nitroarginine and Thrombosis

There is increasing evidence that Chlamydia pneumoniae is linked to atherosclerosis and thrombosis. In this regard, we have recently shown that C. pneumoniae stimulates platelet aggregation and secretion, which may play an important role in the progress of atherosclerosis and in thrombotic vascular occlusion. The aims of the present study were to investigate the effects of C. pneumoniae on platelet-mediated formation of reactive oxygen species (ROS) and oxidation of low-density lipoprotein (LDL) in vitro. ROS production was registered as changes in 2',7'-dichlorofluorescin- fluorescence in platelets with flow cytometry. LDL-oxidation was determined by measuring thiobarbituric acid reactive substances (TBARs). We found that C. pneumoniae stimulated platelet production of ROS. Polymyxin B treatment of C. pneumoniae, but not elevated temperature, abolished the stimulatory effects on platelet ROS-production, which suggests that chlamydial lipopolysaccharide has an important role. Inhibition of nitric oxide synthase with nitro-L-arginine, lipoxygenase with 5,8,11-eicosatriynoic acid and protein kinase C with GF 109203X significantly lowered the production of radicals. In contrast, inhibition of NADPH-oxidase with di-phenyleneiodonium (DPI) did not affect the C. pneumoniae induced ROS-production. These findings suggest that the activities of nitric oxide synthase and lipoxygenase are the sources for ROS and that the generation is dependent of the activity of protein kinase C. The C. pneumoniae-induced ROS-production in platelets was associated with an extensive oxidation of LDL, which was significantly higher compared to the effect obtained by separate exposure of LDL to C. pneumoniae or platelets. In conclusion, C. pneumoniae interaction with platelets leading to aggregation, ROS-production and oxidative damage on LDL, may play a crucial role in the development of atherosclerotic cardiovascular disease.

Topics: Atherosclerosis; Blood Platelets; Cells, Cultured; Chlamydophila pneumoniae; Enzyme Inhibitors; Fatty Acids, Unsaturated; Flow Cytometry; Fluoresceins; Humans; Indoles; Lipopolysaccharides; Lipoproteins, LDL; Lipoxygenase; Lipoxygenase Inhibitors; Maleimides; Nitric Oxide Synthase; Nitroarginine; Oxygen; Polymerase Chain Reaction; Polymyxin B; Protein Kinase C; Reactive Oxygen Species; Signal Transduction; Temperature; Thiobarbituric Acid Reactive Substances; Thrombosis

Effects of DX-9065a ((+)-2S-2-[4-[[(3S)-1-acetimidoyl-3-pyrrolidinyl]oxy]phenyl]-3-[7-amidino-2-naphthyl]propanoic acid hydrochloride pentahydrate), a dibasic propanoic acid and an inhibitor of factor Xa, were compared with those of argatroban, a low molecular weight thrombin inhibitor, on the ellagic acid-induced plantar skin thrombosis in the rats treated with tetrodotoxin and N(omega)-nitro-L-arginine. Plantar skin blood flow was measured with laser Doppler flow meters, and skin temperature of the hindlimb was monitored simultaneously. In order to induce thrombus in plantar skin vasculature, ellagic acid (300 microg, i.a.) was injected into a branch of femoral artery. The formation of thrombus in femoral and plantar vessels was assessed by light microscopy. Ellagic acid decreased plantar skin blood flow and skin temperature. Intravenous injections of DX-9065a (3 mg/kg) and argatroban (1-3 mg/kg) significantly inhibited the ellagic acid-induced disturbance of plantar skin blood flow and lowering skin temperature without affecting bleeding time. The oral administration of DX-9065a (30-100 mg/kg) significantly prevented the decrease in skin temperature induced by ellagic acid, but it partially inhibited the disturbance of plantar skin blood flow. DX-9065a and argatroban also prolonged prothrombin time in a dose-dependent manner. These results suggest that DX-9065a effectively prevented thrombosis produced by ellagic acid in the skin circulation without a risk of bleeding.

Topics: Animals; Dose-Response Relationship, Drug; Ellagic Acid; Factor Xa Inhibitors; Hindlimb; Male; Naphthalenes; Nitroarginine; Propionates; Rats; Rats, Wistar; Skin; Skin Diseases; Tetrodotoxin; Thrombosis

We studied the antithrombotic activity of 2-acetoxybenzoate 2-[1-nitroxy-methyl]-phenyl ester (NCX 4016), a novel nitric oxide (NO)-releasing aspirin derivative, in vivo in different animal models of platelet-dependent and independent pulmonary thromboembolism and compared it with that of aspirin. NCX 4016 protected mice from death induced by the intravenous (i.v.) injection of collagen plus epinephrine, of 9,11-dideoxy-11alpha, 9alpha-epoxymethano-prostaglandin F(2alpha) (U46619) and of thrombin while aspirin was only active against collagen plus epinephrine. The drop in platelet count and number of lung emboli were reduced by NCX 4016 more effectively than aspirin. NCX 4016 protected mice also from mechanical pulmonary embolism (i.v. injection of hardened rat red blood cells) while aspirin was ineffective. In rabbits, NCX 4016 significantly reduced the accumulation of [111In]oxine-labeled platelets in the pulmonary vasculature induced by collagen and by thrombin while aspirin produced reductions which were significant only versus collagen. In conclusion, NCX 4016 exerts a more pronounced antithrombotic activity than aspirin in vivo in two different animal species, largely due to a deeper inhibitory effect on platelets. NCX 4016 may represent a better antithrombotic agent than aspirin.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Aspirin; Disease Models, Animal; Dose-Response Relationship, Drug; Injections, Intravenous; Lung; Male; Mice; Nitroarginine; Platelet Aggregation Inhibitors; Platelet Count; Pulmonary Artery; Pulmonary Embolism; Rabbits; Thrombin; Thrombosis

Topics: Animals; Arginine; Aspirin; Blood Pressure; Cats; Epoprostenol; Extracorporeal Circulation; Fibrinolysis; Heparitin Sulfate; Iloprost; Infusions, Intravenous; Injections, Intravenous; Molsidomine; Nitric Oxide; Nitroarginine; Platelet Aggregation; Streptokinase; Thrombosis; Vasodilator Agents

Inhibition of an endothelium-derived relaxing factor (EDRF) may contribute to the pathogenesis of thrombotic arterial occlusions.. We measured the blood pressure and urinary excretion of protein, sodium, and potassium and histologically examined the brains, hearts, and kidneys in normotensive Wistar Kyoto rats (WKY) and stroke-prone spontaneously hypertensive rats (SHRSP) fed on a diet containing: (a) EDRF inhibitor (L-N-nitroarginine:L-NNA); (b) L-arginine, which reverses the effect of L-NNA; or (c) both L-NNA and L-arginine for 1 to 8 weeks. In addition, we examined L-NNA-treated SHRSP, the blood pressures of which were lowered using hydralazine. Furthermore, we produced and examined Goldblatt's renal hypertensive rats, which are of a different type from those resulting from the L-NNA treatment.. Both WKY and SHRSP rats fed on a diet containing L-NNA suffered from hypertension and cerebral infarctions in a dose-dependent manner. Cerebral infarctions occurred whether or not SHRSP rats were treated with an antihypertensive agent when they were fed a high dosage of L-NNA. In contrast, SHRSP rats, treated simultaneously with both L-NNA and L-arginine, suffered few cerebral infarctions, although they were severely hypertensive. In addition, there were no cerebral infarctions in Goldblatt's renal hypertensive rats, although they suffered from advanced hypertension.. The data indicate that the inhibition of EDRF injures the vessel walls and encourages platelet adhesion to the damaged areas. The adhering platelets narrow the lumen with resultant thrombotic arterial occlusions. Pathophysiologic conditions that decrease EDRF synthesis appear to play an important role in cerebral, renal, and myocardial infarctions.

Topics: Animals; Arginine; Arterial Occlusive Diseases; Blood Pressure; Blood Vessels; Cerebrovascular Disorders; Dose-Response Relationship, Drug; Hydralazine; Hypertension, Renovascular; Incidence; Microscopy, Electron; Nitric Oxide; Nitroarginine; Potassium; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Sodium; Thrombosis; Time Factors; Vascular Diseases

Using a novel method of measuring thrombogenesis on the surface of rabbit aorta endothelium superfused with whole blood, it was demonstrated that the thrombogenic properties of endothelium are potentiated by pretreatment with inhibitors of nitric oxide (NO) synthase (NG-monomethyl-L-arginine, MeArg, 100 mcM) or cyclooxygenase (aspirin, ASA, 60 mcM). Since MeArg is less effective than aspirin and moreover does not influence thrombogenesis after previous inhibition of the synthesis of PGI2, it is concluded that the generation of NO by vascular endothelium complements PGI2 in its antithrombotic activity but NO may be ineffective when the generation of PGI2 has been impaired.

Topics: Animals; Aorta, Thoracic; Arginine; Aspirin; Blood Platelets; Endothelium, Vascular; Epoprostenol; In Vitro Techniques; Nitric Oxide; Nitroarginine; Rabbits; Thrombosis