogen synthase kinase 3beta (GSK-3beta), and by determination of cGMP in plasma and lung tissue. Inhalation of NO partly prevented the loss of activation by I/R and hyperoxic ventilation of ERK, JNK, and AKT, and it reduced the I/R-induced activation of GSK-3beta. The level of cGMP in plasma and lung tissue was increased in the NO group after 4 h reperfusion. In conclusion, application of inhaled NO in the preconditioning mode prevented I/R injury in the rat lung without interfering effects of hyperoxic ventilation. The effects of NO on cellular signaling pathways resemble mechanisms of ischemic preconditioning, but further studies have to evaluate the physiological relevance of these results.

Topics: Administration, Inhalation; Animals; Capillary Permeability; Cyclic GMP; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Glycogen Synthase Kinases; Hyperoxia; Ischemia; Ischemic Preconditioning; Lung; Male; MAP Kinase Signaling System; Nitric Oxide; Proto-Oncogene Proteins c-akt; Pulmonary Gas Exchange; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Signal Transduction

Cerebral microvascular endothelial cells form the anatomical basis of the blood-brain barrier (BBB), and the tight junctions of the BBB are critical for maintaining brain homeostasis and low permeability. Ischemia/reperfusion is known to damage the tight junctions of BBB and lead to permeability changes. Here we investigated the protective role of 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (YC-1), against chemical hypoxia and hypoxia/reoxygenation (H/R)-induced BBB hyperpermeability using adult rat brain endothelial cell culture (ARBEC). YC-1 significantly decreased CoCl2- and H/R-induced hyperpermeability of fluorescein isothiocyanate (FITC)-dextran in cell culture inserts. It was found that the decrease and disorganization of tight junction protein zonular occludens-1 (ZO-1) in response to CoCl2, and H/R was antagonized by YC-1. The protection of YC-1 may result from the inhibition of HIF-1alpha accumulation and production of its downstream target vascular endothelial growth factor (VEGF). VEGF alone significantly increased FITC-dextran permeability and down-regulated mRNA and protein levels of ZO-1 in ARBECs. We further used animal model to examine the effect of YC-1 on BBB permeability after cerebral ischemia/reperfusion. It was found that YC-1 significantly protected the BBB against ischemia/reperfusion-induced injury. Taken together, these results indicate that YC-1 may inhibit HIF-1alpha accumulation and VEGF production, which in turn protect BBB from injury caused by hypoxia.

Topics: Animals; Blood-Brain Barrier; Cell Hypoxia; Cells, Cultured; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Hypoxia-Inducible Factor 1, alpha Subunit; Indazoles; Ischemia; Male; Membrane Proteins; Permeability; Phosphoproteins; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Vascular Endothelial Growth Factor A; Zonula Occludens-1 Protein

Nitric oxide (NO) has been shown as an important signaling messenger involved in cardioprotection of ischemic preconditioning (IPC). To date, most studies suggest that NO might provide its protective effects by regulating the mitochondrial ATP-sensitive potassium (K(ATP)) channel via the classic NO/cGMP-dependent pathway. However, there is emerging data suggesting that NO might also elicit its physiological role through protein S-nitrosylation. Protein S-nitrosylation, the covalent attachment of an NO moiety to sulfhydryl group(s) of cysteine residue(s) of proteins, is a reversible post-translational protein modification involved in redox-based cellular signaling. IPC has been found to increase S-nitrosothiol content and result in increased S-nitrosylation of proteins, which not only induces the structural and functional changes of modified proteins, but also prevents the target cysteine residue(s) from the further oxidative modification. In addition, S-nitrosothiols could elicit pharmacological preconditioning effect and protect against myocardial ischemia-reperfusion injury. Thus, protein S-nitrosylation is emerging as an important contributor to cardioprotection in IPC, providing protection from cellular oxidative and nitrosative stress.

Topics: Cyclic GMP; Heart; Ischemic Preconditioning; Mitochondria; Myocardial Reperfusion Injury; Nitric Oxide; Oxidation-Reduction; Protein S; Reperfusion Injury; Signal Transduction

This study is to investigate the effect of gamma-hydroxybutyric acid receptor (GHBR) on focal cerebral ischemia-reperfusion injury in rats and its mechanism. NCS-356 (the agonist of GHBR) and NCS-382 (the antagonist of GHBR) were adopted as the tool medicine. The ripe male Sprague-Dawley rats weighing 240 - 280 g were randomly divided into seven groups: sham operation group (sham), ischemia-reperfusion group (Isc/R), NCS-356 160 microg x kg(-1) group (N1), NCS-356 320 microg x kg(-1) group (N2), NCS-356 640 microg x kg(-1) group (N3), NCS-382 640 microg x kg(-1) + NCS-356 640 microg x kg(-1) group (NCS-382 + N3), and nimodipine (Nim) 600 microg x kg(-1) group. The middle cerebral artery occlusion (MCAO) model referring to Longa's method with modifications was adopted. The effect of GHBR on behavioral consequence of MCAO rats was studied after 2 h of ischemia-reperfusion. After 24 h of ischemia-reperfusion, part of animals were used to measure the cerebral infarction volume by TTC staining; ischemic cortex of another part of animals were used to measure the content of intracellular free calcium by flow cytometry, the tNOS, iNOS activity and the content of NO by spectrophotometric method, the content of cGMP by radioimmunoassay. The neurological function score and infarction volume rate in Isc/R group rats increased significantly than that in sham group; The content of intracellular calcium ([Ca2+]) of cortex neuron and cGMP, the activities of tNOS and iNOS, and the content of NO in Isc/R group were higher than that in sham group obviously (P < 0.01); These consequence we mentioned of N1, N2, N3 and Nim group were lower than that of Isc/R. NCS-382 + N3 group could significantly antagonize the above effect of N3. Thus, NCS-356 has protective effects against ischemia-reperfusion brain injury by activating GHBR. The neuroprotective effect of GHBR is related with decreasing the content of [Ca2+]i, NO, cGMP and tNOS, iNOS activity in MCAO rats.

Topics: Animals; Benzocycloheptenes; Calcium; Cerebral Cortex; Cerebral Infarction; Cyclic GMP; Infarction, Middle Cerebral Artery; Male; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Random Allocation; Rats; Rats, Sprague-Dawley; Receptors, Cell Surface; Reperfusion Injury

Lung transplantation is severely limited by an inadequate supply of lungs from brain-dead donors. A potential solution is use of lungs from non-heart-beating donors (NHBDs) with retrieval at intervals after circulatory arrest and death. A warm ischemic period with concomitant reperfusion injury is a major limiting factor in the transplantation of lungs retrieved from NHBDs. We hypothesized that the administration of the nitric oxide-donor nitroglycerin to lungs from NHBDs would reduce ischemia-reperfusion injury by activation of guanylate cyclase to form guanosine 3',5'-cyclic monophosphate (cGMP).. An in situ isolated perfused rat lung model was used. Lungs were retrieved from rats at varying intervals after circulatory arrest and death. Lungs were either ventilated with O(2) in situ or not ventilated. Lungs were reperfused at intervals after death with Earle's solution with or without nitroglycerin (0.1 mg/ml). Lung ischemia-reperfusion injury was assessed by capillary filtration coefficient, wet-to-dry lung weight ratio, and pulmonary hemodynamics. Tissue levels of adenine nucleotides and cGMP concentrations were measured by high-performance liquid chromatography and enzyme immunoassay, respectively.. Reperfusion with nitroglycerin decreased capillary filtration coefficient compared with reperfusion without nitroglycerin at all post-mortem ischemic times, irrespective of pre-harvest ventilation. cGMP levels increased significantly with nitroglycerin-reperfusion and attenuated decreases in high-energy adenine nucleotides.. Reperfusion of lungs with nitroglycerin may facilitate safe lung transplantation from NHBDs by reducing capillary leak after reperfusion.

Topics: Animals; Chromatography, High Pressure Liquid; Cyclic GMP; Death; Enzyme Activation; Guanylate Cyclase; Hemodynamics; Lung; Lung Transplantation; Models, Animal; Nitric Oxide; Nitric Oxide Donors; Nitroglycerin; Rats; Reperfusion Injury; Respiration, Artificial; Tissue Donors

Topics: Animals; Antineoplastic Agents; Carbon Monoxide; Cisplatin; Cyclic GMP; Heme Oxygenase-1; Kidney Diseases; Organometallic Compounds; Rats; Reperfusion Injury

A consequence of ischemia/reperfusion (IR) is endothelial barrier dysfunction and intravascular volume loss. The purposes of our study are to explore the impact of: 1) cyclic guanosine monophosphate (cGMP) synthesis inhibition, 2) cyclic adenosine monophosphate (cAMP) synthesis inhibition, 3) treatment with endothelin-1, and 4) endothelin-1 (ET-1)-mediated cAMP changes on IR-induced fluid leak. We hypothesize that IR-mediated microvascular fluid leak results from increased cGMP activity and ET-1 decreases IR-induced fluid leak via cAMP.. A micro-cannulation technique was used to determine fluid leak or hydraulic permeability (Lp) in rat mesenteric venules. Lp was measured during IR and after treatment with 1) cGMP synthesis inhibitor (LY83583,10 micromol/L) 2) cAMP synthesis inhibitor (2',5'dideoxyadenosine,10 micromol/L), 3) ET-1 (80 pM), and 4) cAMP synthesis inhibitor plus ET-1 (n=6 in each group; Lp represented as mean+/-standard error of the mean; units 10-cm/sec/cmH2O).. IR resulted in an increase in Lp (Lp=7.07+/-0.20) sevenfold above baseline (1.05+/-0.31) (p

Topics: Animals; Capillary Leak Syndrome; Capillary Permeability; Cyclic AMP; Cyclic GMP; Endothelin-1; Female; Mesenteric Veins; Microcirculation; Rats; Rats, Sprague-Dawley; Reperfusion Injury

If lungs could be retrieved from non-heart-beating donors, the critical shortage of lungs for transplantation could be alleviated. However, lungs subjected to warm ischemia develop edema when reperfused. We hypothesized that ventilation of rat lungs from non-heart-beating donors with nitric oxide during the period of warm ischemia alone, with reperfusion, or both might reduce ischemia-reperfusion injury.. An isolated perfused rat lung model measured the filtration coefficient and accumulation of lung water by the wet/dry weight ratio. Donor rats were euthanized, and then lungs were retrieved immediately after death or 2 or 3 hours postmortem. Lungs retrieved postmortem were either not ventilated or ventilated with 100% oxygen alone or 40 ppm nitric oxide in oxygen. In the circuit, lungs were ventilated with alveolar gas with or without 40 ppm nitric oxide.. Nitric oxide administration to the non-heart-beating donor or in the perfusion circuit reduced filtration coefficient and wet/dry weight ratio. Lungs retrieved 2 hours postmortem ventilated with nitric oxide or treated with nitric oxide on reperfusion had filtration coefficients and wet/dry weight ratios similar to those of lungs retrieved immediately after death. Nitric oxide was most beneficial when administered both during warm ischemia and at reperfusion in lungs retrieved 3 hours postmortem. Nitric oxide administration in the circuit was associated with increased lung levels of lung cyclic guanosine monophosphate, determined by enzyme-linked immunosorbent assay.. Administration of nitric oxide to non-heart-beating donors during warm ischemia and with reperfusion might facilitate transplantation of lungs from non-heart-beating donors by reducing ischemia-reperfusion injury and capillary leak.

Topics: Adenine Nucleotides; Animals; Chromatography, High Pressure Liquid; Cyclic GMP; Enzyme-Linked Immunosorbent Assay; In Vitro Techniques; Lung Transplantation; Male; Nitric Oxide; Pulmonary Artery; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Vascular Resistance

Previous studies have shown that endothelial nitric oxide (NO) synthase (eNOS)-derived NO is an important signaling molecule in ischemia-reperfusion (I-R) injury. Deficiency of eNOS-derived NO has been shown to exacerbate injury in hepatic and myocardial models of I-R. We hypothesized that transgenic overexpression of eNOS (eNOS-TG) would reduce hepatic I-R injury. We subjected two strains of eNOS-TG mice to 45 min of hepatic ischemia and 5 h of reperfusion. Both strains were protected from hepatic I-R injury compared with wild-type littermates. Because the mechanism for this protection is still unclear, additional studies were performed by using inhibitors and activators of both soluble guanylyl cyclase (sGC) and heme oxygenase-1 (HO-1) enzymes. Blocking sGC with 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) and HO-1 with zinc (III) deuteroporphyrin IX-2,4-bisethyleneglycol (ZnDPBG) in wild-type mice increased hepatic I-R injury, whereas pharmacologically activating these enzymes significantly attenuated I-R injury in wild-type mice. Interestingly, ODQ abolished the protective effects of eNOS overexpression, whereas ZnDPBG had no effect. These results suggest that hepatic protection in eNOS-TG mice may be mediated in part by NO signaling via the sGC-cGMP pathway and is independent of HO-1 signal transduction pathways.

Topics: Animals; Blood Pressure; Cyclic GMP; Deuteroporphyrins; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Guanylate Cyclase; Heart Rate; Heme Oxygenase-1; Liver; Mice; Mice, Transgenic; Nitric Oxide; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Oxadiazoles; Quinoxalines; Receptors, Cytoplasmic and Nuclear; Reperfusion Injury; Severity of Illness Index; Signal Transduction; Soluble Guanylyl Cyclase

Phosphodiesterase-3 inhibition has been shown to attenuate hepatic warm ischemia-reperfusion injury. The aim of this study was to investigate the effect of milrinone, phosphodiesterase-3 inhibitor, on post-ischemic microcirculation of rat livers by intravital microscopy.. Male Wistar rats were randomly assigned to three groups; group A, milrinone pre-treatment; group B, ischemic pre-conditioning; and group C, no pre-treatment. All animals underwent a 60-min warm ischemia of the left lateral liver lobe. Microvascular perfusion and leukocyte-endothelial interaction were observed by intravital videomicroscopy. Hepatocellular viability and cellular damage were quantified by adenosine triphosphate tissue concentration as well as alanine aminotransferase and lactate dehydrogenase blood levels, respectively.. In groups A and B, cyclic AMP hepatic tissue concentration was elevated significantly. After reperfusion, microvascular perfusion in hepatic sinusoids was significantly better maintained, and the number of adherent leukocytes was reduced in sinusoids and in post-sinusoidal venules in these rats. Serum transaminase blood levels were suppressed significantly in these groups compared with controls.. The demonstrated improvement of hepatic microcirculation is certainly derived from milrinone induced cell protection in ischemia reperfusion of the liver. This effect is outlined by improved energy status and reduced liver enzyme liberation and mimics the effect of ischemic pre-conditioning.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Blood Pressure; Cell Communication; Cell Survival; Cyclic AMP; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 3; Endothelial Cells; Heart Rate; Hematocrit; Ischemic Preconditioning; Leukocytes; Liver; Liver Circulation; Liver Transplantation; Male; Microcirculation; Microscopy, Video; Milrinone; Nitric Oxide; Phosphodiesterase Inhibitors; Rats; Rats, Wistar; Reperfusion Injury; Venules

We investigated the effects of nitric oxide (NO) on hepatocellular killing after simulated ischemia/reperfusion and characterized signaling factors triggering cytoprotection by NO. Cultured rat hepatocytes were incubated in anoxic Krebs-Ringer-HEPES buffer at pH 6.2 for 4 hours and reoxygenated at pH 7.4 for 2 hours. During reoxygenation, some hepatocytes were exposed to combinations of NO donors (S-nitroso-N-acetylpenicillamine [SNAP] and others), a cGMP analogue (8-bromoguanosine-3,5-cGMP [8-Br-cGMP]), and a cGMP-dependent protein kinase inhibitor (KT5823). Cell viability was determined by way of propidium iodide fluorometry. Inner membrane permeabilization and mitochondrial depolarization were monitored by confocal microscopy. SNAP, but not oxidized SNAP, increased cGMP during reperfusion and decreased cell killing. Other NO donors and 8-Br-cGMP also prevented cell killing. Both guanylyl cyclase and cGMP-dependent kinase inhibition blocked the cytoprotection of NO. However, 5-hydroxydecanoate and diazoxide- mitochondrial K(ATP) channel modulators-did not affect NO-dependent cytoprotection or reperfusion injury. During reoxygenation, confocal microscopy showed mitochondrial repolarization, followed by depolarization, inner membrane permeabilization, and cell death. In the presence of either SNAP or 8-Br-cGMP, mitochondrial repolarization was sustained after reperfusion preventing inner membrane permeabilization and cell death. In isolated rat liver mitochondria, a cGMP analogue in the presence of a cytosolic extract and adenosine triphosphate blocked the Ca(2+)-induced mitochondrial permeability transition (MPT), an effect that was reversed by KT5823. In conclusion, NO prevents MPT-dependent necrotic killing of ischemic hepatocytes after reperfusion through a guanylyl cyclase and cGMP-dependent kinase signaling pathway, events that may represent the target of NO cytoprotection in preconditioning.

Topics: Adenosine Triphosphate; Animals; Cells, Cultured; Cyclic GMP; Guanylate Cyclase; Hepatocytes; Mitochondria, Liver; Nitric Oxide; Nitric Oxide Donors; Permeability; Rats; Reperfusion Injury; S-Nitroso-N-Acetylpenicillamine

Carbon monoxide (CO) is postulated to protect tissues against several types of injuries. We investigated the role of CO in amelioration of cardiac ischemia-reperfusion injury in vivo and the mechanisms involved in it.. Rats inhaled CO (250 ppm, 500 ppm, or 1000 ppm) for 24 hours in a chamber after myocardial ischemia-reperfusion induced by occluding the left anterior descending coronary artery for 30 minutes. Pre-exposure to 1000 ppm of CO significantly reduced the ratio of infarct areas to risk areas and suppressed the migration of macrophages and monocytes into infarct areas, and the expression of tumor necrosis factor (TNF)-alpha in the heart; however, 250 ppm, 500 ppm of CO, or low barometric pressure hypoxia (0.5 atm) did not affect them. Exposure to 1000 ppm CO resulted in the activation of p38 mitogen-activated protein kinase (p38MAPK), protein kinase Balpha(Akt), endothelial nitric oxide synthase (eNOS), and cyclic guanosine monophosphate (cGMP) in the myocardium. Inhibition of p38MAPK, PI3kinase, NO, and soluble guanylate cyclase with SB203580, wortmannin, N(G)-nitro-L-arginine methyl ester (L-NAME), and methylene blue, respectively, attenuated the cytoprotection by CO.. CO has beneficial effects on cardiac ischemia-reperfusion injury; this effect is mediated by p38MAPK pathway and Akt-eNOS pathway, including production of cGMP.

Topics: Animals; Carbon Monoxide; Cyclic GMP; Hemoglobins; Inhalation; Mitogen-Activated Protein Kinases; Myocardial Ischemia; Myocardium; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Reperfusion Injury; Tumor Necrosis Factor-alpha

Pretreatment with atrial natriuretic peptide (ANP) attenuates ischemia-reperfusion injury of livers via cGMP. Heme oxygenase-1 (HO-1) is known as a protective mediator in ischemia-reperfusion injury. The aim of this study was to investigate whether ANP affects the expression of HO-1.. Rat livers were perfused with KH-buffer with/without ANP or 8-Br-cGMP, kept in UW solution (4 degrees C, 24 h), and reperfused. HO-1 mRNA and protein was determined by Northern and Western blot, in situ hybridization, and immunohistochemistry in livers or isolated liver cells.. ANP significantly elevated HO-1 mRNA expression at the end of the preconditioning period and was without effects at the end of ischemia and during reperfusion. 8-Br-cGMP did not affect HO-1 mRNA expression. In situ hybridization as well as immunohistological double-staining revealed that Kupffer cells but not hepatocytes showed HO-1 mRNA and protein expression. Hepatocytes revealed no changes in HO-1 protein whereas Kupffer cells showed a marked increase in HO-1 protein after ANP treatment. Inhibition of HO-1 did not abrogate hepatoprotection conveyed by ANP.. Our data show the potency of ANP to specifically induce HO-1 in Kupffer cells independently of cGMP. This increased expression of HO-1 is not involved in hepatoprotection conferred by ANP being in line with the knowledge that ANP mediates hepatoprotection via cGMP.

Topics: Animals; Atrial Natriuretic Factor; Cyclic GMP; Gene Expression Regulation, Enzymologic; Guanylate Cyclase; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Ischemic Preconditioning; Kupffer Cells; Rats; Receptors, Atrial Natriuretic Factor; Reperfusion Injury; RNA, Messenger

Many studies in diverse models suggest that nitric oxide (NO) may be protective against liver injury due to ischaemia-reperfusion (IR). We evaluated, in an experimental in vivo model of rat liver partial ischaemia, the effects of pretreatment by an NO donor (spermineNONOate, 5mg/kg), and exogenous cGMP (8Br-cGMP, 16 mg/kg) or an endogenous cGMP producer (ANP, 10 microg/kg), to assess their beneficial effects. After 6h of reperfusion, 8Br-cGMP completely prevented the adverse effect of Nomega-nitro-L-arginine (10mg/kg) and 8Br-cGMP alone showed a protective action on both hepatocytes (AST, -25%, LDH, -55%) and endothelial cells (plasma hyaluronic acid (HA), -30%). ANP caused a marked decrease in AST and LDH activities only after 1h of reperfusion (AST, -30%, LDH, -40%). Pretreatment with spermineNONOate prevented hepatocyte injury after 1 and 6h of reperfusion (AST, -22%, LDH, -27%). However, neither spermineNONOate nor ANP had any protective effect on endothelial cell damage. These results confirm the beneficial effect of an NO donor and strongly suggest the implication of a cGMP pathway that does not involve a blockade of inflammatory cytokines production (IL-6 generation was unaffected by 8Br-cGMP pre-treatment). In our model, 8Br-cGMP showed a greater protective effect than ANP or spermineNONOate and so might be used to prevent hepatic injury after IR. Finally, we propose a schematic representation of the different routes for the actions of NO in protecting the liver against IR damage.

Topics: Animals; Aspartate Aminotransferases; Atrial Natriuretic Factor; Cyclic GMP; Enzyme Inhibitors; Hyaluronic Acid; L-Lactate Dehydrogenase; Liver; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Donors; Nitrogen Oxides; Random Allocation; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Spermine

The stress-responsive mitogen-activated protein kinases (MAPKs) (p38-MAPK, c-Jun NH2-terminal kinase [JNK-1 and JNK-2], and extracellular signal regulated kinases [ERK-1 and ERK-2]) might be involved in angiotensin II (AII)-induced ischemia-reperfusion injury. Cardioprotection induced by AII type 1 (AT1) and type 2 (AT2) receptor blockade during ischemia-reperfusion is associated with protein kinase Cepsilon (PKCepsilon), nitric oxide, and cyclic guanosine monophosphate (cGMP) signaling. Our aim was to assess the effect of selective AT1 and AT2 receptor blockade with losartan and PD123,319, respectively, on MAPK expression after ischemia-reperfusion in isolated working rat hearts.. Groups of six hearts were subjected to global ischemia (30 minutes) followed by reperfusion (30 minutes) and exposed to no drug/no ischemia-reperfusion (control), ischemia-reperfusion/no drug, and ischemia-reperfusion with losartan (1 microM), or PD123,319 (0.3 microM) and additional groups. AT1/AT2 receptor expression, MAPKs, PKCepsilon, and cGMP, and changes in mechanical function were measured. Western blotting was done on left ventricular tissue for AT1/AT2, p38/phosphorylated-p38 (p-p38), phosphorylated (p)-JNK-1/-2, phosphorylated (p)-ERK-1/-2, and PKCepsilon proteins; Northern blots for AT1/AT2 mRNA; and enzyme immunoassay for cGMP.. Compared with controls, ischemia-reperfusion induced significant left ventricular dysfunction, decreased AT2 protein and mRNA, increased p-p38 and p-JNK-1/-2, did not change p-ERK-1/-2 or PKCepsilon, and decreased cGMP. PD123,319 improved left ventricular recovery after ischemia-reperfusion, increased AT2 protein and mRNA, mildly increased p-p38, normalized p-JNK-1, did not change p-ERK-1/-2, and increased PKCepsilon and cGMP. Losartan did not change p-p38, increased p-JNK-1, and did not change pERK-1/-2, PKCepsilon, or cGMP.. The overall results suggest that the activation of p38-MAPK and JNK might be linked to AII signaling and play a significant role in acute ischemia-reperfusion injury as well as in the cardioprotective effect of AT2 receptor blockade.

Topics: Angiotensin II Type 2 Receptor Blockers; Animals; Antihypertensive Agents; Blood Pressure; Cardiac Output; Coronary Circulation; Cyclic GMP; Disease Models, Animal; Enalaprilat; Enzyme Activation; Heart; Losartan; Male; Mitogen-Activated Protein Kinases; Models, Cardiovascular; Phosphorylation; Protein Kinase C; Protein Kinase C-epsilon; Rats; Rats, Sprague-Dawley; Reperfusion Injury; RNA, Messenger

To characterize the effects of ischemia on cGMP synthesis in microvascular endothelium, cultured endothelial cells from adult rat hearts were exposed to hypoxia or normoxia at pH 6.4 or 7.4. Cellular cGMP and soluble (sGC) and membrane guanylyl cyclase (mGC) activities were measured after stimulation of sGC (S-nitroso-N-acetyl-penicillamine) or mGC (urodilatin) or after no stimulation. Cell death (lactate dehydrogenase release) was negligible in all experiments. Hypoxia at pH 6.4 induced a rapid approximately 90% decrease in cellular cGMP after sGC and mGC stimulation. This effect was reproduced by acidosis. Hypoxia at pH 7.4 elicited a less pronounced (approximately 50%) and slower reduction in cGMP synthesis. Reoxygenation after 2 h of hypoxia at either pH 6.4 or 7.4 normalized the response to mGC stimulation but further deteriorated the sGC response; normalization of pH rapidly reversed the effects of acidosis. At pH 7.4, the response to GC stimulation correlated well with cellular ATP. We conclude that simulated ischemia severely depresses cGMP synthesis in microvascular coronary endothelial cells through ATP depletion and acidosis without intrinsic protein alteration.

Topics: Acidosis; Adenosine Triphosphate; Animals; Atrial Natriuretic Factor; Coronary Vessels; Cyclic GMP; Endothelium, Vascular; Energy Metabolism; Enzyme Activation; Guanylate Cyclase; Hydrogen-Ion Concentration; Hypoxia; Male; Microcirculation; Nitric Oxide; Oxidative Stress; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Ischemic preconditioning (IPC) may protect the liver from ischemia reperfusion injury by nitric oxide formation. This study has investigated the effect of ischemic preconditioning on hepatic microcirculation (HM), and the relationship between nitric oxide metabolism and HM in preconditioning. Rats were allocated to 5 groups: 1. sham laparotomy; 2. 45 minutes lobar ischemia followed by 2-hour reperfusion (IR); 3. IPC with 5 minutes ischemia and 10 minutes reperfusion before IR; 4. L-arginine before IR; and 5. L-NAME + IPC before IR. HM was monitored by laser Doppler flowmeter. Liver transaminases, adenosine triphosphate, nitrites + nitrates, and guanosine 3'5'-cyclic monophosphate (cGMP) were measured. Nitric oxide synthase (NOS) distribution was studied using nicotinamide adeninine dinucleotide phosphate (NADPH) diaphorase histochemistry. At the end of reperfusion phase, in the IR group, flow in the HM recovered partially to 25.8% of baseline (P < .05 versus sham), whereas IPC improved HM to 49.5% of baseline (P < .01 versus IR). With L-arginine treatment, HM was 31.6% of baseline (NS versus IR), showing no attenuation of liver injury. In the preconditioned group treated with L-NAME, HM declined to 10.2% of baseline, suggesting not only a blockade of the preconditioning effect, but also an exacerbated liver injury. Hepatocellular injury was reduced by IPC, and L-arginine and was increased by NO inhibition with L-NAME. IPC also increased nitrate + nitrate (NOx) and cGMP concentrations. NOS detected by NADPH diaphorase staining was associated with hepatocytes and vascular endothelium, and was induced by IPC. IPC induced NOS and attenuated HM impairment and hepatocellular injury. These data strongly suggest a role for nitric oxide in IPC.

Topics: Adenosine Triphosphate; Animals; Cyclic GMP; Dihydrolipoamide Dehydrogenase; Hemodynamics; Ischemic Preconditioning; Liver; Liver Circulation; Microcirculation; Models, Animal; Nitric Oxide; Rats; Reperfusion Injury; Time Factors

Preischemic treatment with atrial natriuretic peptide (ANP) attenuates ischemia-reperfusion injury of the rat liver via cyclic guanosine monophosphate (cGMP). The attenuated activation of nuclear factor kappaB (NF-kappaB) seems to contribute to this effect. The aim of this study was to determine whether heat shock proteins are involved in these molecular pathways. Livers of male Sprague-Dawley rats were continuously perfused with Krebs-Henseleit (KH) buffer with or without ANP or 8-Br-cGMP. In different experiments livers were perfused with or without ANP for 20 minutes, kept in cold storage solution for 24 hours, and reperfused. Activation of heat shock transcription factor (HSF) (by electrophoretic mobility shift assay), heat shock protein 70 (HSP70), and glyceraldehyde phosphate dehydrogenase (GAPDH) mRNA (by reverse transcription polymerase chain reaction [RT-PCR]), as well as HSP70 (by Western blot) were investigated in freeze-clamped liver samples. During continuous perfusion ANP as well as 8-Br-cGMP activated HSF, HSP70 protein concentrations paralleled HSF-activation. ANP pretreated livers exhibited elevated HSF after 24 hours of ischemia and elevated HSP70 mRNA levels during reperfusion. ANP prevented the marked decrease of HSP70 protein during reperfusion. Coimmunoprecipitation studies showed increased binding of HSP70 to inhibitory factor kappaB (IkappaB) in ANP-treated livers. In conclusion, we showed the cGMP-mediated activation of HSF by ANP, which resulted in elevated HSP70 mRNA and protein concentrations and correlated with enhanced binding of HSP70 to IkappaB. This could be an important mechanism of ANP-mediated prevention of hepatic preservation damage.

Topics: Adhesins, Bacterial; Animals; Atrial Natriuretic Factor; Cyclic GMP; DNA; Gene Expression Regulation; HSP70 Heat-Shock Proteins; I-kappa B Proteins; Liver; Male; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

A direct role of carbon monoxide (CO), an effector-signaling molecule during heme oxygenase-1 (HO-1) catalysis of heme, in the protection against hepatic ischemia/reperfusion (I/R) injury needs to be established. This study was designed to determine the effects and downstream mechanisms of CO on cold I/R injury in a clinically relevant isolated perfusion rat liver model. After 24 hours of cold storage, rat livers perfused ex vivo for 2 hours with blood supplemented with CO (300 parts per million) showed significantly decreased portal venous resistance and increased bile production, as compared with control livers perfused with blood devoid of CO. These beneficial effects correlated with improved liver function (serum glutamic oxaloacetic transaminase levels) and diminished histological features of hepatocyte injury (Banff's scores). The CO-mediated cytoprotective effects were nitric oxide synthase- and cyclic guanine monophosphate-independent, but p38 mitogen-activated protein kinase (MAPK)-dependent. Moreover, adjunctive use of zinc protoporphyrin, a competitive HO-1 inhibitor, has shown that exogenous CO could fully substitute for endogenous HO-1 in preventing hepatic I/R insult. This study performed in a clinically relevant ex vivo cold ischemia model is the first to provide the evidence that HO-1-mediated cytoprotection against hepatic I/R injury depends on the generation of, and can be substituted by, exogenous CO. The p38 MAPK signaling pathway represents the key downstream mechanism by which CO prevents the I/R insult. In conclusion, regimens that employ exogenous CO should be revisited, as they may have potential applications in preventing/mitigating I/R injury, and thus expanding the liver donor pool for clinical transplantation.

Topics: Animals; Carbon Monoxide; Cold Temperature; Cyclic GMP; Enzyme Activation; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; In Vitro Techniques; Ischemia; Liver; Liver Circulation; Male; Mitogen-Activated Protein Kinases; Nitric Oxide; p38 Mitogen-Activated Protein Kinases; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Nitric oxide (NO) may act as a toxin in several neuropathologies, including the brain damage resulting from cerebral ischaemia. Rat striatal slices were used to determine the mechanism of enhanced NO release following simulated ischaemia and, for estimating the NO concentrations, the activity of guanylyl cyclase served as a biosensor. Exposure of the slices for 10 min to an oxygen- and glucose-free medium caused a 70% fall in cGMP levels. On recovery, cGMP increased 2-fold above basal, where it remained for 40 min before declining. The pattern of changes matched those of cGMP or NO oxidation products measured during and after brain ischaemia in vivo. The increase observed during the recovery period was blocked by inhibition of NO synthase or NMDA receptors and was curtailed by tetrodotoxin, implying that it was caused by glutamate release leading to activation of the NMDA receptor-NO synthase pathway. Calibration of the cGMP levels against NO-stimulated guanylyl cyclase yielded a basal NO concentration of 0.6 nm. The peak NO concentration achieved on recovery from simulated ischaemia was estimated as 0.8 nm. These values are compatible with the low micromolar concentrations of NO oxidation products (chiefly nitrate) found by microdialysis in vivo, providing the NO inactivation rate (forming nitrate) is accounted for. NO at a concentration around 1 nm is unlikely to be toxic to cells. However, if the NO inactivation mechanism were to fail (as it can) the NO production rate normally providing only subnanomolar NO could readily generate toxic (microM) NO concentrations.

Topics: 1-Methyl-3-isobutylxanthine; Animals; Arginine; Biosensing Techniques; Brain Ischemia; Cyclic GMP; Guanylate Cyclase; NADPH Dehydrogenase; Neostriatum; Neurons; Nitric Oxide; Nitric Oxide Synthase; Phosphodiesterase Inhibitors; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury

In a series of canine liver ischemia experiments, we have shown that amelioration of hepatic injury is achievable by the inhibition of vasoconstriction, cytokine production, platelet aggregation, and neutrophil infiltration. Cyclic adenosine diphosphate (cAMP) was considered to be involved in most of these events. In our study, we tested our hypothesis that augmentation of endogenous cAMP by phosphodiesterase (PDE) 3 inhibitor, amrinone (AM), or adenylate cyclase stimulator, NKH477 (NKH), could attenuate ischemia and reperfusion injury of the liver.. Thirty-six beagle dogs were used. They were divided into group CT (untreated control), group AM, group NKH, and group CB (treated by both agents). AM or NKH were administered i.v. 1 hr before ischemia (group preAM and group preNKH) or 15 min before reperfusion (pos-AM and postNKH). Combination group animals were treated only before ischemia. Animal survival, hepatic tissue blood flow, liver enzymes, platelet counts, energy metabolism, hepatic cAMP and cyclic guanosine 3',5'-cyclic monophosphate levels, and histopathology were analyzed.. Two-week animal survival was significantly improved by pre- or posttreatment with either agent. After reperfusion, hepatic tissue blood flow, liver enzyme release, platelet counts, energy metabolism, tissue cAMP levels, and histological architecture were also ameliorated markedly. Combination of both agents induced severe liver damage and lethal hypotension. AM treatment exhibited more protective effects than NKH, particularly when it was given before ischemia. Interestingly, not only cyclic guanosine 3',5'-cyclic monophosphate, were also restored at higher levels after reperfusion by preischemia treatment.. Administration of amrinone or NKH477 maintained hepatic tissue concentrations of cyclic nucleotides, and attenuated ischemia and reperfusion injury of the liver. Thus, regulation of hepatic tissue cyclic nucleotides is an important alternative for prevention of hepatic damage in liver preservation and surgery.

Topics: Adenosine Triphosphate; Amrinone; Animals; Colforsin; Cyclic AMP; Cyclic GMP; Dogs; Energy Metabolism; Female; Hemodynamics; Ischemia; Liver; Liver Circulation; Platelet Count; Portal Vein; Reperfusion Injury

Ischemia is an inciting factor in 50% of incidences of acute renal failure, and it increases the risk of organ rejection after renal transplantation. We have previously demonstrated that resveratrol (RSV) reduces ischemia-reperfusion (I/R) injury of rat kidney both by antioxidant and anti-inflammatory mechanisms. However, a clear morphological demonstration of this activity has not been made. To answer this question we have performed a new set of experiments following the experimental protocol reported below to investigate the effects of I/R injury and RSV pretreatment on kidney morphology by computerized morphometric analysis. Both renal arteries were clamped for 40 minutes in 40 male Wistar rats (b.w. 220 +/- 20 g); 20 rats were pretreated with RSV 1 microM e.v. 40 minutes before clamping. All animals were reperfused for 24 hours and then sacrificed. Histological examination showed tissue conservation in treated rats. I/R-induced glomerular collapse (as revealed by mean glomerular volume and glomerular shape factor) was significantly reduced by RSV pretreatment. Capillary tuft/Bowman's capsule area ratio was enhanced in the I/R group suggesting tubular hypertension. RSV pre-treatments significantly reduced this parameter to the control value. The number of platelet clots in the capillary tuft and tubular necrosis were also reduced by RSV versus I/R group. L-NAME administration worsened both functional and structural damage. Finally, cGMP urinary levels were markedly reduced from 12.1 +/- 8.4 nmol/day to 0.10 +/- 0.10 nmol/day in the I/R group. RSV provided cGMP (5.01 +/- 1.5 nmol/day, P < 0.05). As expected, L-NAME administration significantly reduced cGMP in urine (0.71 +/- 0.6 nmol/day). The present study confirms the protective effect of RSV pretreatment in I/R injury of rat kidney and suggests multiple mechanisms of action.

Topics: Animals; Cyclic GMP; Dose-Response Relationship, Drug; Ischemia; Kidney Diseases; Kidney Glomerulus; Male; Rats; Rats, Wistar; Renal Circulation; Reperfusion Injury; Resveratrol; Stilbenes; Vitis; Wine

The present study aims at investigating the role of nitric oxide (NO) on the oxidative damage in gastric mucosa of rats which received ischemia/reperfusion (I/R) and its relation to mucus. NO synthesis modulators such as L-arginine and N(G)-nitro-L-arginine methyl ester (L-NAME) were injected intraperitoneally to the rats 30 min prior to I/R which was induced by clamping the celiac artery and the superior mesenteric artery for 30 min and reperfusion for 1 h. As a result, I/R increased lipid peroxide production and decreased the contents of glutathione (GSH), cGMP and mucus as well as GSH peroxidase activities of gastric mucosa. I/R decreased the activity and protein of NO synthase (NOS) in gastric mucosa. Pretreatment of L-arginine, a substrate for NOS, prevented I/R-induced alterations of gastric mucosa. However, L-NAME, an NOS inhibitor, deteriorated oxidative damage induced by I/R. In conclusion, NO has an antioxidant defensive role on gastric mucosa by maintaining mucus, GSH and GSH peroxidase, which were related to preservation of cGMP and NOS in gastric mucosa.

Topics: Animals; Arginine; Cyclic GMP; Gastric Mucosa; Glutathione; Glutathione Peroxidase; Lipid Peroxides; Male; Mucus; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Sprague-Dawley; Reperfusion Injury

We examined the effect of 3-isobutyryl-2-isopropylpyrazolo[1,5-a]pyridine (ibudilast), which has been clinically used for bronchial asthma and cerebrovascular disorders, on cell viability induced in a model of reperfusion injury. Ibudilast at 10 - 100 microM significantly attenuated the H(2)O(2)-induced decrease in cell viability. Ibudilast inhibited the H(2)O(2)-induced cytochrome c release, caspase-3 activation, DNA ladder formation and nuclear condensation, suggesting its anti-apoptotic effect. Phosphodiesterase inhibitors such as theophylline, pentoxyfylline, vinpocetine, dipyridamole and zaprinast, which increased the guanosine-3',5'-cyclic monophosphate (cyclic GMP) level, and dibutyryl cyclic GMP attenuated the H(2)O(2)-induced injury in astrocytes. Ibudilast increased the cyclic GMP level in astrocytes. The cyclic GMP-dependent protein kinase inhibitor KT5823 blocked the protective effects of ibudilast and dipyridamole on the H(2)O(2)-induced decrease in cell viability, while the cyclic AMP-dependent protein kinase inhibitor KT5720, the cyclic AMP antagonist Rp-cyclic AMPS, the mitogen-activated protein/extracellular signal-regulated kinase inhibitor PD98059 and the leukotriene D(4) antagonist LY 171883 did not. KT5823 also blocked the effect of ibudilast on the H(2)O(2)-induced cytochrome c release and caspase-3-like protease activation. These findings suggest that ibudilast prevents the H(2)O(2)-induced delayed apoptosis of astrocytes via a cyclic GMP, but not cyclic AMP, signalling pathway.

Topics: Alkaloids; Animals; Animals, Newborn; Apoptosis; Astrocytes; Carbazoles; Cell Survival; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Cytochrome c Group; Dipyridamole; DNA; Dose-Response Relationship, Drug; Enzyme Inhibitors; Hydrogen Peroxide; Indoles; Mitochondria; Pentoxifylline; Peptide Hydrolases; Phosphodiesterase Inhibitors; Purinones; Pyridines; Rats; Rats, Wistar; Reperfusion Injury; Signal Transduction; Theophylline; Vinca Alkaloids

Substitution of the nitric oxide- (NO-) pathway improves early graft function following lung transplantation. We previously demonstrated that 8-Br-cGMP (second messenger of NO) to the flush solution and tetrahydrobiopterin (BH4, coenzyme of NO synthase) given as additive during reperfusion improve post-transplant graft function. In the present study, the combined treatment with 8-Br-cGMP and BH4 was evaluated.. Unilateral left lung transplantation was performed in weight matched outbred pigs (24-31 kg). In group I, grafts were preserved for 30 h (n=5). 8-Br-cGMP (1mg/kg) was added to the flush solution (Perfadex, 1.5l, 1 degrees C) and BH4 (10mg/kg/h) was given to the recipient for 5h after reperfusion. In group II, lungs were transplanted after a preservation time of 30 h (n=3) and prostaglandin E(1) (250 g) was given into the pulmonary artery (PA) prior to flush. In all recipients 1h after reperfusion the contralateral right PA and bronchus were ligated to assess graft function only. Survival time after reperfusion, extravascular lung water index (EVLWI), hemodynamic variables, and gas exchange (PaO(2)) were assessed during a 12h observation period.. All recipients in group I survived the 12h assessment, whereas none of the group II animals survived more than 4h after reperfusion with a rapid increase of EVLWI up to 24.8+/-6.7 ml/kg. In contrast, in group I EVLWI reached up to 8.9+/-1.5 ml/kg and returned to nearly normal levels at 12h (6.1+/-0.8 ml/kg). In two animals of group I the gas exchange deteriorated slightly. The other three animals showed normal arterial oxygenation over the entire observation time.. Our data indicate that the combined substitution of the NO pathway during preservation and reperfusion reduces ischemia/reperfusion injury substantially and that this treatment even allows lung transplantation after 30 h preservation in this model.

Topics: Animals; Biopterins; Cell Movement; Coenzymes; Cyclic GMP; Extravascular Lung Water; Graft Survival; Hemodynamics; Infusions, Intravenous; Lipid Peroxidation; Lung; Lung Transplantation; Neutrophils; Nitric Oxide Synthase; Organ Preservation; Organ Preservation Solutions; Peroxidase; Pulmonary Gas Exchange; Reperfusion Injury; Swine; Thiobarbituric Acid Reactive Substances; Time Factors

Pulmonary edema, owing to an impairment of microvascular barrier function, is an important feature in lung ischemia/reperfusion (IR) injury. Inhalation of nitric oxide (NO) during the period of reperfusion has previously been shown to reduce this leakage response.. We investigated the impact of short-term (30 min) low-dose (10 ppm) pre-ischemic NO inhalation on IR injury in buffer-perfused rabbit lungs, subsequently undergoing 210 min of warm, anoxic-ventilated ischemia.. Far-reaching suppression of the leakage response, reflected by manifold increased capillary filtration coefficients and edema formation, was noted in lungs with pre-ischemic NO administration, corresponding to the beneficial effect of NO inhalation during reperfusion. The effect of NO pre-exposure was not related to vasodilation, because microvascular pressures were unchanged, and was mimicked by pre-ischemic intravascular administration of sodium nitroprusside with subsequent washout of this agent. NO inhalation during reperfusion, but not pre-ischemic, short-term NO administration, provoked a manifold increase in the accumulation of guanosine 3',5'-cyclic monophosphate (cGMP) in the perfusate. The cGMP-analogue, 8-Br-cGMP, mimicked the anti-edematous effect of NO when present during reperfusion, but pre-ischemic, short-term administration of 8-Br-cGMP provided only limited protection. The guanylate cyclase-inhibitor, 1H-[1, 2, 4]-Oxadiazolo-[4,3-a]-quinoxalin-1-one (ODQ), largely antagonized the beneficial effects of NO inhalation during reperfusion but had only minor influence on the effect of NO pre-exposure.. "Preconditioning" of the lung vasculature with short-term NO administration maintains endothelial integrity in a subsequent ischemia/reperfusion maneuver, with nonvasodilatory and non-cGMP-related mechanisms suggested to be largely responsible. This finding may offer interesting perspectives for donor management in clinical lung transplantation.

Topics: Administration, Inhalation; Animals; Cyclic GMP; Female; Ischemic Preconditioning; Lung; Male; Nitric Oxide; Pulmonary Artery; Rabbits; Reperfusion Injury; Weight Gain

The aim of this study was to test the hypothesis that agents which stabilize the mast cell membrane may modulate the phenotype of the vascular wall in a lung ischemia-reperfusion model, including altering expression of endothelial and leukocyte adhesion receptors and the inducible nitric oxide synthase (NOS-2).. Three sets of rats were given either intravenous saline (group A), ketotifen (group B), or cromolyn (group C), respectively. The left pulmonary artery was ligated temporarily and reopened after 2 hours of ischemia. Then, after a 2-hour period of reperfusion, the left lung was excised. ICAM-1 and NOS-2 were measured at the protein level by Western blotting, and cGMP levels were measured by enzyme-linked immunosorbent assay in the lung tissue specimens for each drug group.. ICAM-1 expressions, determined as the intensity of a given band on the Western blot, were 197+/-59 in group B and 195+/-83 in group C versus 369+/-114 in group A (p = 0.002 for analysis of variance). In contrast with ICAM-1, NOS-2 expression was increased by ketotifen or cromolyn treatment (464+/-82 in group B and 507+/-93 in group C, compared with 377+/-44 for group A, p = 0.007). The finding of increased NOS-2 expression in groups B and C is consistent with the observed increase in tissue cGMP levels in the same groups (1.92+/-0.9 pmol/mL for group A versus 7.8+/-3.5 pmol/mL for group B, and 12.4+/-5.8 pmol/mL for group C, p = 0.0004).. These data establish that mast cell stabilizing agents modulate the vascular phenotype in the setting of pulmonary ischemia and reperfusion by decreasing ICAM-1 expression, augmenting expression of NOS-2, and increasing tissue cGMP levels. As decreasing ICAM-1 expression and increasing cGMP levels have proven useful to limit proinflammatory mechanisms of tissue injury, mast cell stabilizing agents may provide a new therapeutic option to improve organ function in the setting of reperfusion.

Topics: Analysis of Variance; Animals; Anti-Asthmatic Agents; Cell Membrane; Cromolyn Sodium; Cyclic GMP; Disease Models, Animal; Endothelium, Vascular; Histamine H1 Antagonists; Intercellular Adhesion Molecule-1; Ischemia; Ketotifen; Lung; Male; Mast Cells; Nitric Oxide Synthase; Phenotype; Rats; Rats, Sprague-Dawley; Reperfusion Injury

Substitution of the NO-pathway reduces ischemia/reperfusion injury following lung transplantation. 8-Br-cGMP is a membrane permeable analogue of cGMP, the second messenger of NO. In this study the effect of continuous administration of 8-Br-cGMP on early graft function was evaluated.. Unilateral left lung transplantation was performed in 10 weight-matched pigs (23-30 kg). Donor lungs were flushed with 1.51 cold (1 degree C) LPD solution and preserved for 20 hours. In Group I (n = 5), 8-Br-cGMP (0.2 mg/kg/h) was given continuously over the entire observation time starting 15 min before reperfusion. Group II served as control, no 8-Br-cGMP was administered. In both groups, 250 microg PGE1 was injected into the pulmonary artery (PA) before flush. One hour after reperfusion the recipients contralateral right PA and bronchus were ligated to assess isolated graft function only. Extravascular lung water index (EVLWI), pulmonary vascular resistance, mean PA pressure, mean systemic arterial pressure and gas exchange were assessed during a 5-hour observation period. Lipid peroxidation as indicator for free radical mediated injury and neutrophil migration to the allograft were measured at the end of the assessment.. EVLWI was significantly reduced in animals treated with 8-Br-cGMP (overall difference P = 0.024) with a peak 2 hours after reperfusion (Group I, 8.2+/-0.3 mg/ml vs Group II, 10.1+/-0.6 mg/ml; P = 0.039). Also in Group I the free radical mediated tissue injury was significantly lower when compared to Group II (Group I, 61.8+/-12.3 pmol/g vs Group II, 120.7+/-7.2 pmol/g; P = 0.006). A tendency towards a reduced neutrophil migration after 8-Br-cGMP infusion was shown; however, the changes in comparison to the control animals were not statistically significant (Group I, 1.0+/-0.2 deltaOD/mg/min vs Group II, 1.7+/-0.3 deltaOD/mg/min; P = 0.13). Pulmonary- and systemic hemodynamics, and allograft gas exchange did not differ between groups.. The results indicate that substitution of the NO pathway by administration of the second messenger cGMP at the time of reperfusion improves post-transplant lung allograft function.

Topics: Animals; Cyclic GMP; Cyclic GMP-Dependent Protein Kinases; Extravascular Lung Water; Free Radicals; Lipid Peroxidation; Lung Transplantation; Postoperative Period; Pulmonary Edema; Random Allocation; Reperfusion Injury; Swine; Thiobarbituric Acid Reactive Substances; Transplantation, Homologous

Prevention of ischemia-reperfusion (IR) injury is crucial for successful lung transplantation. We investigated whether a nitric oxide donor, nitroglycerin (NTG), could suppress the oxidative stress of IR injury and improve pulmonary function after reperfusion in an ex vivo rat lung perfusion model. In Fresh group of animals, the lungs were flushed with perfusate, followed immediately by reperfusion, and no lung injury was observed. In NTG- and NTG+ groups of animals, the lungs were flushed with perfusate alone or perfusate containing NTG, respectively. Harvested lung and heart blocks from these latter two groups were immersed in the corresponding perfusate at 4 degrees C for 15 h, and were then reperfused for 60 min. Reperfusion induced pulmonary edema in the NTG- group, but not in the NTG+ group. Shunt fractions in NTG+ group were significantly lower than in the NTG- group throughout reperfusion. NTG had no effect on pulmonary arterial pressure or myeloperoxidase activity. In contrast, oxidative DNA damage assessed immunohistochemically with a monoclonal antibody against 8-hydroxy-2'-deoxyguanosine (8-OHdG) was significantly increased in the NTG- group, in the order alveolar epithelium > pulmonary endothelium > bronchial epithelium. NTG treatment significantly decreased staining with the anti-8-OHdG antibody in all three areas of tissue. Therefore, administration of NTG attenuates the oxidative stress of IR injury, and may improve pulmonary function after reperfusion.

Topics: Animals; Cyclic GMP; DNA Damage; Immunoenzyme Techniques; Lung; Lung Transplantation; Male; Nitroglycerin; Organ Preservation; Oxidative Stress; Peroxidase; Pulmonary Alveoli; Pulmonary Edema; Rats; Rats, Inbred Lew; Reperfusion Injury; Vasodilator Agents

Disruption of endothelial barrier properties with development of noncardiogenic pulmonary edema is a major threat in lung ischemia-reperfusion (I/R) injury that occurs under conditions of lung transplantation. Inhaled nitric oxide (NO) reduced vascular leakage in lung I/R models, but the efficacy of this agent may be limited. We coadministered NO and zaprinast, a cGMP-specific phosphodiesterase inhibitor, to further augment the NO-cGMP axis. Isolated, buffer-perfused rabbit lungs were exposed to 4.5 h of warm ischemia. Reperfusion provoked a transient elevation in pulmonary arterial pressure and a negligible rise in microvascular pressure followed by a massive increase in the capillary filtration coefficient and severe lung edema formation. Inhalation of 10 parts/million of NO or intravascular application of 100 microM zaprinast on reperfusion both reduced pressor response and moderately attenuated vascular leakage. Combined administration of both agents induced no additional vasodilation at constant microvascular pressures, but additively protected against capillary leakage paralleled by a severalfold increase in perfusate cGMP levels. In conclusion, combining low-dose NO inhalation and phosphodiesterase inhibition may be suitable for the maintenance of graft function in lung transplantation by amplifying the beneficial effect of the NO-cGMP axis and avoiding toxic effects of high NO doses.

Topics: Animals; Blood Pressure; Capillary Permeability; Compliance; Cyclic GMP; Female; In Vitro Techniques; Ischemia; Male; Nitric Oxide; Organ Size; Phosphodiesterase Inhibitors; Pulmonary Artery; Pulmonary Circulation; Purinones; Rabbits; Reperfusion Injury

Alterations in nitric oxide synthesis, endothelial adhesivity and pulmonary hemodynamics are investigated in an animal model of lung ischemia-reperfusion.. Two sets of rats, each containing seven animals, were either subjected to unilateral pulmonary ischemia and reperfusion (Study Group) or underwent the same surgical procedure without ischemia (Control Group). Pulmonary artery pressure (PAP), pulmonary blood flow (PBF) trend, NOS-2, intercellular adhesion molecule-1 (ICAM-1), myeloperoxidase (MPO) and cGMP expression of the reperfused lung tissue and, final paO(2) were compared between the two groups.. ICAM-1 expression was increased (369+/-114 vs. 115+/-65; P=0.02), NOS-2 expression and tissue cGMP levels were decreased (377+/-44 vs. 452+/-54; P=0.03 and 7.8+/-3.5 vs. 9.4+/-2.3 pmol/ml; P=0.03, respectively) and MPO activity was increased (2.7+/-0.9-3.5+/-0.8; P=0.03) in the reperfused lungs. Pulmonary artery pressure was 15+/-7 mmHg in the Control Group vs. 22+/-16 mmHg in the Study Group (P=0.04) at the 30th min of reperfusion. Pulmonary blood flow was greater in the Study Group at the beginning of reperfusion (9.5+/-4.1 vs. 7.1+/-3.1 ml/min at the 30th min) but considerably reduced thereafter (3.2+/-1. 4 vs. 6.2+/-2.1 at the 60th minute and 2.9+/-1.6 vs. 5.8+/-1.9 at the 120th min). At the end of the experiment, paO(2) was 95+/-30 in the Control Group vs. 71+/-32 in the Study Group (P=0.03).. These data establish that nitric oxide synthesis was suppressed after reperfusion. Pulmonary blood flow was first increased and then reduced. A parallel increase in MPO and ICAM-1 indicated proinflammatory reaction. Decreased tissue cGMP level was consistent with the suppressed NOS-2 production. Organ function was negatively influenced as represented by the decreased oxygenation, probably due to no-reflow phenomenon.

Topics: Animals; Biomarkers; Blood Flow Velocity; Blotting, Western; Cell Adhesion; Cyclic GMP; Endothelium, Vascular; Intercellular Adhesion Molecule-1; Leukocytes; Lung; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Peroxidase; Pulmonary Circulation; Pulmonary Wedge Pressure; Rats; Rats, Sprague-Dawley; Reperfusion Injury

To observe the role of NMDA receptor-Nitric Oxide(NOS)-cGMP pathway in ischemic brain injury.. The common cervical arteries of the rats were transient blocked bilaterally, in association with bleeding from their tails and followed by reperfusion. The procedures were repeated once again as above to establish a stable ischemic brain injury model. 3H-MK801 binding, cNOS activity, iNOS activity, and the cGMP content were measured at different time intervals after ischemic injury.. The results indicated that the change of 3H-MK801 binding varied among the cerebral cortex and hippocampus. cNOS activity began to rise in all parts of the brain 24 hours after operation and reached its peak in 3 days. The regions where iNOS and cNOS activity and cGMP content all increased significantly include the hippocamus, striatum and cortex, especially in hippocampus.. The tendency and degree of increase of those four indixes were cosistent during ischemic injury, indicating an important role of NMDA receptor-NO-cGMP in ischemic injury of the hippocampus.

Topics: Animals; Brain; Brain Ischemia; Cyclic GMP; Female; Hippocampus; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type II; Rats; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury

The generation of reactive oxygen species (ROS) by activated Kupffer cells contributes to liver injury following liver preservation, shock, or endotoxemia. Pharmacological interventions to protect liver cells against this inflammatory response of Kupffer cells have not yet been established. Atrial natriuretic peptide (ANP) protects the liver against ischemia-reperfusion injury, suggesting a possible modulation of Kupffer cell-mediated cytotoxicity. Therefore, we investigated the mechanism of cytoprotection by ANP during Kupffer cell activation in perfused rat livers of male Sprague-Dawley rats. Activation of Kupffer cells by zymosan (150 microgram/ml) resulted in considerable cell damage, as assessed by the sinusoidal release of lactate dehydrogenase and purine nucleoside phosphorylase. Cell damage was almost completely prevented by superoxide dismutase (50 U/ml) and catalase (150 U/ml), indicating ROS-related liver injury. ANP (200 nM) reduced Kupffer cell-induced injury via the guanylyl cyclase-coupled A receptor (GCA receptor) and cGMP: mRNA expression of the GCA receptor was found in hepatocytes, endothelial cells, and Kupffer cells, and the cGMP analog 8-bromo-cGMP (8-BrcGMP; 50 microM) was as potent as ANP in protecting from zymosan-induced cell damage. ANP and 8-BrcGMP significantly attenuated the prolonged increase of hepatic vascular resistance when Kupffer cell activation occurred. Furthermore, both compounds reduced oxidative cell damage following infusion of H2O2 (500 microM). In contrast, superoxide anion formation of isolated Kupffer cells was not affected by ANP and only moderately reduced by 8-BrcGMP. In conclusion, ANP protects the liver against Kupffer cell-related oxidant stress. This hormonal protection is mediated via the GCA receptor and cGMP, suggesting that the cGMP receptor plays a critical role in controlling oxidative cell damage. Thus ANP signaling should be considered as a new pharmacological target for protecting liver cells against the inflammatory response of activated Kupffer cells without eliminating the vital host defense function of these cells.

Topics: Animals; Atrial Natriuretic Factor; Catalase; Cyclic GMP; Endothelium, Vascular; Gene Expression; Guanylate Cyclase; Kupffer Cells; L-Lactate Dehydrogenase; Liver; Liver Diseases; Male; Oxidative Stress; Purine-Nucleoside Phosphorylase; Rats; Rats, Sprague-Dawley; Receptors, Atrial Natriuretic Factor; Reperfusion Injury; Superoxide Dismutase

Recent studies have reported that nitric oxide (NO) acts as a cytoprotective mediator in ischemia-reperfusion (IR) lung injury. We hypothesized that the addition of L-arginine to the perfusate would attenuate the increases in microvascular permeability and pulmonary vascular resistance.. Isolated rabbit lungs were reperfused for 60 min after 120 min warm ischemia. Lung injury was assessed using the fluid filtration coefficient (Kf), pulmonary vasucular resistance (PVR) before ischemia and after reperfusion, and a wet-to-dry lung weight ratio (W/D).. The Kf of the control group (without L-arginine) was significantly increased after reperfusion. Lungs perfused with L-arginine showed attenuation of the IR-induced increases in Kf and PVR. Addition of Nomega-nitro-L-arginine (L-NA), a NO synthase inhibitor, to the perfusate reduced the beneficial effects of L-arginine. The lungs perfused with dibutyryl-cyclic GMP (dbcGMP) showed attenuation of IR-induced increases in Kf and PVR. There were no significant differences in the W/D ratio between these groups.. These results demonstrate that L-arginine has beneficial effects on IR lung injury, perhaps due to enhancement of endothelial cGMP levels.

Topics: Animals; Arginine; Body Fluids; Capillary Permeability; Cyclic GMP; Enzyme Inhibitors; Hemodynamics; In Vitro Techniques; Ischemia; Lung; Nitroarginine; Pressure; Pulmonary Circulation; Rabbits; Reperfusion Injury; Trachea; Vascular Resistance

Exogenous nitric oxide reduces ischemia-reperfusion injury after solid organ transplantation. Tetrahydrobiopterin, an essential cofactor for nitric oxide synthases, may restore impaired endothelium-dependent nitric oxide synthesis. We evaluated whether tetrahydrobiopterin administration to the recipient attenuates lung reperfusion injury after transplantation in swine.. Unilateral left lung transplantation was performed in 15 weight-matched pigs (24-31 kg). Donor lungs were flushed with 1.5 L cold (1 degrees C) low-potassium-dextran solution and preserved for 20 hours. Group I animals served as controls. Group II and III animals were treated with a bolus of tetrahydrobiopterin (20 mg/kg). In addition, in group III a continuous infusion of tetrahydrobiopterin (10 mg/kg per hour over 5 hours) was given. One hour after reperfusion, the recipient right lung was occluded. Cyclic guanosine monophosphate levels were measured in the pulmonary venous and central venous blood. Extravascular lung water index, hemodynamic variables, lipid peroxidation, and neutrophil migration to the allograft were assessed.. In group III a significant reduction of extravascular lung water was noted in comparison with the controls (P =.0047). Lipid peroxidation in lung allograft tissue was significantly reduced in group II (P =.0021) and group III ( P =. 0077) in comparison with group I. Pulmonary venous levels of cyclic guanosine monophosphate increased up to 23 +/- 1 pmol/mL at 5 hours in group II and up to 40 +/- 1 pmol/mL in group III (group I, 4.1 +/- 0.5 pmol/mL [I vs III]; P <.001), whereas central venous levels of cyclic guanosine monophosphate were unchanged in all groups.. Tetrahydrobiopterin administration during lung allograft reperfusion may reduce posttransplantation lung edema and oxygen-derived free radical injury in the graft. This effect is mediated by local enhancement of the nitric oxide/cyclic guanosine monophosphate pathway.

Topics: Animals; Antioxidants; Biopterins; Cryopreservation; Cyclic GMP; Dextrans; Disease Models, Animal; Edema; Endothelium, Vascular; Extravascular Lung Water; Free Radical Scavengers; Hemodynamics; Infusions, Intravenous; Injections, Intravenous; Lipid Peroxidation; Lung Diseases; Lung Transplantation; Neutrophil Infiltration; Nitric Oxide; Nitric Oxide Synthase; Plasma Substitutes; Potassium; Pulmonary Veins; Reactive Oxygen Species; Reperfusion Injury; Swine

Substitution of the nitric oxide (NO) pathway reduces ischemia/reperfusion injury after lung transplantation. 8-Br-cGMP is a membrane-permeable analogue of cGMP, the second messenger of NO. In this study, we evaluated the effect of administration of 8-Br-cGMP in the flush solution on early graft function.. Unilateral left lung transplantation was performed in 10 weight-matched pairs of outbred pigs (24 to 31 kg). Donor lungs were flushed with 1.5 L cold (1 degree C) low potassium dextrane (LPD) solution and preserved for 20 hours. In group I (n = 5), 8-Br-cGMP (1 mg/kg) was added to the flush solution. In group II (n = 5), 8 microg/kg prostaglandin E1 (PGE1) was injected into the pulmonary artery (PA) before flush. One hour after reperfusion, the recipients' contralateral right PA and bronchus were ligated to assess graft function only. cGMP levels in the PA and pulmonary vein were measured. Extravascular lung water index (EVLWI), pulmonary vascular resistance, mean PA pressure, and gas exchange (PaO2) were assessed during a 5-hour observation period. Lipid peroxidation (thiobarbituric acid-reactive substance) and neutrophil migration to the allograft (myeloperoxidase activity) were measured at the end of the assessment.. In group I, a significant reduction of EVLWI (group I, 6.7 +/- 1.0 mL/kg vs group II, 10.1 +/- 0.6 ml/kg after 2 hours of reperfusion; p = 0.022), TBARS (group I, 65.6 +/- 10.0 pmol/g vs group II, 120.8 +/- 7.2 pmol/g, p = 0.0039), and MPO activity (group I, 0.8 +/- 0.1 change in optical density, (deltaOD)/mg/min vs group II, 1.7 +/- 0.3 deltaOD/mg/min, p = 0.036) was noted in comparison with group II. PaO2 levels tended to be higher in cGMP-treated animals, but the changes were not significant. Hemodynamic parameters did not differ between groups.. In this large animal model of lung allograft ischemia/reperfusion injury, 8-Br-cGMP as additive to the flush solution improves posttransplant lung edema, lipid peroxidation, and neutrophil migration to the allograft. This effect is not attributable to improved flush by vasodilation, as we compared 8-Br-cGMP with PGE1 given before flush in control animals.

Topics: Alprostadil; Animals; Cyclic GMP; Hemodynamics; Lung; Lung Transplantation; Neutrophils; Organ Preservation; Organ Preservation Solutions; Peroxidase; Pulmonary Gas Exchange; Reperfusion Injury; Swine; Vascular Resistance

This study was done to examine the protective effects of cyclic guanosine monophosphate (cGMP), a second messenger of nitric oxide, for ischemia/reperfusion injury of the liver, since it is known to induce vasodilatation and to inhibit platelet aggregation. Using an experimental model of porcine liver ischemia, 8-bromoguanosine 3',5' monophosphate, a cGMP analog, was continuously administered into the portal vein before ischemia and after reperfusion 30 min for each in the cGMP group (n = 6). Saline water was administered in the same way in the control group (n = 6). The cardiac output (CO), mean arterial blood pressure (MAP), portal venous flow (PVF), hepatic arterial flow (HAF), hepatic tissue blood flow (HTBF), and hepatic tissue cGMP level were determined. Hepatic enzymes and the bile discharge were also assessed as indicators of hepatic function. The hepatic tissue cGMP level was significantly higher, and PVF, HTBF, and the bile discharge were significantly greater in the cGMP group, while there were no remarkable differences between the groups with CO, MAP, HAF, and hepatic enzymes. In conclusion, the continuous supplementation of cGMP into the portal vein was found to be beneficial for preserving both the hepatic circulation and, consequently, the hepatic function after warm ischemia of porcine liver.

Topics: Analysis of Variance; Animals; Cyclic GMP; Disease Models, Animal; Guanosine Monophosphate; Hemodynamics; Injections, Intravenous; Liver Circulation; Liver Function Tests; Nitric Oxide; Portal Vein; Reference Values; Reperfusion Injury; Swine

Nitric oxide (NO) exerts cytoprotective effects against hepatic ischemia-reperfusion damage. This study was designed to evaluate which isoform of NO synthase (NOS) is implicated in the generation of cytoprotective NO and to investigate whether NO effects are mediated by cyclic GMP (cGMP). After partial ischemia for 45 min, liver damage was estimated by the release into plasma of cytolytic enzymes. Ischemia-reperfusion induced marked increases in plasma creatine kinase and lactate dehydrogenase after 1 h of reperfusion and of aminotransferases after 6 h of reperfusion. The pretreatment of ischemic rats with 8-bromo-cGMP (16 mg/kg i.v. 30 min before ischemia) or with L-arginine (the endogenous precursor of NO, 100 mg/kg i.v.) significantly diminished the ischemia-reperfusion-induced release of all these enzymes. This demonstrates that cGMP possesses hepatoprotective properties. By immunohistochemistry, we observed, after 6 h of reperfusion, an increase in endothelial NOS-III immunoreactivity, particularly in the small arteries and sinusoids. This NOS-III accumulation in endothelial cells could protect the liver against ischemia-reperfusion by the local generation of NO probably via cGMP.

Topics: Animals; Arginine; Creatine Kinase; Cyclic GMP; Endothelium, Vascular; Immunohistochemistry; L-Lactate Dehydrogenase; Liver; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Rats; Rats, Sprague-Dawley; Reperfusion Injury

FK409 is the first spontaneous nitric oxide (NO) donor known to increase plasma cyclic guanosine 3',5'monophosphate levels. In this study, we evaluated the effect of FK409 on pulmonary ischemia-reperfusion injury in an in situ warm ischemia canine model. Fourteen dogs were divided into two groups, and the FK409-treated group was given 5 micrograms/kg per min FK409. Warm ischemia was induced for 3 h. The arterial partial pressure of oxygen (PaO2), arterial oxygen saturation (SaO2), cardiac output (CO), left pulmonary vascular resistance (L-PVR), and endothelin-I (ET-I) were measured. A histologic study was performed, and polymorphonuclear neutrophils (PMNs) were also counted. The PaO2, SaO2, and L-PVR levels and PMNs after 30 min of reperfusion, ET-I after 2 h of reperfusion, and the 7-day survival rate were significantly (P < 0.05) better in the FK409-treated group than in the control group. The histologic damage was reduced in the FK409-treated group compared to the control group. FK409 appears to have a protective effect in ischemia-reperfusion injury of the lung.

Topics: Animals; Cardiac Output; Cyclic GMP; Dogs; Drug Evaluation, Preclinical; Endothelin-1; Ischemia; Lung; Nitric Oxide Donors; Nitro Compounds; Oxygen; Partial Pressure; Pulmonary Edema; Reperfusion Injury

We have tested if inhaled nitric oxide (NO) is beneficial in ischaemia-reperfusion (IR) lung injury using an isolated perfused rabbit lung model. Ischaemia for 60 min was followed by reperfusion and ventilation with nitric oxide 40 ppm (n = 6) or without nitric oxide ventilation (n = 6) for 60 min. In the control group (n = 6), the lungs were perfused continuously for 120 min. Permeability coefficient (Kfc) and vascular resistance (PVR) were measured serially for 60 min after reperfusion. We also determined the left lung W/D ratio and measured nitric oxide metabolites (NOx) and cGMP concentrations in bronchoalveolar lavage (BAL) fluid from the right lung. IR increased Kfc, PVR and W/D followed by decreased cGMP. Ventilation with nitric oxide restored these changes by preventing the decrease in cGMP. Differences in NOx concentrations in BAL fluid between the control and IR groups were not statistically significant. Our results indicate that IR impaired pulmonary vascular function and resulted in microvascular constriction and leakage. Ventilation with nitric oxide from the beginning of the reperfusion period improved pulmonary dysfunction such as vasoconstriction and capillary leak by restoring cGMP concentrations.

Topics: Animals; Bronchoalveolar Lavage Fluid; Capillary Permeability; Cyclic GMP; Female; Leukocyte Count; Lung; Nitric Oxide; Rabbits; Reperfusion Injury; Vascular Resistance; Vasodilator Agents

The aim of our studies was to investigate hormonal prevention of hepatic preservation damage by the atrial natriuretic peptide (ANP) and the mechanisms involved. Isolated perfusion of rat livers was performed in a nonrecirculating fashion. Twenty minutes of preischemic perfusion was performed with or without different concentrations of ANP, followed by 24-hour storage in cold University of Wisconsin (UW) solution. Two hundred nanomoles of ANP prevented hepatocellular damage during a 2-hour reperfusion period as indicated by a marked attenuation of the sinusoidal efflux of lactate dehydrogenase (LDH) and purine nucleoside phosphorylase (PNP), and by reduced Trypan blue uptake. Furthermore, postischemic bile flow as an indicator of liver function was significantly improved by about 60% with 200 nmol/L ANP. No protection was conveyed by 20 nmol/L ANP nor by pretreatment with 200 nmol/L ANP for only 10 minutes. The effects of ANP seemed to be mediated by the guanylate cyclase-coupled A (GC-A) receptor and cyclic guanosine monophosphate (cGMP): whereas expression of both GC-A and GC-B receptors as well as of the GC-C receptor was found, cGMP did protect from ischemia-reperfusion damage, but selective ligands of the B and C receptor did not. To begin to determine the mechanisms of ANP-mediated protection, different parameters were investigated: ANP had no effect on portal pressure as an indicator of hepatic circulation, nor on intracellular energy depletion determined by adenosine nucleotide concentration. However, the marked augmentation of nuclear factor kappaB (NF-kappaB) binding activity during reperfusion was prevented in ANP-pretreated livers. In conclusion, pretreatment with ANP protects the rat liver from cold ischemia-reperfusion damage. This effect is mediated via the GC-A receptor and cGMP, and may be linked to an influence of ANP on NF-kappaB activation. Thus, ANP signaling via the GC-A receptor should be considered as a new pharmacological target to prevent preservation injury of the liver.

Topics: Animals; Atrial Natriuretic Factor; Cold Temperature; Cyclic GMP; Guanylate Cyclase; Ischemia; L-Lactate Dehydrogenase; Liver; Male; NF-kappa B; Purine-Nucleoside Phosphorylase; Rats; Rats, Sprague-Dawley; Receptors, Atrial Natriuretic Factor; Reperfusion Injury

Cyclic nucleotides mediate intracellular signal transduction of several vasodilators. In addition to its vascular relaxant effects, cAMP is known to protect endothelial cells and to suppress Kupffer cell activation. On the other hand, cGMP potently ameliorates adhesion of leukocytes and platelets. We tested the effects of two analogs of cyclic nucleotides (8bromo cyclic adenosine monophosphate [8br-cAMP] and 8bromo cyclic guanosine monophosphate [8br-cGMP]) in rat liver preservation.. In experiment 1, either analog (0.1-1.0 mM) alone was added to University of Wisconsin (UW) solution in a survival study. In experiment 2, donors and recipients were also treated with 8br-cAMP or 8br-cGMP, with the following three groups tested: group 1=control; group 2=administration of 8br-cAMP to donors, UW solution, and recipients; group 3=administration of 8br-cGMP to donors, UW solution, and recipients. Experiment 3 tested combined treatments: group 4=administration of 8br-cGMP to donors and UW solution, and cAMP to recipients; group 5=administration of 8br-cAMP to donors and UW solution, and 8br-cGMP to recipients. To elucidate the roles of each nucleotide, two further groups were tested: group 6=administration of 8br-cAMP to donors and UW solution; group 7=administration of 8br-cGMP to recipients. In experiment 4, rats in groups 1, 5, 6, and 7 were killed at several time points after reperfusion, and percent graft blood flow (%BF), number of accumulated neutrophils, plasma levels of tumor necrosis factor-alpha and interleukin-1, and serum alanine aminotransferase levels were examined.. In experiments 1 and 2, no significant effect was observed on animal survival. In experiment 3, a significant increase in animal survival was observed only in group 5 (100%, 7/7, P=0.0004 vs. group 1: 16.7%, 2/12). In group 5, no improvement of %BF was observed during the early phase of reperfusion (15 and 30 min) compared with that in group 1. On the other hand, the %BF of group 5 was significantly higher in the later phase (6 hr), consistent with the decrease in accumulation of neutrophils observed then. Production of tumor necrosis factor-alpha and serum alanine aminotransferase levels were also reduced with this treatment. Histologically, the bleeding and segmental necrosis, observed in group 1, were completely prevented in group 5.. We conclude that restoration of grafts with cAMP and administration of cGMP to recipients led to successful transplantation, and that the two analogs acted synergistically in opposing preservation and reperfusion injury without improvement of graft blood flow during the early phase of reperfusion. The effect was due to their regulation of neutrophil activation and sequestration.

Topics: 8-Bromo Cyclic Adenosine Monophosphate; Adenosine; Allopurinol; Animals; Cyclic GMP; Drug Combinations; Drug Synergism; Glutathione; Insulin; Interleukin-1; Liver; Liver Circulation; Male; Organ Preservation; Organ Preservation Solutions; Postoperative Period; Raffinose; Rats; Rats, Inbred Lew; Reperfusion Injury; Tissue Donors; Tumor Necrosis Factor-alpha

In this study, the N-Methyl-D-Aspartate (NMDA) receptor-dependent nitric oxide and cyclic GMP (cGMP) synthesis in the course of reperfusion after 5 min of ischemia in gerbil brain hemispheres and cerebellum were investigated. Moreover, the role of the neuronal isoform of nitric oxide (NO) synthase (nNOS) in liberation of NO in postischemic brain and the involvement of NO in membrane lipoperoxidations activated during reperfusion were evaluated. Enhancement of Ca2+/calmodulin-regulated NOS activity and cGMP level in brain hemispheres and in cerebellum during reperfusion was found to be coupled to the activation of the NMDA receptor. cGMP concentration 40% above the control level was observed to persist up to 7 days after ischemia. The amount of conjugated double bounds in membrane lipids and the level of thiobarbituric acid reactive substances were increased exclusively in brain hemispheres, indicating activation of lipid peroxidation. The NMDA receptor antagonist, MK-801, eliminated, and a rather selective nNOS inhibitor, 7-Nitroindazole (7-NI) attenuated, NMDA receptor-evoked enhancement of NOS activity and cGMP level in brain hemispheres and in cerebellum during reperfusion. Moreover, 7-NI decreased significantly membrane lipid peroxidation during the early time of reperfusion. Histological examination demonstrated that 7-NI protects against death a selected population of neuronal cells in CA1 layer of hippocampus. It is suggested that NMDA receptor dependence of NO release during reperfusion is responsible for the degeneration of some populations of neurons and that the effect is mediated by activation of free radical formation and lipid peroxidation. Moreover, in cerebellum, ischemia-evoked activation of glutamatergic system stimulates NO-dependent signal transmission. Our results indicated that 7-NI has a significant ameliorating effect on biochemical alterations evoked by ischemia, suggesting nNOS inhibitors as a potential therapeutic agents in reperfusion injury.

Topics: Animals; Brain Ischemia; Cerebellum; Cerebral Cortex; Cyclic GMP; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Antagonists; Gerbillinae; Indazoles; Lipid Peroxidation; Male; Membrane Lipids; Neuroprotective Agents; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Prosencephalon; Receptors, N-Methyl-D-Aspartate; Reperfusion Injury; Signal Transduction; Thiobarbituric Acid Reactive Substances

Cyclic guanosine monophosphate (cGMP) is a potent second messenger for the nitric oxide pathway in the pulmonary vasculature. Increased cytosolic cGMP levels elicit pulmonary vasodilatation resulting in decreased pulmonary vascular resistance and maximized pulmonary function after ischemia-reperfusion injury. We hypothesized that the addition of a membrane-permeable cGMP analogue (8-bromo-cGMP) to a Euro-Collins (EC) preservation solution would ameliorate pulmonary reperfusion injury better than prostaglandin E1 injection alone after prolonged hypothermic ischemia.. All lungs from New Zealand White rabbits (weight, 3 to 3.5 kg) were harvested en bloc, flushed with EC solution, and reperfused with whole blood for 30 minutes. Group 1 lungs (immediate control) were immediately reperfused. Group 2 lungs (control) were stored inflated at 4 degrees C for 18 hours before reperfusion. Groups 3 and 4 lungs were flushed with EC solution containing 200 micromol/L 8-bromo-cGMP and stored at 4 degrees C for 18 and 30 hours, respectively. Fresh, nonrecirculated venous blood was used to determine single-pass pulmonary venous-arterial oxygen gradients at 10-minute intervals. Assays for cGMP, cyclic adenosine monophosphate, nitric oxide synthase activity, and myeloperoxidase were performed on all lung tissue samples. Wet to dry weight ratios were determined after 2 weeks of passive desiccation.. Oxygenation (venous-arterial oxygen gradient), pulmonary artery pressure, pulmonary vascular resistance, and edema formation were significantly improved in groups 3 and 4 (addition of 8-bromo-cGMP to EC plus 18 or 30 hours of hypothermic ischemia). Hypothermic storage (groups 2, 3, and 4) decreased both nitric oxide synthase activity and myeloperoxidase levels compared with immediate reperfusion (group 1).. These results suggest that the addition of a membrane-permeable cGMP analogue to an EC pulmonary flush solution improves pulmonary function after prolonged storage compared with EC and prostaglandin (E1) preservation alone. The finding of myeloperoxidase reduced levels after hypothermic storage and subsequent reperfusion may suggest a more important role for pulmonary hemodynamic control in mitigating pulmonary reperfusion injury.

Topics: Alprostadil; Animals; Blood Pressure; Cyclic AMP; Cyclic GMP; Hypertonic Solutions; Lung; Nitric Oxide Synthase; Organ Preservation; Organ Preservation Solutions; Oxygen; Peroxidase; Pulmonary Artery; Pulmonary Circulation; Rabbits; Reperfusion Injury; Vascular Resistance

Orthotopic liver transplantation (OLT) is often associated with hemodynamic instability upon reperfusion, recognized as postreperfusion syndrome. Changes in vascular tone due to humoral factors released upon reperfusion of the graft have been suggested as a possible mechanism. In this study, we looked at the perioperative changes in cyclic guanosine monophosphate (cGMP), a mediator of vascular smooth muscle relaxation, and investigated its relationship with hemodynamic parameters. cGMP was measured in the plasma of 14 patients undergoing OLT by radioimmunoassay serially at the preanhepatic and anhepatic phases, and after reperfusion at 30, 60, and 120 min. Hemodynamic data recorded were systemic and pulmonary arterial pressures, cardiac output, and pulmonary and systemic vascular resistance. cGMP decreased markedly after reperfusion from a baseline level of 5.33+/-0.7 ng/ml to 1.63+/-0.5 ng/ml (P<0.01). Pulmonary arterial pressure increased from 17+/-1.21 mmHg to 23.5+/-1.9 mmHg (P<0.05), and pulmonary vascular resistance increased from 62.8 +/-12.9 dynes/sec/cm5 to 135+/-42.7 dynes/sec/cm5 (P<0.01). Changes in cardiac output and systemic vascular resistance were not significant. The changes in cGMP correlated with pulmonary arterial pressure (r=0.74, P=0.005) and pulmonary vascular resistance (r=0.7, P=0.01). These data confirm the occurrence of hemodynamic changes during OLT, and provide evidence to suggest that the reduction in cGMP after reperfusion may mediate the vascular changes.

Topics: Blood Pressure; Cyclic GMP; Female; Hemodynamics; Humans; Liver Transplantation; Male; Middle Aged; Pulmonary Artery; Reperfusion Injury; Vascular Resistance

Plasma L-arginine is usually deficient immediately after hepatic reperfusion in orthotopic liver transplantation, which may also contribute to the occurrence of either hepatic ischemia-reperfusion injury or pulmonary hypertension. In this study, exogenous L-arginine was thus experimentally used to reverse the deficient status of the L-arginine/NO pathway. An in vivo model of 1 hr hepatic ischemia and reperfusion was thus tested in both rats (Experiment A) and pigs (Experiment B). In Experiment A, 10 mg/kg of L-arginine (group 1, n = 7), D-arginine (group 2, n = 7), or saline (group 3, n = 7) was administered through the portal vein. The hepatic tissue blood flow, at 20 min after reperfusion, improved in group 1 (70.7 +/- 7.0% of the preclamp levels) compared to groups 2 and 3. The serum glutamate oxaloacetate transaminase levels at 24 hr after reperfusion were also lower in group 1 (320 +/- 22.2 IU/L) than in either group 2 or group 3. The intrahepatic NO levels showed a temporal burst (> 15,000 pA current) after reperfusion only in group 1. In Experiment B, 10 mg/kg of L-arginine (group 4, n = 5), D-arginine (group 5, n = 5), or 10 ml of saline (group 6, n = 5) was administered through the portal vein. In group 4, the MPAP (mean pulmonary arterial pressure)/MAP (mean arterial pressure) was lower than that observed in groups 5 and 6. In conclusion, exogenous L-arginine administered from the portal vein was thus found to be effective in mitigating both portal hypertension and reperfusion injury by producing an increased amount of NO immediately after reperfusion.

Topics: Animals; Arginine; Aspartate Aminotransferases; Blood Pressure; Citrulline; Cyclic GMP; Female; Hemodynamics; Liver; Male; Nitrates; Nitric Oxide; Nitrites; Rats; Rats, Wistar; Reperfusion Injury; Swine; Time Factors

Topics: Animals; Arginine; Citrulline; Cyclic GMP; Liver; Liver Circulation; Male; Nitrates; Nitric Oxide; Nitrites; Rats; Rats, Wistar; Regional Blood Flow; Reperfusion; Reperfusion Injury

Topics: Adenosine; Allopurinol; Analysis of Variance; Animals; Arginine; Cyclic GMP; Enzyme Inhibitors; Glutathione; Insulin; Kidney; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Organ Preservation; Organ Preservation Solutions; Raffinose; Rats; Rats, Wistar; Reperfusion Injury; Temperature

Topics: Animals; Arginine; Cyclic GMP; Enzyme Inhibitors; Kidney; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Organ Preservation; Rats; Rats, Wistar; Reperfusion Injury

This study was aimed to examine properties and changes in nitric oxide synthase (NOS) activity and cGMP level during reperfusion after 5 min of brain ischemia in gerbils. Animals were treated 5 min before ischemia with NOS inhibitors: N-Nitro-L-arginine (NNLA), or 7-Nitroindazole (7-NI), or with the inhibitor of guanylate cyclase, LY 83583, or with hydrocortisone for 7 days before ischemia. Northern blot analysis was performed using specific cDNA for inducible NOS. It was observed that ischemia significantly enhances NOS activity and cGMP level. During reperfusion, biphasic increase in NOS activity and cGMP level took place with two peaks 15 min and 2 h after ischemia. NNLA, 7-NI, and LY 83583 eliminated enhancements of NOS activity and cGMP level, whereas glucocorticoid remained without effect. There was no activation of gene encoding inducible NOS (iNOS). Our results indicate that ischemia-reperfusion activates constitutive NOS. It is suggested that nitric oxide (NO) production during reperfusion is related to neuronal degeneration and that inhibitor of NOS offers a new therapeutical strategies.

Topics: Animals; Brain; Brain Chemistry; Brain Ischemia; Cyclic GMP; Gerbillinae; Male; Nitric Oxide Synthase; Reperfusion Injury

The goal of these experiments was to determine whether the perturbation of ischemia-reperfusion has an age-dependent effect on subsequent endothelial cell production of nitric oxide. Three- and 35-d-old swine in the experimental group were exposed to 1-h partial ischemia (90% flow reduction) and 2-h reperfusion in vivo by creation and then removal of a mesenteric artery coarctation. Control subjects underwent exposure of the mesenteric artery only. After reperfusion, gut vascular resistance had increased 44 +/- 6% in 3-d-old, but had decreased 41 +/- 4% in 35-d-old subjects. At the completion of the in vivo portion of the protocol mesenteric artery was removed, and nitric oxide production was estimated in vitro, by measuring cGMP production by vessel segments or by measuring relaxation of phenylephrine-precontracted rings, both after stimulation of nitric oxide production by substance P or the calcium ionophore A23187. Compared with control, mesenteric artery segments from 3-d-old subjects demonstrated reductions in basal, substance P-stimulated (10(-8) M) and A23187-stimulated (10(-7) M) cGMP accumulation of 50 +/- 7%, 66 +/- 6% and 78 +/- 7%. Mesenteric artery segments from 35-d-old subjects demonstrated increases in basal, substance P-stimulated, or A23187-stimulated cGMP accumulations of 114 +/- 14%, 92 +/- 8%, or 78 +/- 9%. Compared with control, I/R rings from 3-d-old subjects demonstrated reductions in substance P-induced (10(-8) M) or A23187-induced (10(-7) M) relaxations of 56 +/- 7% or 30 +/- 7%. In contrast, 35-d-old ischemia-reperfusion rings demonstrated increases in substance P- or A23187-induced relaxation of 36 +/- 8% or 98 +/- 11%. It is concluded that ischemia-reperfusion has an age-dependent effect on endothelial production of NO within in vitro postnatal mesenteric artery and that these changes mirror the effects of ischemia-reperfusion on gut vascular resistance in vivo.

Topics: Age Factors; Animals; Calcimycin; Cyclic GMP; Endothelium; Hemodynamics; Intestinal Mucosa; Ionophores; Mesenteric Arteries; Nitric Oxide; Reperfusion Injury; Substance P; Swine

A major hemodynamic feature of acute lung injury is pulmonary hypertension caused by pulmonary vasoconstriction. Impairment of the mechanisms of pulmonary vasorelaxation may contribute to this pulmonary vasoconstriction. This study examined the effect of mesenteric ischemia/reperfusion (I/R) on lung neutrophil accumulation and endothelial-dependent and -independent cyclic 3'-5' guanosine monophosphate-mediated pulmonary vasorelaxation in rats.. Rats were studied after 1 hour of superior mesenteric artery occlusion and 2 hours of reperfusion. Lung neutrophil accumulation was determined by myeloperoxidase assay (MPO). The following mechanisms of pulmonary vasorelaxation were studied in isolated pulmonary artery rings by generating dose response curves (10(-9) to 10(-6)mol/L): (1) receptor-dependent, endothelial-dependent relaxation (response to acetylcholine), (2) receptor-independent, endothelial-dependent relaxation (response to the calcium ionophore, A23187), and (3) endothelial-independent relaxation (response to sodium nitroprusside [SNP]).. Lung MPO activity was significantly increased from 2.4 +/- 0.2 units/gm lung weight in controls to 10.3 +/- 0.4 after mesenteric I/R (p < 0.05). The vasorelaxation response to SNP was not different after mesenteric I/R, but vasorelaxation by both acetylcholine and A23187 were significantly impaired.. Endothelial-dependent pulmonary vasorelaxation is significantly impaired after mesenteric I/R. Such impairment of pulmonary vasorelaxation may help tip the net balance of pulmonary vasomotor tone toward vasoconstriction and contribute to the pulmonary hypertension seen in acute lung injury.

Topics: Acetylcholine; Animals; Calcimycin; Cyclic GMP; Endothelium, Vascular; Hypertension, Pulmonary; In Vitro Techniques; Lung; Lung Injury; Male; Neutrophils; Nitroprusside; Peroxidase; Pulmonary Artery; Pulmonary Circulation; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Splanchnic Circulation; Vasodilation; Vasodilator Agents

Ischaemia/reperfusion (I/R) of the intestine causes mucosal injury associated with a high death rate in rats.. To investigate whether nitric oxide (NO) might be implicated in the recovery of the intestinal mucosa after ischaemic insult.. Wistar rats were subjected to mesenteric artery occlusion for 90 minutes. The animals were given either L-arginine, the substrate of NO synthase, or molsidomine, a NO donor. The controls received casein hydrolysate. The compounds were administered by gavage 19, 16, and 1.5 hours before ischaemia. Mucosal barrier permeability and cGMP content were determined 24 hours after ischaemia.. Survival after I/R was 50% in the control group. Animals treated with L-arginine or molsidomine exhibited a higher survival rate (70% and 83% respectively). Mucosal barrier permeability was decreased in rats receiving L-arginine or molsidomine compared with controls (4.0 (0.9) and 2.6 (0.6) v 11.2 (1.6) 14C-PEG pmol/segment, p < 0.05). Increased cGMP content was seen in the mucosa of the L-arginine group.. The findings suggest that pretreatment with L-arginine or molsidomine ameliorates survival after intestinal I/R and improves mucosal barrier function.

Topics: Amino Acids, Essential; Analysis of Variance; Animals; Arginine; Cyclic GMP; Intestinal Mucosa; Male; Mesenteric Arteries; Molsidomine; Nitric Oxide Synthase; Permeability; Polyethylene Glycols; Rats; Rats, Wistar; Reperfusion Injury; Survival Rate; Treatment Outcome; Vasodilator Agents

Nitric oxide (NO), identified as a mediator of endothelium-dependent relaxation of vascular smooth muscle, is known to cause a number of inflammatory diseases, especially ischemia-reperfusion injury. This experimental study, using a rabbit epigastric island flap, was designed to investigate whether skin flap ischemia followed by reperfusion influences serum NO concentrations. In addition, the author investigated the effects of NO synthase inhibitors and heparin on skin flap ischemia. Serum NO concentrations after 15, 30, 45, and 60 minutes of ischemia followed by reperfusion were significantly increased compared with non-ischemic controls and elevated flaps. On the other hand, serum NO concentrations were suppressed in nitro-amino-methyl-L-arginine- and aminoguanidine-treated animals. Furthermore, administration of heparin increased serum NO concentrations in controls and animals with elevated flaps, but decreased serum NO concentrations in ischemic flaps with subsequent reperfusion. These results suggest that NO is one of the factors responsible for ischemia-reperfusion injury and that NO synthase inhibitors and heparin may protect against such injury.

Topics: Animals; Cyclic GMP; Enzyme Inhibitors; Guanidines; Heparin; Ischemia; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Rabbits; Reperfusion Injury; Skin; Surgical Flaps

To gain further insights into the greater susceptibility to acute ischemic renal injury (ARF, 45 minutes of renal arteries clamping) of old rats (O, 24-26 months) as against young ones (Y, 3-4 months), and the possible role of endothelium derived relaxing factor (EDRF) and its influence on ischemic renal function.. Basal renal dynamics was compared between O and Y.. Two hours after ischemia, the decrease of GFR RPF, and UV was more severe in O than in Y. Urine cGMP was significantly dropped in both groups. Intravenous administration of L-arginine increasld urine cGMP of the O as well as the Y, with that of the Y higher than that in O. GFR, RPF, UV were elevated significantly in the O. Simultaneous administration of L-NNA, the analogue of L-Arginine could partially antagonize its effect on elevating RPF. RT-PCR technique was used to detect renal inducible nitric oxide synthase (iNOS) expression. Although the ischemic kidney was capable of expressing iNOS mRNA in the presence of L-arginine after ischemia, the production of NO in the old may not be regulated at the transcriptional level, other factors such as NOS enzyme activity, availability of L-arginine and O2, metabolism of NO after its production were suspected to be involved.

Topics: Aging; Animals; Arginine; Cyclic GMP; Ischemia; Kidney; Male; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Wistar; Renal Agents; Reperfusion Injury

Topics: Analysis of Variance; Animals; Arginine; Cyclic GMP; Ischemia; Kidney; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Rats; Rats, Wistar; Reperfusion Injury; Time Factors

Pulmonary vascular resistance is significantly increased in the transplanted lung. We hypothesized that the ischemic or reperfusion injuries incurred by the transplanted lung may produce pulmonary vasomotor dysfunction, which in turn may produce increased pulmonary vascular resistance. In a dog model of autologous lung transplantation, the purpose of this study was to examine the following mechanisms of pulmonary vasomotor control and to relate each of them to cold ischemia and to reperfusion: (1) endothelium-dependent cyclic guanosine monophosphate-mediated vasorelaxation (response to acetylcholine 10(-6) mol/L), (2) endothelium-independent cyclic guanosine monophosphate-mediated vasorelaxation (response to sodium nitroprusside 10(-6) mol/L), and beta-adrenergic cyclic adenosine monophosphate-mediated vasorelaxation (response to isoproterenol 10(-6) mol/L). Autologous right lung transplantation was performed in five dogs. At each of three times, two third-order pulmonary arteries were dissected from each transplanted lung and studied: control (immediately after harvest), cold ischemia (3 hours in 4 degrees C saline solution), and cold ischemia plus reperfusion (1 hour after lung reimplantation). The vasorelaxing effects of acetylcholine, sodium nitroprusside, and isoproterenol were studied in isolated pulmonary arterial rings, suspended on fine wire tensiometers in individual organ chambers. Statistical analysis was by analysis of variance. Results demonstrated significant dysfunction of beta-adrenergic cyclic adenosine monophosphate-mediated relaxation after cold ischemia alone, and this dysfunction was exacerbated by reperfusion. Endothelium-dependent cyclic guanosine monophosphate-mediated relaxation was not impaired by cold ischemia alone but was significantly impaired by reperfusion. Endothelium-independent cyclic guanosine monophosphate-mediated relaxation was not impaired by cold ischemia or reperfusion. We conclude that cold ischemia and reperfusion each produce different patterns of pulmonary vasomotor dysfunction. Cumulatively, such dysfunction may contribute to increased pulmonary vascular resistance in the transplanted lung.

Topics: Animals; Cyclic AMP; Cyclic GMP; Dogs; Ischemia; Lung; Lung Transplantation; Muscle, Smooth, Vascular; Reperfusion Injury; Vascular Resistance; Vasomotor System

Recent data have demonstrated that inhibition of nitric oxide synthesis exacerbated the mucosal injury associated with reperfusion of the postischemic intestine. In this study, using a feline 1-h intestinal ischemia followed by reperfusion model, we tested the possibility that exogenous sources of nitric oxide may prevent the reperfusion-induced mucosal barrier disruption and examined the mechanisms involved. Mucosal barrier integrity was assessed by determining 51Cr-EDTA clearance from blood to lumen. Intestinal blood flow and resistance were also determined. Reperfusion after 1 h of ischemia significantly increased 51Cr-EDTA clearance (0.05 +/- 0.01 to 0.35 +/- 0.07 ml.min-1.100 g-1) and decreased intestinal blood flow by 50%. Exogenous sources of nitric oxide including SIN-1, CAS-754, and nitroprusside as well as exogenous L-arginine all reduced reperfusion-induced mucosal barrier dysfunction without improving intestinal blood flow. Inhibition of endogenous nitric oxide with NG-nitro-L-arginine methyl ester between 1 and 2 h of reperfusion further augmented the rise in mucosal permeability associated with ischemia-reperfusion. Addition of the permeable analogue of guanosine 3',5'-cyclic monophosphate, 8-bromoguanosine 3',5'-cyclic monophosphate, improved reperfusion-induced intestinal blood flow significantly but did not provide protection against mucosal barrier disruption associated with the first hour of ischemia-reperfusion. Exogenous sources of nitric oxide can reduce reperfusion-induced mucosal barrier dysfunction independent of alterations in intestinal blood flow.

Topics: Animals; Cats; Cyclic GMP; Intestinal Mucosa; Intestines; Ischemia; Molsidomine; Nitric Oxide; Nitroprusside; Permeability; Regional Blood Flow; Reperfusion Injury; Sydnones