3-nitrotyrosine and Nervous-System-Diseases

Trans-sodium crocetinate (TSC) is a novel synthetic carotenoid compound that improves diffusion of small molecules, including oxygen, in solutions. TSC provides neuroprotection in healthy rats and rabbits. This study seeks to determine whether TSC is neuroprotective in obese mice. Sixteen-week-old CD-1 male mice that had been fed a high-fat diet for 10 weeks were subjected to a 90-min middle cerebral arterial occlusion (MCAO). They received TSC by two boluses through a tail vein 10 min after the onset of MCAO and reperfusion, respectively, with doses of 0.14, 0.28, and 0.7 mg/kg or by a bolus-infusion-bolus strategy with a dose of 0.14 mg/kg during MCAO. The neurological outcome was evaluated 72 hr after MCAO. Brain tissues were harvested 24 hr after MCAO to measure nitrotyrosine-containing proteins, 4-hydroxy-2-nonenal, matrix metalloproteinase (MMP)-2 and -9 activity and expression, and inflammatory cytokines. TSC given in the two-bolus strategy did not improve the neurological outcome. The bolus-infusion-bolus strategy significantly reduced brain edema, infarct volume, and hemorrhagic transformation and improved neurological functions. TSC reduced nitrotyrosine-containing proteins, MMP-9 activity and expression, and inflammatory cytokines in ischemic brain tissues. Our results indicate that TSC delivered by the bolus-infusion-bolus strategy provides neuroprotection in obese mice. This protection may occur through reduction of oxidative stress, MMP-9 activity, or inflammatory cytokines in the ischemic brain tissues.

Topics: Aldehydes; Analysis of Variance; Animals; Brain; Brain Ischemia; Carotenoids; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Male; Matrix Metalloproteinases; Mice; Nervous System Diseases; Neuroprotective Agents; Obesity; Oxidative Stress; Reperfusion; Tyrosine; Vitamin A

Substantial experimental evidence supports that reactive species mediate secondary damage after traumatic spinal cord injury (SCI) by inducing oxidative stress. Removal of reactive species may reduce secondary damage following SCI. This study explored the effectiveness of a catalytic antioxidant - Mn (III) tetrakis (4-benzoic acid) porphyrin (MnTBAP) - in removing reactive oxygen species (ROS), reducing oxidative stress, and improving functional recovery in vivo in a rat impact SCI model. The efficiency of MnTBAP was also compared with that of methylprednisolone - the only drug used clinically in treating acute SCI.. In vivo measurements of time courses of ROS production by microdialysis and microcannula sampling in MnTBAP, methylprednisolone, and saline (as vehicle control)-treated SCI rats showed that both agents significantly reduced the production of hydrogen peroxide, but only MnTBAP significantly reduced superoxide elevation after SCI. In vitro experiments further demonstrated that MnTBAP scavenged both of the preceding ROS, whereas methylprednisolone had no effect on either. By counting the immuno-positive neurons in the spinal cord sections immunohistochemically stained with anti-nitrotyrosine and anti-4-hydroxy-nonenal antibodies as the markers of protein nitration and membrane lipid peroxidation, we demonstrated that MnTBAP significantly reduced the numbers of 4-hydroxy-nonenal-positive and nitrotyrosine-positive neurons in the sections at 1.55 to 2.55 mm and 1.1 to 3.1 mm, respectively, rostral to the injury epicenter compared to the vehicle-treated animals. By behavioral tests (open field and inclined plane tests), we demonstrated that at 4 hours post-SCI treatment with MnTBAP and the standard methylprednisolone regimen both significantly increased test scores compared to those produced by vehicle treatment. However, the outcomes for MnTBAP-treated rats were significantly better than those for methylprednisolone-treated animals.. This study demonstrated for the first time in vivo and in vitro that MnTBAP significantly reduced the levels of SCI-elevated ROS and that MnTBAP is superior to methylprednisolone in removing ROS. Removal of ROS by MnTBAP significantly reduced protein nitration and membrane lipid peroxidation in neurons. MnTBAP more effectively reduced neurological deficits than did methylprednisolone after SCI - the first most important criterion for assessing SCI treatments. These results support the therapeutic potential of MnTBAP in treating SCI.

Topics: Aldehydes; Analysis of Variance; Animals; Blood-Brain Barrier; Cell Count; Disease Models, Animal; Hydrogen Peroxide; Locomotion; Male; Metalloporphyrins; Methylprednisolone; Microdialysis; Nervous System Diseases; Neuroprotective Agents; Oxidative Stress; Psychomotor Performance; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Recovery of Function; Spinal Cord Injuries; Time Factors; Tyrosine

Cerebral ischemia exacerbates neuronal death and neurological dysfunction. Evidence supports the involvement of oxidative/nitrative stress in the pathophysiology of cerebral ischemia. Heme oxygenase-1 (HO-1) is a rate-limiting enzyme in heme catabolism, possessing potent anti-oxidant and anti-apoptosis effects. In transgenic mice, HO-1 overproduction is neuroprotective against cerebral ischemia injury, but by unclear mechanisms. The present study determined whether treatment with adenoviral vector overexpressing HO-1 (Ad-HO-1) attenuates post-ischemic brain damage via reduction of oxidative/nitrative stress. After focal cerebral ischemia, Ad-HO-1 reduced lipid peroxidation and protein nitration, decreased infarct volume, and attenuated neurologic deficits. Zinc protoporphyrin IX (ZnPP IX, a specific HO-1 inhibitor) blocked Ad-HO-1 mediated effects against ischemic brain damage. Although Ad-HO-1 slightly reduced ischemic brain NO concentrations, Ad-HO-1 treatment significantly inhibited cerebral expression of iNOS protein expression, without significant effect upon nNOS or eNOS expression compared to vehicle after focal cerebral ischemia. Ad-HO-1 preserved NO bioavailability by increasing eNOS phosphorylation during ischemia compared to vehicle. Together, our results suggest that Ad-HO-1 attenuates post-ischemic brain damage via simultaneous reduction of oxidative/nitrative stress and preservation of NO bioavailability.

Topics: Animals; Blotting, Western; Brain Ischemia; Dependovirus; Genetic Therapy; Genetic Vectors; Heme Oxygenase-1; Infarction, Middle Cerebral Artery; Male; Nervous System Diseases; Nitric Oxide; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Phosphorylation; Rats; Rats, Sprague-Dawley; Reactive Nitrogen Species; Reactive Oxygen Species; Superoxides; Tyrosine; Up-Regulation

In this study, we evaluated the time course of changes in inducible nitric oxide synthase (iNOS) in the brain by using the rat model of sepsis induced by cecal ligation and puncture (CLP) and examined whether selective iNOS inhibition can prevent the hemodynamic and neurological changes induced by sepsis. Male Wistar rats were randomly divided into four groups: control, sham, CLP, and CLP + the selective iNOS inhibitor L-N6-(1-iminoethyl)-lysine (L-NIL). Septic shock was induced in the rats by CLP under pentobarbital anesthesia, and then we measured hemodynamic variables, neurological indicators, blood gases, plasma levels of nitrate/nitrite (an indicator of the biosynthesis of NO), and brain iNOS activity and nitrotyrosine levels after 1, 6, 12, and 24 h. Plasma nitrite was increased at 12 and 24 h in the CLP group. The activity of iNOS in the brain was increased at 12 and 24 h after CLP (at 12 h: control, 0.3 +/- 0.05; sham, 0.3 +/- 0.1; CLP, 1.3 +/- 0.08*; CLP + L-NIL, 0.33 +/- 0.1 fmol x mg(-1) x min(-1); at 24 h: control, 0.27 +/- 0.08; sham, 0.31 +/- 0.1; CLP, 1.0 +/- 0.3*; CLP + L-NIL, 0.34 +/- 0.1 fmol x mg(-1) x min(-1); mean +/- SD; *P < 0.05). Brain nitrotyrosine was increased at 24 h after CLP (at 24 h: control, 6.7 +/- 0.4; sham, 6.7 +/- 0.5; CLP, 11.2 +/- 2.8*; CLP + L-NIL, 7.52 +/- 0.5 densitometric units; means +/- SD; *P < 0.01). In contrast, in both the CLP and CLP + L-NIL groups, the consciousness reflex was significantly decreased at 24 h after CLP. Selective iNOS inhibition restored the hemodynamic changes induced by sepsis but could not improve neurological dysfunction.

Topics: Animals; Blood Pressure; Blotting, Western; Calcium; Cecum; Cerebral Cortex; Enzyme Inhibitors; Heart Rate; Hemodynamics; Interleukin-1; Lysine; Male; Nervous System Diseases; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rats; Rats, Wistar; Shock, Septic; Tumor Necrosis Factor-alpha; Tyrosine

Acute inflammatory reactions cause neuronal damage in cerebral ischemia-reperfusion. Urinary trypsin inhibitor (UTI), a serine protease inhibitor, is cytoprotective against ischemia-reperfusion injury in the liver, intestine, kidney, heart, and lung through its antiinflammatory activity. Neuroprotective action of UTI on transient global cerebral ischemia has been documented. This is the first study to determine whether UTI is neuroprotective against transient focal cerebral ischemia.. Adult male Wistar rats were randomly assigned to the following treatment groups: 0.9% saline (control, n = 9); 100,000 U/kg UTI (n = 9); and 300,000 U/kg UTI (n = 9). Treatments were performed intravenously 10 min before right middle cerebral artery occlusion for 2 h and subsequent reperfusion. Ninety-six hours after the onset of reperfusion, the motor neurologic deficit and the cerebral infarct size were evaluated. Furthermore, immunohistochemical staining for myeloperoxidase and nitrotyrosine to count infiltrating neutrophils and nitrated cells, respectively, was performed on the brain sections.. Infarct volume in the 300,000 U/kg UTI group was smaller than in the 100,000 U/kg UTI and saline control groups (P < 0.05). Treatment with 300,000 U/kg UTI showed a trend to improve neurologic outcome but did not reach statistical significance (P = 0.0693). The significant decrease in neutrophil infiltration was observed in the ischemic hemisphere treated with 300,000 U/kg UTI compared with saline control (P < 0.05). Nitrotyrosine deposition in the ischemic hemisphere was significantly reduced in the 300,000 U/kg UTI group compared with saline control and 100,000 U/kg UTI groups (P < 0.05).. Intravenous pretreatment with 300,000 U/kg UTI reduces focal ischemia-reperfusion injury in the rat brain, potentially opening a novel therapeutic avenue for the treatment of cerebral ischemia.

Topics: Animals; Behavior, Animal; Brain; Cell Count; Cerebrovascular Circulation; Glycoproteins; Hypoxia-Ischemia, Brain; Immunohistochemistry; Infarction, Middle Cerebral Artery; Laser-Doppler Flowmetry; Male; Nervous System Diseases; Neuroprotective Agents; Neutrophils; Peroxidase; Rats; Rats, Wistar; Reperfusion Injury; Trypsin Inhibitors; Tyrosine

All processes of oxygen activation include very reactive intermediates. Therefore, aerobic cells must cope with--and to some extent also adapt to--oxidative stress provoked for example by infections or intoxications, where these reactive intermediates accumulate. All inflammatory processes include such oxygen activating processes where reactive oxygen species (ROS) are produced. Dependent on the strength of these impact(s), several symptoms indicate the deviation from normal, steady-state metabolism. Intrinsic radical scavenging processes or compounds administered with food thus have to warrant metabolic control within certain limits. Antioxidants which in many cases are free radical scavengers or quenchers of activated states comprise a vast number of classes of organic molecules including phenolics as the most prominent ones. In this publication the activities of extracts from Fraxinus excelsior, Populus tremula and Solidago virgaurea as components of the drug Phytodolor and their mechanisms of protection from oxidative damage are summarized. In addition, new results on tyrosine nitration, a process characteristic for sites of inflammation, and its inhibition by these plant extracts, is reported.

Topics: Animals; Antioxidants; Cells, Cultured; Chromatography, High Pressure Liquid; Enzymes; Free Radical Scavengers; Humans; Indicators and Reagents; Inflammation; Inflammatory Bowel Diseases; Lipoproteins, LDL; Lipoxygenase; Models, Biological; Nervous System Diseases; Oxidative Stress; Peroxidase; Plant Extracts; Plants, Medicinal; Respiratory Burst; Respiratory Tract Diseases; Tyrosine; Xanthine Oxidase

Human African trypanosomiasis, or sleeping sickness, evolves toward a meningoencephalitic stage, with a breakage in the blood-brain barrier, perivascular infiltrates, and astrocytosis. The involvement of nitric oxide (NO) has been evoked in the pathogenic development of the illness, since NO was found to be increased in the brain of animals infected with Trypanosoma brucei (T. b.) brucei. An excessive NO production can lead to alterations of neuronal signaling and to cell damage through the cytotoxicity of NO and its derivatives, especially peroxynitrites. In African trypanosomiasis, the sites of NO production and its role in the pathogenicity of lesions in the central nervous system (CNS) are unknown. In a chronic model of African trypanosomiasis (mice infected with T. b. brucei surviving with episodic suramin administration), NADPH-diaphorase staining of brain slides revealed that NO synthase (NOS) activity is located not only in endothelial cells, choroid plexus ependymal cells, and neurons as in control mice but also in mononuclear inflammatory cells located in perivascular and parenchyma infiltrates. An immunohistochemical study showed that the mononuclear inflammatory cells expressed an inducible NOS activity. Furthermore, the presence of nitrotyrosine in inflammatory lesions demonstrated an increased NO production and the intermediate formation of peroxynitrites. The detection of extensive formation of nitrotyrosine in the CNS parenchyma was observed in mice having shown neurological disorders, suggesting the role of peroxynitrites in the appearance of neurological troubles. In conclusion, this study confirmed the increased NO synthesis in the CNS of mice infected with T. b. brucei and suggests a deleterious role for NO, through the formation of peroxynitrites, in the pathogenesis of African CNS trypanosomiasis.

Topics: Animals; Brain; Coloring Agents; Female; Fluorescent Antibody Technique; Immunohistochemistry; Mice; NADPH Dehydrogenase; Nervous System Diseases; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Trypanosoma brucei brucei; Trypanosomiasis, African; Tyrosine

An HPLC method was used for quantification of 3-nitrotyrosine (3-NT) in human postmortem brain tissue. A peak with similar retention time to 3-NT was detected in brain tissue from patients with Parkinson's disease, Huntington's chorea, multiple system atrophy, and Alzheimer's disease but not in control tissue. The peak was lost on reduction with dithionite, a criterion often used to identify 3-NT. Tissue from the same neurodegenerative diseases was analysed by HPLC using a photodiode array detector in series with an amperometric electrochemical detector, but the peak was found not to be 3-NT. The absorbance spectrum, fragmentation pattern on mass spectroscopy, and electrochemical profile of this peak do not match authentic 3-NT. A search of the mass spectroscopy databases failed to reveal its identity. The presence of this closely eluting, dithionite-reducible peak could confound analysis of human tissues for 3-NT. In vitro experiments showed that high concentrations of peroxynitrite were needed to achieve detectable levels of 3-NT in human brain tissue.

Topics: Artifacts; Brain; Cadaver; Chromatography, High Pressure Liquid; Humans; Nerve Degeneration; Nervous System Diseases; Reference Values; Tyrosine

Peroxynitrite, generated by the reaction of nitric oxide (NO) with superoxide at sites of inflammation, is a strong oxidant capable of damaging tissues and cells. Detection of nitrotyrosine (NT) at inflammatory sites serves as a biochemical marker for peroxynitrite-mediated damage. In this study, NT was detected immunohistochemically within autopsied CNS tissues from six of nine multiple sclerosis (MS) patients, and in most of the MS sections displaying inflammation. Nitrite and nitrate, the stable oxidation products of NO and peroxynitrite, respectively, were measured in cerebrospinal fluid samples obtained from MS patients and controls. Levels of nitrate were elevated significantly during clinical relapses of MS. These data suggest that peroxynitrite formation is a major consequence of NO produced in MS-affected CNS and implicate a role for this powerful oxidant in the pathogenesis of MS.

Topics: Central Nervous System; Humans; Immunohistochemistry; Inflammation; Multiple Sclerosis; Nervous System Diseases; Nitrates; Nitrites; Recurrence; Tyrosine