2-2--(hydroxynitrosohydrazono)bis-ethanamine and Disease-Models--Animal

Depression and Type 2 diabetes mellitus are interrelated conditions, but the underlying neurobiology is insufficiently understood. The current study compared the effects of a pharmacological manipulation with 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) that targets neurobiological processes by adenosine 5'-monophosphate-activated protein kinase activation versus exercise on depression-like behavior and nitric oxide (NO)-related measures.. A mouse model of a depression-like and insulin-resistant state, induced by the co-treatment of high-fat diet and corticosterone administration, was used to examine the antidepressant action of AICAR and exercise.. Data showed that AICAR was a putative antidepressant in the depression-like and insulin-resistant mice (total ambulatory distance in the open-field test was 5120.69 ± 167.47 cm, mobility duration in the forced swim test was 17.61 ± 1.54 seconds, latency to feed in the novelty suppressed feeding test was 255.67 ± 37.80 seconds; all p values < .05). Furthermore, the antidepressant actions of AICAR required endothelial nitric oxide synthase activity with increased NO production in the prefrontal cortex, whereas corticosterone-induced expression of neuronal nitric oxide synthase and NO production may increase the risk of depression. In contrast to the traditional antidepressants such as ketamine and imipramine, AICAR interfered with the effects of insulin in skeletal muscle in the context of high-fat diet, consistent with the potential antidepressant effects of AICAR. Exercise also resulted in activation of adenosine 5'-monophosphate-activated protein kinase, nitric oxide synthase, and NO production (all p values < .01), which in turn may be implicated in the antidepressant effects of exercise.. These findings suggest that NO is an essential signal mediating the antidepressant actions of AICAR. Ultimately, the concurrent effects of AICAR on brain insulin action and mitochondrial function suggest a potential of neural insulin resistance, which may contribute to our understanding of the comorbidity of depression and Type 2 diabetes.

Topics: Adenylate Kinase; Aminoimidazole Carboxamide; Animals; Antidepressive Agents; Combined Modality Therapy; Corticosterone; Depressive Disorder; Diet, High-Fat; Disease Models, Animal; Drug Evaluation, Preclinical; Enzyme Activation; Fluoxetine; Imipramine; Insulin Resistance; Ketamine; Male; Mice; Mice, Inbred C57BL; Nerve Tissue Proteins; NG-Nitroarginine Methyl Ester; Nitric Oxide; Physical Conditioning, Animal; Prefrontal Cortex; Ribonucleotides; Triazenes

We investigated efficacy of nitric oxide (NO) against Leishmania donovani. NO is a mediator of host response to infection, with direct parasiticidal activity in addition to its role in signalling to evoke innate macrophage responses. However, it is short-lived and volatile, and is therefore difficult to introduce into infected cells and maintain inracellular concentrations for meaningful periods of time. We incorporated diethylenetriamine NO adduct (DETA/NO), a prodrug, into poly(lactide-co-glycolide) particles of ∼200 nm, with or without amphotericin B (AMB). These particles sustained NO levels in mouse macrophage culture supernatants, generating an area under curve (AUC0.08-24h) of 591.2 ± 95.1 mM × h. Free DETA/NO resulted in NO peaking at 3 h and declining rapidly to yield an AUC of 462.5 ± 193.4. Particles containing AMB and DETA/NO were able to kill ∼98% of promastigotes and ∼76% of amastigotes in 12 h when tested in vitro. Promastigotes and amastigotes were killed less efficiently by particles containing a single drug- either DETA/NO (∼42%, 35%) or AMB (∼90%, 50%) alone, or by equivalent concentrations of drugs in solution. In a pre-clinical efficacy study of power >0.95 in the hamster model, DETA/NO particles were non-inferior to Fungizone® but not Ambisome®, resulting in significant (∼73%) reduction in spleen parasites in 7 days. Particles containing both DETA/NO and AMB were superior (∼93% reduction) to Ambisome®. We conclude that NO delivered to the cytosol of macrophages infected with Leishmania possesses intrinsic activity and adds significantly to the efficacy of AMB.

Topics: Amphotericin B; Animals; Antiprotozoal Agents; Cell Survival; Cricetinae; Disease Models, Animal; Drug Therapy, Combination; Leishmania donovani; Leishmaniasis, Visceral; Macrophages; Mice; Nanoparticles; Nitroso Compounds; Treatment Outcome

Thrombus formation is determined by the balance between pro- thrombotic mediators and anti-thrombotic factors.High-density lipoprotein (HDL) from healthy subjects exerts anti-thrombotic properties. Whether this is also the case for HDL from patients with stable coronary heart disease (CHD) or acute coronary syndrome (ACS) is unknown.In human aortic endothelial cells in culture,HDL (50 µg/ml) from healthy subjects (HS) inhibited thrombin-induced tissue factor (TF) expression and activity, while HDL (50 µg/ml) from CHD and ACS patients did not. Similarly, only healthy HDL increased endothelial tissue factor pathway inhibitor (TFPI) expression and tissue plasminogen activator (tPA) release, while HDL from CHD and ACS patients had no effect. Healthy HDL inhibited thrombin-induced plasminogen activator inhibitor type 1 (PAI-1) expression, while HDL from ACS patients enhanced endothelial PAI-1 expression. Inhibition of nitric oxide (NO) formation with L-NAME (100 µmol/l) abolished the anti-thrombotic effects of healthy HDL on TF, TFPI, and tPA expression. The exogenous nitric oxide donor, DETANO, mimicked the effects of healthy HDL and counterbalanced the loss of anti-thrombotic effects of HDL from CHD and ACS patients in endothelial cells. In line with this observation, healthy HDL, in contrast to HDL from CHD and ACS patients, increased endothelial NO production. In the laser-injured carotid artery of the mouse, thrombus formation was delayed in animals treated with healthy HDL compared with mice treated with vehicle or HDL from patients with CHD or ACS. In conclusion, HDL from CHD and ACS patients loses the ability of healthy HDL to suppress TF and to increase TFPI and t-PA and instead enhances PAI-1 and arterial thrombus formation.

Topics: Acute Coronary Syndrome; Adult; Aged; Animals; Aorta; Blood Coagulation; Carotid Artery Injuries; Cells, Cultured; Coronary Disease; Disease Models, Animal; Endothelial Cells; Humans; Lipoproteins; Lipoproteins, HDL; Mice; Middle Aged; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitroso Compounds; Plasminogen Activator Inhibitor 1; Platelet Aggregation; Thrombin; Thromboplastin; Tissue Plasminogen Activator

We tested the hypothesis that a nitric oxide donor, DETA-NONOate, up-regulates stromal cell-derived factor-1 (SDF1) and angiopoietin 1 (Ang1) in the ischemic brain and their respective receptors chemokine CXC motif receptor 4 (CXCR4) and Tie2 in the subventricular zone (SVZ) and thereby promote SVZ neuroblast cell migration after stroke. C57BL/6J mice were subjected to middle cerebral artery occlusion (MCAo), and 24 hr later DETA-NONOate (0.4 mg/kg) or phosphate-buffered solution was intravenously administered. Mice were sacrificed at 14 days for histological assessment or sacrificed at 3 days for analysis by real-time polymerase chain reaction and migration after MCAo. To elucidate whether SDF1/CXCR4 and Ang1/Tie2 pathways mediate DETA-NONOate-induced SVZ migration after stroke, SDF1alpha, Ang1 peptide, a specific antagonist of CXCR4 (AMD3100), and a neutralizing antibody of Tie2 (anti-Tie2) were used in vitro. DETA-NONOate significantly increased the percentage area of doublecortin (DCX, a marker of migrating neuroblasts)-immunoreactive cells in the SVZ and ischemic boundary zone. DETA-NONOate significantly increased the expression of SDF1 and Ang1 in the ischemic border and up-regulated CXCR4 and Tie2 in the SVZ compared with MCAo control. DCX-positive cell migration from SVZ explants was significantly increased in the DETA-NONOate treatment group compared with MCAo-alone animals. In vitro, SDF1alpha and Ang1 significantly increased SVZ explants cell migration. In addition, inhibition of CXCR4 or Tie2 significantly attenuated DETA-NONOate-induced SVZ cell migration. Our data indicate that treatment of stroke with a nitric oxide donor up-regulates SDF1/CXCR4 and Ang1/Tie2 pathways and thereby likely increases SVZ neuroblast cell migration.

Topics: Adult Stem Cells; Analysis of Variance; Angiopoietin-1; Animals; Cell Movement; Cerebral Ventricles; Chemokine CXCL12; Disease Models, Animal; Doublecortin Protein; In Vitro Techniques; Infarction, Middle Cerebral Artery; Male; Mice; Mice, Inbred C57BL; Nitric Oxide Donors; Nitroso Compounds; Receptor, TIE-2; Receptors, CXCR4; Up-Regulation

Arteriogenesis supports restored perfusion in the ischemic brain and improves long-term functional outcome after stroke. We investigate the role of endothelial nitric oxide synthetase (eNOS) and a nitric oxide (NO) donor, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl) amino] diazen-1-ium-1, 2-diolate (DETA-NONOate), in promoting arteriogenesis after stroke. Adult wild-type (WT, n=18) and eNOS-knockout (eNOS(-/-), n=36) mice were subjected to transient (2.5 h) right middle cerebral artery occlusion (MCAo) and were treated with or without DETA-NONOate (0.4 mg/kg) 24 h after MCAo. Functional evaluation was performed. Animals were sacrificed 3 days after MCAo for arterial cell culture studies, or 14 days for immunohistochemical analysis. Consistent with previous studies, eNOS(-/-) mice exhibited a higher mortality rate (P<0.05, n=18/group) and more severe neurological functional deficit after MCAo than WT mice (P<0.05, n=12/group). Decreased arteriogenesis, was evident in eNOS(-/-) mice compared with WT mice, as demonstrated by reduced vascular smooth muscle cell (VSMC) proliferation, arterial density and diameter in the ischemic brain. eNOS(-/-) mice treated with DETA-NONOate had a significantly decreased mortality rate and improved functional recovery, and exhibited enhanced arteriogenesis identified by increased VSMC proliferation, and upregulated arterial density and diameter compared to eNOS(-/-) mice after stroke (P<0.05, n=12/group). To elucidate the mechanisms underlying eNOS/NO mediated arteriogenesis, VSMC migration was measured in vitro. Arterial cell migration significantly decreased in the cultured common carotid artery (CCA) derived from eNOS(-/-) mice 3 days after MCAo compared to WT arterial cells. DETA-NONOate-treatment significantly attenuated eNOS(-/-)-induced decrease of arterial cell migration compared to eNOS(-/-) control artery (P<0.05; n=6/group). Using VSMC culture, DETA-NONOate significantly increased VSMC migration, while inhibition of NOS significantly decreased VSMC migration (P<0.05; n=6/group). Our data indicated that eNOS not only promotes vascular dilation but also increases VSMC proliferation and migration, and thereby enhances arteriogenesis after stroke. Therefore, increase eNOS may play an important role in regulating of arteriogenesis after stroke.

Topics: Actins; Animals; Carotid Artery, Common; Cell Movement; Cell Proliferation; Cells, Cultured; Disease Models, Animal; Enzyme Inhibitors; Infarction, Middle Cerebral Artery; Ki-67 Antigen; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocytes, Smooth Muscle; Neovascularization, Physiologic; Neurologic Examination; NG-Nitroarginine Methyl Ester; Nitric Oxide Donors; Nitric Oxide Synthase Type III; Nitroso Compounds; Recovery of Function; Time Factors

Increasing evidence suggests that depression may be associated with a lack of hippocampal neurogenesis. Our recent study shows that endogenous nitric oxide (NO) contributes to chronic mild stress (CMS)-induced depression by suppressing hippocampal neurogenesis.. The aim of this study was to investigate the effects of exogenous NO in CMS-induced depression in young adult mice.. In normal mice, administration of a pure NO donor (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl) aminio] diazen-1-ium-1,2-diolate (DETA/NONOate; 0.4 mg/kg, i.p., for 7 days) produced an antidepressant-like effect and significantly increased hippocampal neurogenesis. The mice exposed to CMS exhibited behavioral changes typical of depression and impaired neurogenesis in the hippocampus. Treatment with DETA/NONOate (0.4 mg/kg, i.p., for 7 days) reversed CMS-induced behavioral despair and hippocampal neurogenesis impairment. We treated mice with a telomerase inhibitor 3'-azido-deoxythymidine (AZT; 100 mg/kg, i.p., for 14 days) to disrupt neurogenesis. From day 4 to day 11 of AZT treatment, mice were injected with DETA/NONOate (0.4 mg/kg, i.p., for 7 days). Disrupting hippocampal neurogenesis blocked the antidepressant effect of DETA/NONOate.. Our findings suggest that exogenous NO benefits chronic stress-induced depression by stimulating hippocampal neurogenesis and may represent a novel approach for the treatment of depressive disorders.

Topics: Animals; Antidepressive Agents; Behavior, Animal; Depression; Disease Models, Animal; Hippocampus; Male; Mice; Mice, Knockout; Neurogenesis; Nitric Oxide Donors; Nitric Oxide Synthase Type I; Nitroso Compounds; Stress, Psychological; Zidovudine

The addition of a nitric oxide (NO)-releasing moiety to prednisolone was shown to enhance the anti-inflammatory activity of this glucocorticoid in some experimental conditions, but its effectiveness in the context of eosinophilic inflammation remains to be elucidated.. This study compared the anti-inflammatory effect of prednisolone to a NO-releasing derivative of prednisolone, NCX-1015, using a model of allergen-evoked eosinophil recruitment in rats. The efficacy of a NO-donor compound, DETA-NONOate, was also assessed for comparison.. Wistar rats were actively sensitized with Al(OH)(3) plus ovalbumin and 14 days later challenged with antigen intrapleurally. Treatments were performed locally 1 h before challenge. Cysteinyl-leucotrienes (Cys-LT) and eotaxin were measured by ELISA.. Antigen challenge induced an eosinophil infiltration at 12 h, maximal at 24 h. It also caused an increase in the levels of Cys-LTs in the pleural exudate and in the expression of 5-lipoxygenase (5-LO) in infiltrated leucocytes at 6 h, peaking at 12 h and persisting for at least 24 h. Treatment with equimolar doses of prednisolone and NCX-1015 inhibited the late eosinophil infiltration, although the dose required to produce maximal inhibition was about one-tenth that of prednisolone. Cys-LT generation and 5-LO expression were inhibited by NCX-1015 but not by prednisolone. Treatment with prednisolone combined with the NO-donor DETA-NONOate led to a greater inhibition of the eosinophilia and Cys-LT generation as compared with either drug alone. Administration of the steroid receptor antagonist RU 486, 1 h before prednisolone and NCX-1015, abolished the inhibitory effect of the former, under conditions where it only partially affected the latter.. Our findings indicate that NCX-1015 provided a greater anti-inflammatory effect than prednisolone on the allergic eosinophil recruitment in rats, suggesting that NO-releasing steroids can be considered as a promising therapeutic approach to allergic diseases.

Topics: Animals; Anti-Inflammatory Agents; Arachidonate 5-Lipoxygenase; Chemokine CCL11; Cysteine; Disease Models, Animal; Drug Therapy, Combination; Eosinophilia; Eosinophils; Hypersensitivity; Leukocytes; Leukocytes, Mononuclear; Leukotrienes; Male; Mifepristone; Neutrophils; Nitric Oxide Donors; Nitroso Compounds; Ovalbumin; Pleural Cavity; Pleurisy; Prednisolone; Rats; Rats, Wistar; Receptors, Glucocorticoid

Neovascularization may contribute to functional recovery after neural injury. Combination treatment of stroke with a nitric oxide donor, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl) amino] diazen-1-ium-1, 2-diolate (DETA-NONOate) and bone marrow stromal cells promotes functional recovery. However, the mechanisms underlying functional improvement have not been elucidated. In this study, we tested the hypothesis that combination treatment upregulates angiopoietin-1 and its receptor Tie2 in the ischemic brain and bone marrow stromal cells, thereby enhancing cerebral neovascularization after stroke. Adult wild type male C57BL/6 mice were i.v. administered PBS, bone marrow stromal cells 5x10(5), DETA-NONOate 0.4 mg/kg or combination DETA-NONOate with bone marrow stromal cells (n=12/group) after middle cerebral artery occlusion. Combination treatment significantly upregulated angiopoietin-1/Tie2 and tight junction protein (occludin) expression, and increased the number, diameter and perimeter of blood vessels in the ischemic brain compared with vehicle control (mean+ or -S.E., P<0.05). In vitro, DETA-NONOate significantly increased angiopoietin-1/Tie2 protein (n=6/group) and Tie2 mRNA (n=3/group) expression in bone marrow stromal cells. DETA-NONOate also significantly increased angiopoietin-1 protein (n=6/group) and mRNA (n=3/group) expression in mouse brain endothelial cells (P<0.05). Angiopoietin-1 mRNA (n=3/group) was significantly increased in mouse brain endothelial cells treated with DETA-NONOate in combination with bone marrow stromal cell-conditioned medium compared with cells treated with bone marrow stromal cell-conditioned medium or DETA-NONOate alone. Mouse brain endothelial cell capillary tube-like formation assays (n=6/group) showed that angiopoietin-1 peptide, the supernatant of bone marrow stromal cells and DETA-NONOate significantly increased capillary tube formation compared with vehicle control. Combination treatment significantly increased capillary tube formation compared with DETA-NONOate treatment alone. Inhibition of angiopoietin-1 significantly attenuated combination treatment-induced tube formation. Our data indicated that combination treatment of stroke with DETA-NONOate and bone marrow stromal cells promotes neovascularization, which is at least partially mediated by upregulation of the angiopoietin-1/Tie2 axis.

Topics: Angiopoietin-1; Animals; Bone Marrow Transplantation; Capillaries; Cells, Cultured; Cerebral Arteries; Coculture Techniques; Disease Models, Animal; Endothelium, Vascular; Male; Mice; Mice, Inbred C57BL; Neovascularization, Physiologic; Nitric Oxide Donors; Nitroso Compounds; Receptor, TIE-2; Regeneration; RNA, Messenger; Stroke; Stromal Cells; Tight Junctions; Up-Regulation

Type II diabetes mellitus (DM) and metabolic syndrome are associated with accelerated restenosis following vascular interventions due to neointimal hyperplasia. The efficacy of nitric oxide (NO)-based therapies is unknown in these environments. Therefore, the aim of this study is to examine the efficacy of NO in preventing neointimal hyperplasia in animal models of type II DM and metabolic syndrome and examine possible mechanisms for differences in outcomes. Aortic vascular smooth muscle cells (VSMC) were harvested from rodent models of type II DM (Zucker diabetic fatty), metabolic syndrome (obese Zucker), and their genetic control (lean Zucker). Interestingly, NO inhibited proliferation and induced G0/G1 cell cycle arrest to the greatest extent in VSMC from rodent models of metabolic syndrome and type II DM compared with controls. This heightened efficacy was associated with increased expression of cyclin-dependent kinase inhibitor p21, but not p27. Using the rat carotid artery injury model to assess the efficacy of NO in vivo, we found that the NO donor PROLI/NO inhibited neointimal hyperplasia to the greatest extent in type II DM rodents, followed by metabolic syndrome, then controls. Increased neointimal hyperplasia correlated with increased reactive oxygen species (ROS) production, as demonstrated by dihydroethidium staining, and NO inhibited this increase most in metabolic syndrome and DM. In conclusion, NO was surprisingly a more effective inhibitor of neointimal hyperplasia following arterial injury in type II DM and metabolic syndrome vs. control. This heightened efficacy may be secondary to greater inhibition of VSMC proliferation through cell cycle arrest and regulation of ROS expression, in addition to other possible unidentified mechanisms that deserve further exploration.

Topics: Animals; Aorta, Abdominal; Apoptosis; Carotid Artery Injuries; Cell Cycle; Cell Proliferation; Cells, Cultured; Diabetes Mellitus, Type 2; Disease Models, Animal; DNA Replication; Dose-Response Relationship, Drug; Hyperplasia; Male; Metabolic Syndrome; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Nitric Oxide; Nitric Oxide Donors; Proline; Rats; Rats, Zucker; Reactive Oxygen Species; S-Nitroso-N-Acetylpenicillamine; Triazenes; Tyrosine

We aimed to elucidate the effects of exogenous nitric oxide application via myrigotomized rat tympanic membranes on tympanosclerosis.. Forty male rats were included in the study. After myringotomy, nitric oxide was given to the left tympanic cavities and saline was given to the right as the control group. The procedure was repeated 3 times with weekly intervals. Three months after the first procedure, otomicroscopic examination was made and the rats were sacrificed for histologic examination.. Neither otomicroscopic examination, nor histologic examination of middle ears and tympanic membranes revealed any significant difference between the right and left sides. Tympanic membrane thicknesses were also similar on both sides without showing any significant difference.. Data from this study indicate that exogenous nitric oxide application did not change the occurence of tympanosclerosis in the rat model. Other cytokine interactions are needed to start the reaction sequence leading to tympanosclerosis.

Topics: Animals; Disease Models, Animal; Ear, Middle; Male; Microscopy; Nitric Oxide Donors; Otosclerosis; Rats; Rats, Sprague-Dawley; Triazenes; Tympanic Membrane

The impact of obesity on nitric oxide (NO)-mediated coronary microvascular responses is poorly understood. Thus NO-mediated vasomotor responses were investigated in pressurized coronary arterioles ( approximately 100 microm) isolated from lean (on normal diet) and obese (fed with 60% of saturated fat) rats. We found that dilations to acetylcholine (ACh) were not significantly different in obese and lean rats (lean, 83 +/- 4%; and obese, 85 +/- 3% at 1 microM), yet the inhibition of NO synthesis with N(omega)-nitro-l-arginine methyl ester reduced ACh-induced dilations only in vessels of lean controls. The presence of the soluble guanylate cyclase (sGC) inhibitor oxadiazolo-quinoxaline (ODQ) elicited a similar reduction in ACh-induced dilations in the two groups of vessels (lean, 60 +/- 11%; and obese, 57 +/- 3%). Dilations to NO donors, sodium nitroprusside (SNP), and diethylenetriamine (DETA)-NONOate were enhanced in coronary arterioles of obese compared with lean control rats (lean, 63 +/- 6% and 51 +/- 5%; and obese, 78 +/- 5% and 70 +/- 5%, respectively, at 1 microM), whereas dilations to 8-bromo-cGMP were not different in the two groups. In the presence of ODQ, both SNP and DETA-NONOate-induced dilations were reduced to a similar level in lean and obese rats. Moreover, SNP-stimulated cGMP immunoreactivity in coronary arterioles and also cGMP levels in carotid arteries were enhanced in obese rats, whereas the protein expression of endothelial NOS and the sGC beta1-subunit were not different in the two groups. Collectively, these findings suggest that in coronary arterioles of obese rats, the increased activity of sGC leads to an enhanced sensitivity to NO, which may contribute to the maintenance of NO-mediated dilations and coronary perfusion in obesity.

Topics: Acetylcholine; Adaptation, Physiological; Animals; Arterioles; Blotting, Western; Coronary Vessels; Cyclic GMP; Dietary Fats; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Activation; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Guanylate Cyclase; Immunohistochemistry; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Donors; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Nitroprusside; Nitroso Compounds; Obesity; Rats; Rats, Wistar; Receptors, Cytoplasmic and Nuclear; Soluble Guanylyl Cyclase; Vasodilation; Vasodilator Agents

Leukocyte activation, inflammatory up-regulation, and microcirculatory disruption associated with ischemia-reperfusion injury are hallmarks in the pathogenesis of acute pancreatitis (AP). NO donors ensure microvascular integrity, while glucocorticoids act as anti-inflammatory and immune modulator drugs. AP was induced by the biliopancreatic duct outlet exclusion-closed duodenal loops (BPDOE-CDLs) model. Treatment with hydrocortisone (6 mg/kg) or prednisolone (0.5 mg/kg) alone or together with DETA-NO (0.5 mg/kg) was done (a)1 hr pre or (b)1 hr post, or (c) 1 hr pre and 4 hr post ,or (d) 4 hr post triggering AP. NOS inhibition by L-NAME (15 mg/kg) and glucocorticoid receptor blockage by mifepristone (3 mg/kg) was considered. AP severity was assessed by biochemical and histopathological analyses. Treatment with glucocorticoids together with DETA-NO 1 hr pre and 4 hr post BPDOE-CDLs reduced serum amylase, lipase, C-reactive protein, IL-6, IL-10, hsp72, and 8-isoprostane as well as pancreatic and lung myeloperoxidase. Acinar and fat necrosis, hemorrhage, and neutrophil infiltrate were also decreased. Hydrocortisone together with DETA-NO rendered the best results. We conclude that AP severity was significantly diminished by glucocorticoids associated with DETA-NO, with the optimal dose and time point of administration being crucial to provide adequate protection against AP.

Topics: Animals; Biomarkers; Disease Models, Animal; DNA; Drug Therapy, Combination; Edema; Glucocorticoids; Hydrocortisone; Leukocyte Count; Male; Nitric Oxide Synthase; Pancreatitis; Prednisolone; Rats; Rats, Wistar; Severity of Illness Index; Time Factors; Triazenes

Nitric oxide (NO) is an important cell-signaling molecule, and its generators, nitric oxide synthases, are expressed temporospatially in fetal rat lung. Recently, NO has been reported to modulate branching of the fetal rat lung lobe in vitro. We designed this study to evaluate the effect of NO on the morphogenesis of hypoplastic lung using nitrofen-induced rat lung explant model.. A hypoplastic fetal lung model and a normal control lung model were induced by feeding a pregnant rat with nitrofen (100 mg) or olive oil on day 9.5 of gestation, respectively. Fetal lungs were harvested on day 13.5 and placed in organ culture containing serum-free medium Dulbecco modified Eagle medium. An NO donor, DETA NONOate (DETA/NO), was added daily in the culture medium. The lung cultures were divided into 4 groups: group 1 (n = 8), normal controls without DETA/NO; group 2 (n = 22), normal controls with DETA/NO; group 3 (n = 13), hypoplastic lungs without DETA/NO; group 4 (n = 22), hypoplastic lungs with DETA/NO. The fetal lungs were incubated for 48 hours at 37 degrees C with 5% CO2. Lung bud count and area of the specimens were measured under computer-assisted digital tracings. The rate of increase in bud count and lung area was calculated as the ratio of each value at 48 hours minus each value at 0 hour, divided by the value at 0 hour.. The lung bud count was significantly increased in group 2 compared with group 1 at a concentration of 50 micromol/L DETA/NO (P < .05). In the nitrofen group, the lung bud count was significantly increased in group 4 compared with group 3 at 100 micromol/L DETA/NO added (P < .05). There was no significant difference in the rate of increase in whole lung area among the 4 groups. The peak increase rates of lung area and bud count were significantly lower in group 4 compared with group 2.. This study demonstrates that the NO donor, DETA/NO, promotes branching of the nitrofen-induced hypoplastic fetal lung explant. These data suggest that NO may modulate the development of the nitrofen-induced hypoplastic lung.

Topics: Animals; Disease Models, Animal; Female; Fetal Organ Maturity; In Vitro Techniques; Lung; Lung Diseases; Nitric Oxide; Nitric Oxide Donors; Nitroso Compounds; Oxidation-Reduction; Pesticides; Phenyl Ethers; Pregnancy; Rats; Rats, Sprague-Dawley; Signal Transduction; Teratogens

Liver ischaemia-reperfusion (I/R) occurs during resuscitation from haemorrhagic shock, hepatic transplantation and anatomic resection of the liver. This injury is associated with hepatocellular enzyme release and hepatocyte necrosis. The impact of chronic illnesses such as diabetes mellitus (DM) on hepatic I/R is unknown. This study determines the effect of DM on liver I/R using a murine model of type II DM in which the leptin receptor is defective. Preliminary studies suggest that animal models of DM have impaired endothelial nitric oxide (NO) release. Other studies suggest that NO attenuates hepatic I/R in phenotypically normal animals. We postulated that DM exacerbates hepatic I/R and that exogenous NO administration will attenuate hepatocellular injury.. Non-diabetic and diabetic (db/db) mice were anaesthetized and underwent laparotomy with the placement of a microvascular clip on the hepatic artery and portal vein supplying the medial and left lateral lobes of the liver rendering about 70% of the liver ischaemic. Hepatic ischaemia was maintained for 45 min after which time the clip was removed and the liver segments reperfused. The abdomen was closed and the animals maintained for 5 h of reperfusion. Hepatic injury was then assessed by measuring serum alanine and aspartate transaminases (ALT, AST) spectrophotometrically. Sections of liver reperfused for 24 h were stained with haematoxylin and eosin and the percentage of hepatocyte necrosis evaluated using morphometric techniques. Other animals undergoing hepatic I/R received the NO donor (DETA 100 micro g/kg, i.v. 5 min prior to reperfusion). Time-matched, sham-operated animals served as controls. The data are expressed as mean +/- SEM and analysed by ANOVA.. Serum AST and ALT levels were significantly higher in db/db animals vs. non-diabetics, even in the absence of hepatic I/R (P < 0.01). Serum AST and ALT levels in db/db mice undergoing hepatic I/R were nearly five times greater than that of non-diabetic animals (P < 0.01). Histologic examination of the livers of the diabetic animals undergoing I/R demonstrated significantly greater hepatocellular necrosis (zone III; 30-40%) when compared with non-diabetic animals sustaining the same injury (zone III; 3-10%). The NO donor DETA totally prevented the increase in serum ALT and AST release associated with I/R in both the diabetic and non-diabetic mice when compared with animals not receiving this agent (P < 0.01).. This is the first study suggesting that DM exacerbates hepatic I/R and that NO donors will prevent this hepatocellular injury in the diabetic. Sixteen million Americans have DM. Understanding the effect of this chronic illness on the inflammatory response to injury is essential to improving clinical outcomes in these medically compromised patients.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Disease Models, Animal; Liver; Mice; Mice, Inbred C57BL; Mice, Inbred NOD; Nitric Oxide Donors; Nitroso Compounds; Reperfusion Injury

Vasospasm after subarachnoid hemorrhage (SAH) may result from hemoglobin-mediated removal of nitric oxide (NO) from the arterial wall. We tested the ability of the long-acting, water-soluble, NO donor (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-1,2-diolate (DETA/NO), delivered via continuous intracisternal infusion, to prevent vasospasm in a nonhuman primate model of SAH.. First, vasorelaxation in response to DETA/NO was characterized in vitro by using monkey basilar artery rings under isometric tension. Next, monkeys were randomized to undergo angiography, unilateral SAH, and no treatment (SAH only, n = 4) or treatment with DETA/NO (1 mmol/L, 12 ml/d, n = 4) or decomposed DETA/NO (at the same dose, n = 4). Vasospasm was assessed by angiography, which was performed on Day 0 and Day 7. Levels of cyclic adenosine monophosphate and cyclic guanosine monophosphate (cGMP) were measured in cerebral arteries on Day 7.. DETA/NO produced significant relaxation of monkey arteries in vitro, which reached a maximum at concentrations of 10(-5) mol/L. In monkeys, angiography demonstrated significant vasospasm of the right intradural cerebral arteries in all three groups, with no significant difference in vasospasm among the groups (P > 0.05, analysis of variance). The ratios of cGMP or cyclic adenosine monophosphate levels in the right and left middle cerebral arteries were not different among the groups (P > 0.05, analysis of variance). There was no significant correlation between arterial cGMP contents and the severity of vasospasm.. DETA/NO did not prevent vasospasm. There was no correlation between the severity of vasospasm and cyclic adenosine monophosphate and cGMP levels in the cerebral arteries. These results suggest that events downstream of cyclic nucleotides may be abnormal during vasospasm.

Topics: Animals; Cerebral Angiography; Cyclic AMP; Cyclic GMP; Disease Models, Animal; Macaca fascicularis; Middle Cerebral Artery; Nitric Oxide Donors; Severity of Illness Index; Subarachnoid Hemorrhage; Triazenes; Vasospasm, Intracranial

We investigated the effects of NO on angiogenesis and the synthesis of vascular endothelial growth factor (VEGF) in a model of focal embolic cerebral ischemia in the rat. Compared with control rats, systemic administration of an NO donor, DETANONOate, to rats 24 hours after stroke significantly enlarged vascular perimeters and increased the number of proliferated cerebral endothelial cells and the numbers of newly generated vessels in the ischemic boundary regions, as evaluated by 3-dimensional laser scanning confocal microscopy. Treatment with DETANONOate significantly increased VEGF levels in the ischemic boundary regions as measured by ELISA. A capillary-like tube formation assay was used to investigate whether DETANONOate increases angiogenesis in ischemic brain via activation of soluble guanylate cyclase. DETANONOate-induced capillary-like tube formation was completely inhibited by a soluble guanylate cyclase inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ). Blocking VEGF activity by a neutralized antibody against VEGF receptor 2 significantly attenuated DETANONOate-induced capillary-like tube formation. Moreover, systemic administration of a phosphodiesterase type 5 inhibitor (Sildenafil) to rats 24 hours after stroke significantly increased angiogenesis in the ischemic boundary regions. Sildenafil and an analog of cyclic guanosine monophosphate (cGMP) also induced capillary-like tube formation. These findings suggest that exogenous NO enhances angiogenesis in ischemic brain, which is mediated by the NO/cGMP pathway. Furthermore, our data suggest that NO, in part via VEGF, may enhance angiogenesis in ischemic brain.

Topics: 3',5'-Cyclic-GMP Phosphodiesterases; Animals; Brain; Bromodeoxyuridine; Cell Division; Cyclic GMP; Cyclic Nucleotide Phosphodiesterases, Type 5; Disease Models, Animal; Endothelial Growth Factors; Endothelium, Vascular; Enzyme Inhibitors; Guanylate Cyclase; Intercellular Signaling Peptides and Proteins; Lymphokines; Male; Neovascularization, Physiologic; Nitric Oxide; Nitric Oxide Donors; Nitroso Compounds; Piperazines; Purines; Rats; Rats, Wistar; Receptors, Cytoplasmic and Nuclear; Sildenafil Citrate; Soluble Guanylyl Cyclase; Stroke; Sulfones; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2; Vascular Endothelial Growth Factors

Neurogenesis, which is upregulated by neural injury in the adult mammalian brain, may be involved in the repair of the injured brain and functional recovery. Therefore, the authors sought to identify agents that can enhance neurogenesis after brain injury, and they report that (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)amino]diazen-1-ium-1,2-diolate (DETA/NONOate), a nitric oxide donor, upregulates neurogenesis and reduces functional deficits after traumatic brain injury (TBI) in rats.. The agent DETA/NONOate (0.4 mg/kg) was injected intraperitoneally into 16 rats daily for 7 days, starting 1 day after TBI induced by controlled cortical impact. Bromodeoxyuridine (100 mg/kg) was also injected intraperitoneally daily for 14 days after TBI to label the newly generated cells in the brain. A neurological functional evaluation was performed in all rats and the animals were killed at 14 or 42 days postinjury. Immunohistochemical staining was used to identify proliferating cells.. Compared with control rats, the proliferation, survival, migration and differentiation of neural progenitor cells were all significantly enhanced in the hippocampus, subventricular zone, striatum, corpus callosum, and the boundary zone of the injured cortex, as well as in the contralateral hemisphere in rats with TBI that received DETA/ NONOate treatment. Neurological functional outcomes in the DETA/NONOate-treated group were also significantly improved compared with the untreated group. These data indicate that DETA/NONOate may be useful in the treatment of TBI.

Topics: Animals; Brain Injuries; Bromodeoxyuridine; Cell Division; Cell Movement; Cerebral Cortex; Corpus Callosum; Corpus Striatum; Disease Models, Animal; Drug Administration Schedule; Hippocampus; Injections, Intraperitoneal; Male; Nitric Oxide Donors; Nitroso Compounds; Psychomotor Disorders; Radiation-Sensitizing Agents; Random Allocation; Rats; Rats, Wistar; Recovery of Function; Stem Cells; Time Factors; Up-Regulation

Endogenous and exogenous nitric oxide (NO) may have important antibacterial effects in patients with pneumonia. NO administration has been limited to the continuous inhalation of gas-phase NO (ie, inhaled NO [iNO]). Intermittent nebulization of NONOates, novel NO donors, may permit the continuous intrapulmonary delivery of NO. Thus, we assessed the effects of nebulized diethylenetetraamine-NONOate (DETA-NO) in a model of acute Pseudomonas aeruginosa pneumonia.. Randomized, controlled study.. Male C57Bl/6 mice.. Pneumonia was induced by intratracheal instillation of P aeruginosa (3 x 10(7) CFU in 50 microL). Pneumonia and sham mice were randomized to receive no treatment, nebulized DETA-NO (12.5 or 125 micromol) at 4 h and 12 h, or continuous iNO for 24 h (10 or 40 ppm) until they were killed at 24 h.. The nebulization of DETA-NO was associated with a marked increase in mean (+/- SEM) exhaled NO levels (after nebulization, 484 +/- 34 parts per billion [ppb]; baseline, 13.4 +/- 0.4 ppb; p < 0.01) and plasma levels of nitrites/nitrates (after nebulization, 73 +/- 28 microM; at baseline, 14 +/- 3 microM; p < 0.05). Nebulized DETA-NO decreased the pulmonary bacterial load in mice with pneumonia by 65 +/- 19% (p < 0.05 vs untreated mice) but had no effect on pulmonary leukocyte infiltration. Although the growth of P aeruginosa colonies in vitro was impaired on exposure to DETA-NO, growth was similarly impaired by exposure to DETA nucleophile/backbone alone.. The nebulization of DETA-NO provides a method for the prolonged intrapulmonary delivery of NO. The antibacterial effect of DETA-NO in vivo and in vitro is due, in large part, to the DETA nucleophile moiety and is independent of NO, suggesting a limited therapeutic role for exogenous NO in pneumonia.

Topics: Acute Disease; Aerosols; Animals; Disease Models, Animal; Hydrazines; Lung; Male; Mice; Mice, Inbred C57BL; Nitrates; Nitric Oxide; Nitrites; Nitroso Compounds; Pneumonia, Bacterial; Pseudomonas aeruginosa; Pseudomonas Infections; Random Allocation

The adult rodent brain is capable of generating neuronal progenitor cells in the subventricular zone, and in the dentate gyrus of the hippocampus, throughout the life of the animal. Signals that regulate progenitor cell proliferation, differentiation, and migration are not well known. We report that administration of a nitric oxide donor, (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl) aminio]diazen-1-ium-1,2-diolate (DETA/NONOate), to young adult rats significantly increases cell proliferation and migration in the subventricular zone and the dentate gyrus. Treatment with DETA/ NONOate also increases neurogenesis in the dentate gyrus. Furthermore, administration of DETA/NONOate to rats subjected to embolic middle cerebral artery occlusion significantly increases cell proliferation and migration in the subventricular zone and the dentate gyrus, and these rats exhibit significant improvements of neurological outcome during recovery from ischemic stroke. Administration of DETA/NONOate significantly increases cortical levels of guanosine monophosphate both in ischemic and nonischemic rats, supporting the role of nitric oxide in promoting cell proliferation and neurogenesis. Thus, our data indicate that nitric oxide is involved in the regulation of progenitor cells and neurogenesis in the adult brain. This suggests that nitric oxide delivered to the brain well after stroke may have therapeutic benefits.

Topics: Animals; Bromodeoxyuridine; Cell Division; Cell Movement; Cerebral Cortex; Cyclic GMP; Dentate Gyrus; Disease Models, Animal; Infarction, Middle Cerebral Artery; Lateral Ventricles; Male; Neurons; Nitric Oxide Donors; Nitroso Compounds; Rats; Rats, Wistar; Stroke; Treatment Outcome

Topics: Animals; Apoptosis; Disease Models, Animal; Drug Evaluation, Preclinical; Endothelium, Vascular; Heat-Shock Proteins; HSP72 Heat-Shock Proteins; Nitric Oxide; Stress, Physiological; Swine; Triazenes

Intrathecal bolus administration of (Z)-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)aminio]diazen++ +-1-ium-1,2-diolate (DETA/NO), a long half-life diazeniumdiolate-class nitric oxide (NO) donor, was evaluated for safety and efficacy in the treatment of delayed cerebral vasospasm in a canine model of subarachnoid hemorrhage (SAH).. The baseline basilar artery (BA) diameter of 25 dogs was measured with the aid of angiography on Day 0. Vasospasm was then induced by intracisternal injection of autologous arterial blood on Days 0 and 2. Repeated arteriography on Day 7 revealed an average BA diameter of 58% of baseline. Each dog was then randomized to one of four groups: a pathology control group (SAH only, four animals); a treatment control group (SAH plus 2 micromol of the inactive drug carrier DETA, eight animals); a low-dose treatment group (SAH plus 0.2 micromol DETA/NO, six animals); or a high-dose treatment group (SAH plus 2 micromol DETA/NO, six animals). The drugs were administered in a 2-ml intrathecal bolus via the cisterna magna. Arterial caliber was monitored by angiography over the subsequent 4 hours. A 2-micromol dose of the drug was then given and serial arteriography continued for an additional hour to screen for tachyphylaxis. Intracranial pressure and respiratory and hemodynamic parameters were continuously monitored. Histopathological analyses of the animals' brains were performed after the dogs were killed on Day 8. The drug DETA/NO produced reversal of vasospasm in a dose-dependent fashion that roughly followed a double exponential time course. Doses of 2 micromol DETA/NO resulted in restoration of the angiographically monitored BA diameter to the prevasospasm size at 1.5 hours posttreatment, and this was sustained at 88% of baseline at 4 hours (p < 0.01, independent samples t-test). By contrast, the treatment control group remained on average at 54% of baseline diameter. The low-dose treatment group achieved only partial and more transitory relaxation. Histopathological analyses showed findings consistent with chronic SAH but did not demonstrate any toxicity associated with the NO donor. No adverse physiological changes were seen.. This study indicates that long-acting NO donors are potentially useful as agents to restore circulation in patients suffering from cerebral vasospasm.

Topics: Analysis of Variance; Animals; Basilar Artery; Blood Pressure; Brain; Central Venous Pressure; Cerebral Angiography; Cisterna Magna; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Drug Carriers; Heart Rate; Image Processing, Computer-Assisted; Injections, Spinal; Intracranial Pressure; Ischemic Attack, Transient; Nitric Oxide; Random Allocation; Respiration; Safety; Subarachnoid Hemorrhage; Tachyphylaxis; Triazenes; Vasodilator Agents