3-nitrotyrosine and Pneumonia

Topics: Animals; Asthma; Bronchoalveolar Lavage Fluid; Cystic Fibrosis; Eosinophils; Humans; Nitrates; Nitric Oxide; Pneumonia; Tyrosine

Oxidative stress is a modifiable risk-factor in infection causing damage to human cells. As an adaptive response, cells catabolize Tyrosine to 3-Nitrotyrosine (Tyr-NO2) by nitrosylation. We investigated whether a more efficient reduction in oxidative stress, mirrored by a lowering of Tyrosine, and an increase in Tyr-NO2 and the Tyrosine/Tyr-NO2 ratio was associated with better clinical outcomes in patients with community-acquired pneumonia (CAP).. We measured Tyrosine and Tyr-NO2 in CAP patients from a previous randomized Swiss multicenter trial. The primary endpoint was adverse outcome defined as death or ICU admission within 30-days; the secondary endpoint was 6-year mortality.. Of 278 included CAP patients, 10.4% experienced an adverse outcome within 30 days and 45.0% died within 6 years. After adjusting for the pneumonia Severity Index [PSI], BMI and comorbidities, Tyrosine nitrosylation was associated with a lower risk for short-term adverse outcome and an adjusted OR of 0.44 (95% CI 0.20 to 0.96, p = 0.039) for Tyr-NO2 and 0.98 (95% CI 0.98 to 0.99, p = 0.043) for the Tyrosine/Tyr-NO2 ratio. There were no significant associations for long-term mortality over six-years for Tyr-NO2 levels (adjusted hazard ratio 0.81, 95% CI 0.60 to 1.11, p = 0.181) and Tyrosine/Tyr-NO2 ratio (adjusted hazard ratio 1.00, 95% CI 0.99 to 1.00, p = 0.216).. Tyrosine nitrosylation in our cohort was associated with better clinical outcomes of CAP patients at short-term, but not at long term. Whether therapeutic modulation of the Tyrosine/Tyr-NO2 pathway has beneficial effects should be evaluated in future studies.. ISRCTN95122877. Registered 31 July 2006.

Topics: Aged; Aged, 80 and over; Cohort Studies; Community-Acquired Infections; Female; Follow-Up Studies; Humans; Intensive Care Units; Male; Metabolic Networks and Pathways; Middle Aged; Mortality; Patient Admission; Pneumonia; Prognosis; Risk Factors; Switzerland; Time Factors; Tyrosine

Systemic sclerosis (SSc) is a connective-tissue disease characterized by vascular injury, immune-system disorders, and excessive fibrosis of the skin and multiple internal organs. Recent reports found that RhoA/Rho-kinase (ROCK) pathway is implicated in various fibrogenic diseases. Intradermal injection of hypochlorous acid (HOCl)-generating solution induced inflammation, autoimmune activation, and fibrosis, mimicking the cutaneous diffuse form of SSc in humans. Our study aimed firstly to describe pulmonary inflammation and fibrosis induced by HOCl in mice, and secondly to determine whether fasudil, a selective inhibitor of ROCK, could prevent lung and skin fibroses in HOCl-injected mice.. Female C57BL/6 mice received daily intradermal injection of hypochlorous acid (HOCl) for 6 weeks to induce SSc, with and without daily treatment with fasudil (30 mg·kg(-1)·day(-1)) by oral gavage.. HOCl intoxication induced significant lung inflammation (macrophages and neutrophils infiltration), and fibrosis. These modifications were prevented by fasudil treatment. Simultaneously, HOCl enhanced ROCK activity in lung and skin tissues. Inhibition of ROCK reduced skin fibrosis, expression of α-smooth-muscle actin and 3-nitrotyrosine, as well as the activity of ROCK in the fibrotic skin of HOCl-treated mice, through inhibition of phosphorylation of Smad2/3 and ERK1/2. Fasudil significantly decreased the serum levels of anti-DNA-topoisomerase-1 antibodies in mice with HOCl-induced SSc.. Our findings confirm HOCl-induced pulmonary inflammation and fibrosis in mice, and provide further evidence for a key role of RhoA/ROCK pathway in several pathological processes of experimental SSc. Fasudil could be a promising therapeutic approach for the treatment of SSc.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Actins; Animals; Disease Models, Animal; Female; Hypochlorous Acid; Lung; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Pneumonia; Pulmonary Fibrosis; rho-Associated Kinases; Scleroderma, Systemic; Skin; Smad2 Protein; Smad3 Protein; Tyrosine

Acute lung injury (ALI) and sepsis are major contributors to the morbidity and mortality of critically ill patients. The current study was designed further evaluate the mechanism of pulmonary vascular hyperpermeability in sheep with these injuries.. Sheep were randomized to a sham-injured control group (n=6) or ALI/sepsis group (n=7). The sheep in the ALI/sepsis group received inhalation injury followed by instillation of Pseudomonas aeruginosa into the lungs. These groups were monitored for 24 h. Additional sheep (n=16) received the injury and lung tissue was harvested at different time points to measure lung wet/dry weight ratio, vascular endothelial growth factor (VEGF) mRNA and protein expression as well as 3-nitrotyrosine protein expression in lung homogenates.. The injury induced severe deterioration in pulmonary gas exchange, increases in lung lymph flow and protein content, and lung water content (P<0.01 each). These alterations were associated with elevated lung and plasma nitrite/nitrate concentrations, increased tracheal blood flow, and enhanced VEGF mRNA and protein expression in lung tissue as well as enhanced 3-nitrotyrosine protein expression (P<0.05 each).. This study describes the time course of pulmonary microvascular hyperpermeability in a clinical relevant large animal model and may improve the experimental design of future studies.

Topics: Acute Lung Injury; Animals; Capillary Permeability; Disease Models, Animal; Female; Lung; Microcirculation; Nitric Oxide; Pneumonia; Pseudomonas aeruginosa; Pseudomonas Infections; Pulmonary Circulation; Pulmonary Edema; Pulmonary Gas Exchange; RNA, Messenger; Sepsis; Sheep; Smoke Inhalation Injury; Time Factors; Tyrosine; Vascular Endothelial Growth Factor A

Adrenomedullin (AM), a 52-amino acid ringed-structure peptide with C-terminal amidation, was originally isolated from human pheochromocytoma. AM are widely distributed in various tissues and acts as a local vasoactive hormone in various conditions.. In the present study, we investigated the efficacy of AM on the animal model of bleomycin (BLM)-induced lung injury. Mice were subjected to intratracheal administration of BLM and were assigned to receive AM daily by an intraperitoneal injection of 200 ngr/kg.. Myeloperoxidase activity, lung histology, immunohistochemical analyses for cytokines and adhesion molecules expression, inducible nitric oxide synthase (iNOS), nitrotyrosine, and poly (ADP-ribose) polymerase (PARP) were performed one week after fibrosis induction. Lung histology and transforming growth factor beta (TGF-β) were performed 14 and 21 days after treatments. After bleomycin administration, AM-treated mice exhibited a reduced degree of lung damage and inflammation compared with BLM-treated mice, as shown by the reduction of (1) myeloperoxidase activity (MPO), (2) cytokines and adhesion molecules expression, (3) nitric oxide synthase expression, (4) the nitration of tyrosine residues, (5) poly (ADP-ribose) (PAR) formation, a product of the nuclear enzyme poly (ADP-ribose) polymerase (PARP) (6) transforming growth factor beta (TGF-β) (7)and the degree of lung injury.. Our results indicate that AM administration is able to prevent bleomycin induced lung injury through the down regulation of proinflammatory factors.

Topics: Adrenomedullin; Animals; Anti-Inflammatory Agents; Bleomycin; Cell Adhesion Molecules; Cytokines; Disease Models, Animal; Inflammation Mediators; Injections, Intraperitoneal; Lung; Male; Mice; Mice, Inbred ICR; Nitric Oxide Synthase Type II; Peroxidase; Pneumonia; Poly(ADP-ribose) Polymerases; Pulmonary Fibrosis; Severity of Illness Index; Time Factors; Transforming Growth Factor beta; Tyrosine

Adenosine A(2A) receptor agonists may be important regulators of inflammation. The aim of this study was to investigate the effects of CGS 21680 (0.1mg/kgi.p.), an agonist of the adenosine (A(2A)) receptor, in a mouse model of carrageenan-induced pleurisy. Injection of carrageenan into the pleural cavity of mice elicited an acute inflammatory response characterised by: infiltration of neutrophils in lung tissues and subsequent lipid peroxidation, increased production of nitric oxide (NO), cytokines such as tumour necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) and increased expression of intercellular adhesion molecule (ICAM-1) and platelet-adhesion molecule (P-selectin). Furthermore, carrageenan induced the expression of nuclear factor-κB (NF-κB), inducible nitric oxide synthase (iNOS), nitrotyrosine, the activation of poly-ADP-ribosyl polymerase (PARP), as well as induced apoptosis (FAS-ligand expression, Bax and Bcl-2 expression) in the lung tissues. Administration of CGS 21680, 30 min prior to challenge with carrageenan, caused a significant reduction of all the parameters of inflammation measured. In addition, to confirm the anti-inflammatory effect of CGS 21680, we have also evaluated the effects of CGS 21680 post-treatment (30 min after the challenge with carrageenan) and we have demonstrated that also it caused a reduction of neutrophil infiltration and the degree of lung injury. Thus, based on these findings we propose that adenosine A(2A) receptor agonists such as CGS 21680 may be useful in the treatment of various inflammatory diseases.

Topics: Acute Disease; Adenosine; Adenosine A2 Receptor Agonists; Animals; bcl-2-Associated X Protein; Carrageenan; Cytokines; Enzyme Activation; Fas Ligand Protein; Gene Expression Regulation, Enzymologic; I-kappa B Proteins; Intercellular Adhesion Molecule-1; Lipid Peroxidation; Male; Mice; NF-KappaB Inhibitor alpha; Nitrates; Nitric Oxide Synthase Type II; Nitrites; P-Selectin; Peroxidase; Phenethylamines; Pleurisy; Pneumonia; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-bcl-2; Receptor, Adenosine A2A; Transcription Factor RelA; Tyrosine

Exposure to small size particulates is regarded as a risk factor for cardiovascular diseases.. We exposed young and aged apolipoprotein E knockout mice (apoE-/-) to carbon black (Printex 90, 14 nm) by intratracheal instillation, with different dosing and timing, and measured vasomotor function, progression of atherosclerotic plaques, and VCAM-1, ICAM-1, and 3-nitrotyrosine in blood vessels. The mRNA expression of VCAM-1, ICAM-1, HO-1, and MCP-1 was examined in lung tissue.. Young apoE-/- mice exposed to two consecutive 0.5 mg/kg doses of carbon black exhibited lower acetylcholine-induced vasorelaxation in aorta segments mounted in myographs, whereas single doses of 0.05-2.7 mg/kg produced no such effects. The phenylephrine-dependent vasocontraction response was shifted toward a lower responsiveness in the mice exposed once to a low dose for 24 hours. No effects were seen on the progression of atherosclerotic plaques in the aged apoE-/- mice or on the expression of VCAM-1 and ICAM-1 and the presence of 3-nitrotyrosine in the vascular tissue of either young or aged apoE-/- mice. The expression of MCP-1 mRNA was increased in the lungs of young apoE-/- mice exposed to 0.9-2.7 mg/kg carbon black for 24 hours and of aged apoE-/- mice exposed to two consecutive 0.5 mg/kg doses of carbon black seven and five weeks prior to sacrifice.. Exposure to nano-sized carbon black particles is associated with modest vasomotor impairment, which is associated neither with nitrosative stress nor with any obvious increases in the expression of cell adhesion proteins on endothelial cells or in plaque progression. Evidence of pulmonary inflammation was observed, but only in animals exposed to higher doses.

Topics: Administration, Inhalation; Aging; Animals; Apolipoproteins E; Bronchoalveolar Lavage Fluid; Chemokine CCL2; Heme Oxygenase-1; In Vitro Techniques; Intercellular Adhesion Molecule-1; Lung; Mice; Mice, Knockout; Muscle Contraction; Muscle, Smooth, Vascular; Nanoparticles; Oxidative Stress; Plaque, Atherosclerotic; Pneumonia; Soot; Time Factors; Tyrosine; Vascular Cell Adhesion Molecule-1

In this study, we evaluated the protective effect and therapeutic potential of the prostaglandin E(2) (PGE(2)) synthetic analog 16,16-dimethyl-PGE(2) (dmPGE(2)) in the animal model of pulmonary fibrosis induced by bleomycin. Mice subjected to intratracheal administration of bleomycin (1 mg/kg) received a dmPGE(2) dose of 30 microg/kg/day by continuous subcutaneous infusion. Bronchoalveolar lavage (BAL); immunohistochemical analysis for IL-1, TNF-alpha, and nitrotyrosine; measurement of fluid content in lung; myeloperoxidase activity assay; and lung histology were performed 1 week later. Lung histology and Sircol assay for collagen deposition were performed 3 weeks after treatments. Changes of body weight and survival rate were also evaluated at 1 and 3 weeks. Compared with bleomycin-treated mice, dmPGE(2) co-treated mice exhibited a reduced degree of body weight loss and mortality rate as well as of lung damage and inflammation, as shown by the significant reduction of: (1) lung infiltration by leukocytes; (2) myeloperoxidase activity; (3) IL-1, TNF-alpha, and nitrotyrosine immunostaining; (4) lung edema; and (5) histologic evidence of lung injury and collagen deposition. In a separate set of experiments, dmPGE(2) treatment was started 3 days after bleomycin administration, and the evaluation of lung damage and inflammation was assessed 4 days later. Importantly, delayed administration of dmPGE(2) also was able to protect from inflammation and lung injury induced by bleomycin. These results, indicating that dmPGE(2) is able to prevent and to reduce bleomycin-induced lung injury through its regulatory and anti-inflammatory properties, encourage further research to find new options for the treatment of pulmonary fibrosis.

Topics: 16,16-Dimethylprostaglandin E2; Animals; Bleomycin; Body Weight; Bronchoalveolar Lavage Fluid; Collagen; Disease Models, Animal; Infusions, Subcutaneous; Interleukin-1beta; Lung; Lung Injury; Male; Mice; Peroxidase; Pneumonia; Protective Agents; Pulmonary Edema; Pulmonary Fibrosis; Time Factors; Tumor Necrosis Factor-alpha; Tyrosine

Hyperoxia disrupts postnatal lung development in part through inducing inflammation. To determine the contribution of leukocyte-derived reactive oxygen species, we exposed newborn wild-type and NADPH oxidase p47(phox) subunit null (p47(phox-/-)) mice to air or acute hyperoxia (95% O(2)) for up to 11 days. Hyperoxia-induced pulmonary neutrophil influx was similar in wild-type and p47(-/-) mice at postnatal days (P) 7 and 11. Macrophages were decreased in wild-type hyperoxia-exposed mice compared with p47(phox-/-) mice at P11. Hyperoxia impaired type II alveolar epithelial cell and bronchiolar epithelial cell proliferation, but depression of type II cell proliferation was significantly less in p47(-/-) mice at P3 and P7, when inflammation was minimal. We found reciprocal results for the expression of the cell cycle inhibitor p21(cip/waf) in type II cells, which was induced in 95% O(2)-exposed wild-type mice, but significantly less in p47(phox-/-) littermates at P7. Despite partial preservation of type II cell proliferation, deletion of p47(phox) did not prevent the major adverse effects of hyperoxia on alveolar development estimated by morphometry at P11, but hyperoxia impairment of elastin deposition at alveolar septal crests was significantly worse in wild-type vs. p47(phox-/-) mice at P11. Since we found that p47(phox) is expressed in a subset of alveolar epithelial cells, its deletion may protect postnatal type II alveolar epithelial proliferation from hyperoxia through effects on epithelial as well as phagocyte-generated superoxide.

Topics: Aging; Air; Animals; Animals, Newborn; Body Weight; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p21; Epithelial Cells; Gene Deletion; Hyperoxia; Mice; NADPH Oxidases; Oxygen; Pneumonia; Protein Transport; Pulmonary Alveoli; Pulmonary Surfactant-Associated Protein C; Survival Analysis; Tyrosine

Methicillin-resistant Staphylococcus aureus (MRSA)-related pneumonia and/or sepsis are a frequent serious menace. The aim of the study was to establish a standardized and reproducible model of MRSA-induced septic pneumonia to evaluate new therapies. Sheep were operatively prepared for chronic study. After 5 days' recovery, tracheostomy was performed under anesthesia, and smoke injury was induced by inhalation of cotton smoke (48 breaths, <40 degrees C). Methicillin-resistant S. aureus (AW6) (approximately 2.5x10(11) colony-forming units) was instilled into the airway by a bronchoscope. After the injury, animals were awakened and maintained on mechanical ventilation by 100% oxygen for first 3 h, and thereafter, oxygen concentration was adjusted according to blood gases. The sheep were resuscitated by lactated Ringer solution with an initial rate of 2 mL kg(-1) h(-1) that was further adjusted according to hematocrit. Study groups include (1) sham (noninjured, nontreated; n=6), (2) S+MRSA (exposed to smoke inhalation and MRSA, nontreated; n=6), and (3) smoke (exposed to smoke inhalation alone; n=6). Injured (S+MRSA) animals showed the signs of severe sepsis-related multiple organ failure 3 h after insult. Cardiovascular morbidity was evidenced by severe hypotension, with increased heart rate, cardiac output, left atrial pressure and severely decreased systemic vascular resistance index, and left ventricle stroke work index. Pulmonary dysfunction was characterized by deteriorated gas exchange (PaO2/FIO2 and pulmonary shunt) and increased ventilatory pressures. The S+MRSA group showed significantly greater lung tissue water content, myeloperoxidase activity, and cytokine production compared with uninjured sham animals. Microvascular hyperpermeability was evidenced by marked fluid retention (fluid net balance), decreased plasma protein with decreased plasma oncotic pressure, and increased pulmonary microvascular pressure. All these changes were accompanied by 6- to 7-fold increase in plasma nitrite/nitrate and increased production of reactive nitrogen species in lung. The smoke inhalation alone had a little or no effect on these variables. This model closely mimics hyperdynamic human sepsis. The excessive production of NO may be extensively involved in the pathogenic process.

Topics: Animals; Disease Models, Animal; Female; Hematocrit; Hemodynamics; Lung; Methicillin; Methicillin Resistance; Pneumonia; Sepsis; Sheep; Smoke Inhalation Injury; Staphylococcus aureus; Time Factors; Tyrosine

In the present study, we show that an aqueous extract of Arbutus unedo L. (AuE), a Mediterranean endemic plant widely employed as an astringent, diuretic and urinary anti-septic, in vitro down-regulates the expression of some inflammatory genes, such as those encoding inducible nitric oxide synthase (iNOS) and intracellular adhesion molecule-(ICAM)-1, exerting a inhibitory action on both IFN-gamma-elicited STAT1 activation and IL-6-elicited STAT3 activation. To evaluate further the effect of AuE in animal models of acute inflammation, we examined whether AuE administration attenuates inflammatory response of murine induced by intrapleural injection of carrageenan. For this purpose we studied: (1) STAT1/3 activation, (2) TNF-alpha, IL-1beta and IL-6 production in pleural exudate, (3) lung iNOS, COX-2 and ICAM-1 expression, (4) neutrophil infiltration, (5) the nitration of cellular proteins by peroxynitrite, (6) lipid peroxidation, (7) prostaglandin E2 and nitrite/nitrate levels and (8) lung injury. We show that AuE strongly down-regulates STAT3 activation induced in the lung by carrageenan with concomitant attenuation of all parameters examined associated with inflammation, suggesting that STAT3 should be a new molecular target for anti-inflammatory treatment. This study demonstrates that acute lung inflammation is significantly attenuated by the treatment with AuE.

Topics: Animals; Anti-Inflammatory Agents; Carrageenan; Cell Line; Dinoprostone; Ericaceae; Humans; Intercellular Adhesion Molecule-1; Interferon-gamma; Interleukin-6; Lipid Peroxidation; Male; Mice; Nitric Oxide; Nitric Oxide Synthase Type II; Plant Extracts; Plant Leaves; Pleurisy; Pneumonia; STAT1 Transcription Factor; STAT3 Transcription Factor; Tumor Necrosis Factor-alpha; Tyrosine

A(2A) adenosine receptor (A(2A)AR) has potent anti-inflammatory properties, which may be important in the regulation of airway reactivity and inflammation. Inflammatory cells that possess A(2A)AR also produce nitrosative stress, which is associated with pathophysiology of asthma, so we hypothesized that A(2A)AR deficiency may lead to increased airway reactivity and inflammation through nitrosative stress. Thus the present study was carried out to investigate the role of A(2A)AR on airway reactivity, inflammation, NF-kappaB signaling, and nitrosative stress in A(2A)AR knockout (KO) and wild-type (WT) mice using our murine model of asthma. Animals were sensitized intraperitoneally on days 1 and 6 with 200 microg of ragweed, followed by aerosolized challenges with 0.5% ragweed on days 11, 12, and 13, twice a day. On day 14, airway reactivity to methacholine was assessed as enhanced pause (Penh) using whole body plethysmography followed by bronchoalveolar lavage (BAL) and lung collection for various analyses. Allergen challenge caused a significant decrease in expression of A(2A)AR in A(2A) WT sensitized mice, with A(2A)AR expression being undetected in A(2A) KO sensitized group leading to decreased lung cAMP levels in both groups. A(2A)AR deletion/downregulation led to an increase in Penh to methacholine and influx of total cells, eosinophils, lymphocytes, and neutrophils in BAL with highest values in A(2A) KO sensitized group. A(2A) KO sensitized group further had increased NF-kappaB expression and nitrosative stress compared with WT sensitized group. These data suggest that A(2A)AR deficiency leads to airway inflammation and airway hyperresponsiveness, possibly via involvement of nitrosative stress in this model of asthma.

Topics: Ambrosia; Animals; Asthma; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cyclic AMP; Disease Models, Animal; I-kappa B Proteins; Leukocytes; Lipid Peroxidation; Male; Methacholine Chloride; Mice; Mice, Inbred Strains; Mice, Knockout; NF-KappaB Inhibitor alpha; Nitric Oxide Synthase Type II; Phosphorylation; Plethysmography, Whole Body; Pneumonia; Reactive Nitrogen Species; Receptor, Adenosine A2A; Reverse Transcriptase Polymerase Chain Reaction; Transcription Factor RelA; Tyrosine

Pulmonary manifestations of oxygen toxicity were studied and quantified in rats breathing >98% O(2) at 1, 1.5, 2, 2.5, and 3 ATA to test our hypothesis that different patterns of pulmonary injury would emerge, reflecting a role for central nervous system (CNS) excitation by hyperbaric oxygen. At 1.5 atmosphere absolute (ATA) and below, the well-recognized pattern of diffuse pulmonary damage developed slowly with an extensive inflammatory response and destruction of the alveolar-capillary barrier leading to edema, impaired gas exchange, respiratory failure, and death; the severity of these effects increased with time over the 56-h period of observation. At higher inspired O(2) pressures, 2-3 ATA, pulmonary injury was greatly accelerated but less inflammatory in character, and events in the brain were a prelude to a distinct lung pathology. The CNS-mediated component of this lung injury could be attenuated by selective inhibition of neuronal nitric oxide synthase (nNOS) or by unilateral transection of the vagus nerve. We propose that extrapulmonary, neurogenic events predominate in the pathogenesis of acute pulmonary oxygen toxicity in hyperbaric oxygenation, as nNOS activity drives lung injury by modulating the output of central autonomic pathways.

Topics: Animals; Behavior, Animal; Blood Gas Analysis; Body Fluids; Bronchoalveolar Lavage Fluid; Hyperoxia; L-Lactate Dehydrogenase; Lung; Lung Diseases; Male; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitrites; Oxygen; Pneumonia; Pulmonary Edema; Rats; Rats, Sprague-Dawley; Survival Analysis; Tyrosine; Vagotomy

Surfactant protein (SP)-D, a 43-kD multifunctional collagen-like lectin, is synthesized and secreted by the airway epithelium. SP-D knockout (SP-D [-/-]) mice exhibit an increase in the number and size of airway macrophages, peribronchiolar inflammation, increases in metalloproteinase activity, and development of emphysema. Nitric oxide (NO) is involved in a variety of signaling processes, and because altered NO metabolism has been observed in inflammation, we hypothesized that alterations in its metabolism would underlie the proinflammatory state observed in SP-D deficiency. Examination of the bronchial alveolar lavage (BAL) from SP-D (-/-) mice reveals a significant increase in protein and phospholipid content and total cell count. NO production and inducible NO synthase expression were increased in the BAL; however, there was a decline in S-nitrosothiol (SNO) content in the BAL and a loss of SNO immunoreactivity within the tissue. This decline in SNO was accompanied by an increase in nitrotyrosine staining. We conclude that inflammation that occurs in SP-D deficiency results in an increase in NO production and a shift in the chemistry and targets of NO. We speculate that the proinflammatory response due to SP-D deficiency results, in part, from a disruption of NO-mediated signaling within the innate immune system.

Topics: Animals; Bronchoalveolar Lavage Fluid; Cell Count; Female; Homozygote; Immunity, Innate; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Phospholipids; Pneumonia; Pulmonary Surfactant-Associated Protein D; Pulmonary Surfactants; S-Nitrosothiols; Tyrosine

Acute lung injury (ALI) is characterized by increased alveolar cytokines, inflammatory cell infiltration, oxidative stress, and alveolar cell apoptosis. Previous work suggested that xanthine oxidoreductase (XOR) may contribute to oxidative stress in ALI as a product of the vascular endothelial cell. We present evidence that cytokine induced lung inflammation and injury involves activation of XOR in the newly recruited mononuclear phagocytes (MNP). We found that XOR was increased predominantly in the MNP that increase rapidly in the lungs of rats that develop ALI following intratracheal cytokine insufflation. XOR was recovered from the MNP largely converted to its oxygen radical generating, reversible O-form, and alveolar MNP exhibited increased oxidative stress as evidenced by increased nitrotyrosine staining. Cytokine insufflation also increased alveolar cell apoptosis. A functional role for XOR in cytokine-induced inflammation was demonstrated when feeding rats two different XOR inhibitors, tungsten and allopurinol, decreased MNP XOR induction, nitrotyrosine staining, inflammatory cell infiltration, and alveolar cell apoptosis. Transfer of control or allopurinol treated MNP into rat lungs confirmed a specific role for MNP XOR in promoting lung inflammation. These data indicate that XOR can contribute to lung inflammation by its expression and conversion in a highly mobile inflammatory cell population.

Topics: Allopurinol; Animals; Apoptosis; Cell Differentiation; Cytokines; Enzyme Induction; Enzyme Inhibitors; Interferon-gamma; Interleukin-1; Lung; Male; Phagocytes; Pneumonia; Pulmonary Alveoli; Rats; Rats, Sprague-Dawley; Respiratory Distress Syndrome; Tungsten; Tyrosine; Xanthine Oxidase

1. The aim of this study was to investigate the effect of GW274150, a novel, potent and selective inhibitor of inducible nitric oxide synthase (iNOS) activity in a model of lung injury induced by carrageenan administration in the rats. 2. Injection of carrageenan into the pleural cavity of rats elicited an acute inflammatory response characterized by: fluid accumulation in the pleural cavity which contained a large number of polymorphonuclear cells (PMNs) as well as an infiltration of PMNs in lung tissues and subsequent lipid peroxidation, and increased production of nitrite/nitrate (NO(x)), tumour necrosis factor alpha (TNF-alpha) and interleukin-1beta (IL-1beta). 3. All parameters of inflammation were attenuated in a dose-dependent manner by GW274150 (2.5, 5 and 10 mg x kg(-1) injected i.p. 5 min before carrageenan). 4. Carrageenan induced an upregulation of the intracellular adhesion molecules-1 (ICAM-1), as well as nitrotyrosine and poly (ADP-ribose) (PAR) as determined by immunohistochemical analysis of lung tissues. 5. The degree of staining for the ICAM-1, nitrotyrosine and PAR was reduced by GW274150. These results clearly confirm that NO from iNOS plays a role in the development of the inflammatory response by altering key components of the inflammatory cascade. 6. GW274150 may offer a novel therapeutic approach for the management of various inflammatory diseases where NO and related radicals have been postulated to play a role.

Topics: Animals; Carrageenan; Disease Models, Animal; Enzyme Inhibitors; Intercellular Adhesion Molecule-1; Interleukin-1; Interleukin-1beta; Lung; Male; Malondialdehyde; Nitrates; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Peptide Fragments; Peroxidase; Pleuropneumonia; Pneumonia; Poly(ADP-ribose) Polymerases; Rats; Rats, Sprague-Dawley; Sulfides; Tumor Necrosis Factor-alpha; Tyrosine

Although nitrated proteins have been repeatedly used as markers of lung injury, little is known about their formation and metabolism under hyperoxia. We therefore measured 3-nitrotyrosine (3NTYR) concentrations in lung tissue and serum of rats with carrageenan-induced pneumonia exposed to hyperoxia. Twenty-nine Wistar male rats were assigned to one of 4 groups. Two experimental groups were treated by intratracheal application of carrageenan (0.5 ml of 0.7 % solution) and then one was exposed to hyperoxia for 7 days (FIO2 0.8), the other to air. Rats of two control groups breathed either hyperoxic gas mixture or air for 7 days. At the end of exposure the ventilation was determined in anesthetized, intubated animals in which 3NTYR concentrations were measured in the lung tissue and nitrites and nitrates (NOx) were estimated in the serum. Carrageenan instillation increased 3NTYR concentrations in lung tissue (carrageenan-normoxic group 147+/-7 pmol/g protein, control 90+/-10 pmol/g protein) and NOx concentration in the serum (carrageenan-normoxic group 126+/-13 ppb, control 78+/-9 ppb). Hyperoxia had no effect on lung tissue 3NTYR concentration in controls (control-hyperoxic 100+/-14 pmol/g protein) but blocked the increase of lung tissue 3NTYR in carrageenan-treated rats (carrageenan-hyperoxic 82+/-13 pmol/g protein), increased NOx in serum (control-hyperoxic 127+/-19 ppb) and decreased serum concentration of 3NTYR in both hyperoxic groups (carrageenan-hyperoxic 51+/-5 pmol/g protein, control-hyperoxic 67+/-7 pmol/g protein, carrageenan-normoxic 82+/-9 pmol/g protein, control 91+/-7 pmol/g protein). The results suggest that hyperoxia affects nitration of tyrosine residues, probably by increasing 3NTYR degradation.

Topics: Animals; Carrageenan; Hyperoxia; Lung; Male; Pneumonia; Rats; Rats, Wistar; Tyrosine

Mice deficient in inducible nitric oxide synthase (iNOS; C57Bl/6Ai-[KO]NOS2 N5) or wild-type C57Bl/6 mice were exposed to 1 part/million of ozone 8 h/night or to filtered air for three consecutive nights. Endpoints measured included lavagable total protein, macrophage inflammatory protein (MIP)-2, matrix metalloproteinase (MMP)-9, cell content, and tyrosine nitration of whole lung proteins. Ozone exposure caused acute edema and an inflammatory response in the lungs of wild-type mice, as indicated by significant increases in lavage protein content, MIP-2 and MMP-9 content, and polymorphonuclear leukocytes. The iNOS knockout mice showed significantly greater levels of lung injury by all of these criteria than did the wild-type mice. We conclude that iNOS knockout mice are more susceptible to acute lung damage induced by exposure to ozone than are wild-type C57Bl/6 mice and that protein nitration is associated with the degree of inflammation and not dependent on iNOS-derived nitric oxide.

Topics: Acute Disease; Animals; Bronchoalveolar Lavage Fluid; Chemokine CXCL2; Chemokines; Disease Susceptibility; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ozone; Pneumonia; Proteins; Pulmonary Edema; Tyrosine

Inhalation of ozone causes Type I epithelial cell necrosis and Type II cell hyperplasia and proliferation. This is associated with an accumulation of activated macrophages in the lower lung, which we have demonstrated contribute to tissue injury. Nitric oxide (NO) is a highly reactive cytotoxic macrophage-derived mediator that has been implicated in lung damage. In the present studies we used knockout mice with a targeted disruption of the gene for inducible nitric oxide synthase (NOSII) to analyze the role of NO in ozone-induced lung inflammation and tissue injury. Treatment of wild-type control mice with ozone (0.8 ppm) for 3 h resulted in a time-dependent increase in protein and cells in bronchoalveolar lavage fluid, which reached a maximum 24-48 h after exposure. Alveolar macrophages isolated from animals treated with ozone were found to produce increased amounts of NO, as well as peroxynitrite. This was correlated with induction of NOSII protein and nitrotyrosine staining of lung macrophages in tissue sections and in culture. Production of superoxide anion and prostaglandin (PG)E2 by alveolar macrophages was also increased after ozone inhalation. In contrast, alveolar macrophages from NOSII knockout mice did not produce reactive nitrogen intermediates even after ozone inhalation. Moreover, production of PGE2 was at control levels. NOSII knockout mice were also protected from ozone-induced inflammation and tissue injury, as measured by bronchoalveolar lavage protein and cell number. There was also no evidence of peroxynitrite-mediated lung damage in these animals. Taken together, these data demonstrate that NO, produced via NOSII, and potentially, its reactive oxidative product peroxynitrite, play a critical role in ozone-induced release of inflammatory mediators and in tissue injury.

Topics: Administration, Inhalation; Animals; Bronchoalveolar Lavage Fluid; Cells, Cultured; Dinoprostone; Female; Interferon-gamma; Lipopolysaccharides; Lung; Macrophages, Alveolar; Mice; Mice, Inbred Strains; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Oxidants, Photochemical; Ozone; Peroxynitrous Acid; Pneumonia; Superoxides; Tyrosine

Topics: Administration, Inhalation; Animals; Endothelium, Vascular; Epoprostenol; Hemoglobins; Humans; Hydroxyl Radical; Hypertension, Pulmonary; Nitrates; Nitric Oxide; Nitrogen Dioxide; Oxidation-Reduction; Pneumonia; Prostaglandin H2; Prostaglandins H; Signal Transduction; Superoxides; Tyrosine

In a model of idiopathic pneumonia syndrome after bone marrow transplantation (BMT), injection of allogeneic T cells induces nitric oxide (.NO), and the addition of cyclophosphamide (Cy) generates superoxide (O.) and a tissue-damaging nitrating oxidant. We hypothesized that.NO and O. balance are major determinants of post-BMT survival and inflammation. Inducible nitric oxide synthase (iNOS) deletional mutant mice (-/-) given donor bone marrow and spleen T cells (BMS) exhibited improved survival compared with matched BMS controls. Bronchoalveolar lavage fluids obtained on day 7 post-BMT from iNOS(-/-) BMS mice contained less tumor necrosis factor-alpha and interferon-gamma, indicating that.NO stimulated the production of proinflammatory cytokines. However, despite suppressed inflammation and decreased nitrotyrosine staining, iNOS(-/-) mice given both donor T cells and Cy (BMS + Cy) died earlier than iNOS-sufficient BMS + Cy mice. Alveolar macrophages from iNOS(-/-) BMS + Cy mice did not produce.NO but persisted to generate strong oxidants as assessed by the oxidation of the intracellular fluorescent probe 2',7'-dichlorofluorescin. We concluded that.NO amplifies T cell-dependent inflammation and addition of Cy exacerbates.NO-dependent mortality. However, the lack of.NO during Cy-induced oxidant stress decreases survival of T cell-recipient mice, most likely by generation of.NO-independent toxic oxidants.

Topics: Animals; Bone Marrow Transplantation; Bronchoalveolar Lavage Fluid; Cyclophosphamide; Female; Immunosuppressive Agents; Interferon-gamma; Macrophages; Mice; Mice, Congenic; Mice, Inbred C57BL; Mice, Knockout; Monocytes; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrites; Oxidative Stress; Pneumonia; Survival Rate; T-Lymphocytes; Tumor Necrosis Factor-alpha; Tyrosine