3-nitrotyrosine and Body-Weight

Obstructive sleep apnea (OSA) in children is associated with cardiovascular morbidity such as systemic and pulmonary hypertension. However, it remains unclear whether endothelial dysfunction occurs in pediatric OSA and whether it is reversible on effective treatment of OSA.. Consecutive nonobese children (aged 6 to 11 years) who were diagnosed with OSA after overnight polysomnography and control children matched on the basis of age, gender, ethnicity, and body mass index underwent blood draw the next morning for soluble CD40 ligand, asymmetric dimethylarginine (ADMA), and nitrotyrosine levels, as well as 2 iterations of 60-second cuff-occlusion tests for assessment of endothelial function. These tests were repeated 4 to 6 months after adenotonsillectomy. OSA children showed blunted reperfusion kinetics after release of occlusion, which completely normalized in 20 of 26 patients after adenotonsillectomy. All 6 children in whom no improvements occurred had a strong family history of cardiovascular disease (versus 2 of the remaining 20 patients; P<0.04). Plasma nitrotyrosine and ADMA levels were similar in OSA and control children; however, soluble CD40 ligand levels were higher in OSA children and were reduced after treatment, particularly in those with normalized hyperemic responses.. Postocclusive hyperemia is consistently blunted in children with OSA, and such altered endothelial function is reversible 4 to 6 months after treatment, particularly if a family history of cardiovascular disease is not present. Although no evidence for either nitric oxide-dependent oxidative/nitrosative stress or for the increased presence of the circulating nitric oxide synthase inhibitor ADMA was found in children with OSA, soluble CD40 ligand levels were increased in OSA and reflected the changes in endothelial function after treatment.

Topics: Adenoidectomy; Arginine; Body Weight; CD40 Ligand; Child; Endothelium, Vascular; Female; Humans; Hyperemia; Hypoxia; Male; Nitric Oxide; Polysomnography; Sleep Apnea, Obstructive; Tonsillectomy; Tyrosine; Vasculitis

Bladder dysfunction in diabetes progresses gradually over time. However, the mechanisms of the development are not clear. We tested the hypothesis that oxidative stress plays a key role in the development of diabetic bladder dysfunction using an inducible smooth muscle (SM)-specific superoxide dismutase 2 (

Topics: Animals; bcl-2-Associated X Protein; Body Weight; Caspase 3; Diabetes Mellitus, Experimental; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth; Organ Size; Oxidative Stress; Proto-Oncogene Proteins c-bcl-2; Superoxide Dismutase; Tyrosine; Urinary Bladder Diseases

The present study assessed the beneficial skeletal muscle‑preserving effects of extracellular polysaccharides from Aureobasidium pullulans SM‑2001 (Polycan) (EAP) on dexamethasone (DEXA)‑induced catabolic muscle atrophy in mice. To investigate whether EAP prevented catabolic DEXA‑induced muscle atrophy, and to examine its mechanisms of action, EAP (100, 200 and 400 mg/kg) was administered orally, once a day for 24 days. EAP treatment was initiated 2 weeks prior to DEXA treatment (1 mg/kg, once a day for 10 days) in mice. Body weight alterations, serum biochemistry, calf thickness, calf muscle strength, gastrocnemius muscle thickness and weight, gastrocnemius muscle antioxidant defense parameters, gastrocnemius muscle mRNA expression, histology and histomorphometry were subsequently assessed. After 24 days, DEXA control mice exhibited muscle atrophy according to all criteria indices. However, these muscle atrophy symptoms were significantly inhibited by oral treatment with all three doses of EAP. Regarding possible mechanisms of action, EAP exhibited favorable ameliorating effects on DEXA‑induced catabolic muscle atrophy via antioxidant and anti‑inflammatory effects; these effects were mediated by modulation of the expression of genes involved in muscle protein synthesis (AKT serine/threonine kinase 1, phosphatidylinositol 3‑kinase, adenosine A1 receptor and transient receptor potential cation channel subfamily V member 4) and degradation (atrogin‑1, muscle RING‑finger protein‑1, myostatin and sirtuin 1). Therefore, these results indicated that EAP may be helpful in improving muscle atrophies of various etiologies. EAP at 400 mg/kg exhibited favorable muscle protective effects against DEXA‑induced catabolic muscle atrophy, comparable with the effects of oxymetholone (50 mg/kg), which has been used to treat various muscle disorders.

Topics: Aldehydes; Animals; Antioxidants; Ascomycota; Body Weight; Catalase; Dexamethasone; Extracellular Space; Glutathione; Male; Malondialdehyde; Mice, Inbred ICR; Muscle Fibers, Skeletal; Muscle Strength; Muscle, Skeletal; Muscular Atrophy; Nitric Oxide Synthase Type II; Organ Size; Poly(ADP-ribose) Polymerases; Polysaccharides; Reactive Oxygen Species; RNA, Messenger; Superoxide Dismutase; Tyrosine

The purpose of this study was to use histological and biochemical methods in order to evaluate changes taking place in the ovarian of rats exposed to the effect of a 900-megahertz (MHz) electromagnetic field (EMF) in middle and late adolescence.. Twenty-four 34-d-old female Sprague-Dawley rats were assigned equally to control, sham and EMF groups. EMF group rats were exposed to the effect of a 900-MHz EMF for 1 h a day, at the same time every day between postnatal days 35 and 59, while inside an EMF cage. Sham group rats were kept inside the EMF cage for the same time between postnatal days 35 and 59 without being exposed to any EMF effect. At the end of the study, rats' ovarian were removed and blood specimens were taken. Right ovarium tissues were subjected to routine histological procedures and stained with hematoxylin and eosin, periodic acid shift and Masson's trichrome. Follicles were counted in ovarian sections stained with hematoxylin and eosin. The TUNEL method was used to evaluate apoptosis. Left ovarian tissue and blood specimens were investigated biochemically.. Histopathological examination of EMF group ovarian tissue revealed thinning in the zona granulosa and theca layers, shrinking in granulosa cells, reduced mitotic activity and leukocyte infiltration in the follicles and stroma. Secondary follicle numbers in the EMF group were significantly lower than in the other groups. In terms of biochemistry, EMF and sham group superoxide dismutase, catalase and anti-Mullerian hormone levels and EMF group 3-nitrotyrosine values increased significantly compared to the control group. EMF and sham group serum catalase and 8-hydroxy-deoxiguanosine values increased significantly compared to the control group, and EMF group total oxidant status and oxidative stress index values were significantly higher compared to the sham and control groups.. A total of 900-MHz EMF applied in middle and late adolescence may cause changes in the morphology and biochemistry of the rat ovarium.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Body Weight; Catalase; Electromagnetic Fields; Female; Granulosa Cells; Guanine; Humidity; Leukocytes; Mitosis; Organ Size; Ovary; Oxidative Stress; Rats; Rats, Sprague-Dawley; Temperature; Theca Cells; Time Factors; Tyrosine

Lipid accumulation in the liver and pancreas is primarily caused by combined hyperlipidemia. However, the effect of isolated hypercholesterolemia without hypertriglyceridemia is not fully described. Therefore, our aim was to investigate whether hypercholesterolemia alone leads to alterations both in hepatic and pancreatic lipid panel and histology in rats.. Male Wistar rats were fed with 2% cholesterol +0.25% cholate-supplemented diet or standard chow for 12 weeks. Blood was collected at weeks 0, 4, 8 and 12 to measure serum cholesterol and triglyceride levels. At week 12, both the pancreas and the liver were isolated for further histological and biochemical analysis. Hepatic and plasma fatty acid composition was assessed by gas chromatography. Expression of mRNA of major enzymes involved in saturated/unsaturated fatty acid synthesis was analyzed by qPCR. In separate experiments serum enzyme activities and insulin levels were measured at week 9.. At week 12, rats fed with 2% cholesterol +0.25% cholate-supplemented diet were characterized by elevated serum cholesterol (4.09 ± 0.20 vs. 2.89 ± 0.22 mmol/L, *p < 0.05) while triglyceride (2.27 ± 0.05 vs. 2.03 ± 0.03 mmol/L) and glucose levels (5.32 ± 0.14 vs. 5.23 ± 0.10 mmol/L) remained unchanged. Isolated hypercholesterolemia increased hepatic lipid accumulation, hepatic cholesterol (5.86 ± 0.22 vs. 1.60 ± 0.15 ng/g tissue, *p < 0.05) and triglyceride contents (19.28 ± 1.42 vs. 6.78 ± 0.71 ng/g tissue, *p < 0.05), and hepatic nitrotyrosine level (4.07 ± 0.52 vs. 2.59 ± 0.31 ng/mg protein, *p < 0.05). The histology and tissue lipid content of the pancreas was not affected. Serum total protein level, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities remained unchanged in response to isolated hypercholesterolemia while serum alkaline phosphatase activity (ALP) significantly increased. Plasma insulin levels did not change in response to isolated hypercholesterolemia suggesting an intact endocrine function of the pancreas. Isolated hypercholesterolemia caused a significantly increased hepatic and serum fatty acid level associated with a marked alteration of fatty acid composition. Hepatic expression of Δ9-desaturase (SCD1) was increased 4.92×, while expression of Δ5-desaturase and Δ6-desaturase were decreased (0.447× and 0.577×, respectively) due to isolated hypercholesterolemia.. Isolated hypercholesterolemia leads to hepatic steatosis and marked alterations in the hepatic lipid profile without affecting the pancreas. Altered fatty acid profile might mediate harmful effects of cholesterol in the liver.

Topics: Animals; Blood Glucose; Body Weight; Cholesterol; Enzymes; Fatty Acids; Fatty Liver; Hypercholesterolemia; Insulin; Liver; Male; Nitrosative Stress; Organ Size; Oxidative Stress; Pancreas; Rats, Wistar; RNA, Messenger; Triglycerides; Tyrosine

Antidiabetic agents per se, apart from their glucose-lowering effect, can have an important impact on modifying the cardiovascular risk. The present study was undertaken to determine whether the known cardio-protective effects of metformin are linked to its potential ability to affect activities of HDL's paraoxonase (PON1) and platelet activating factor acetylohydrolase (PAF-AH) or via its interaction with the asymmetric dimethylarginine (ADMA)- dimethylarginine dimethylaminohydrolase (DDAH) axis. Normal and streptozotocin (STZ)-induced diabetic rats were treated with metformin (300mg/kg; 4 weeks). The activity of PON1, PAF-AH and DDAH were measured spectrophotometrically. The plasma ADMA level was determined by ELISA method. In STZ-induced diabetic rats the long-term administration of metformin normalized reduced PON1 activity assayed toward paraoxon (+42.5%, P<0.05), phenyl acetate (+22.35%, P<0.05) and γ-decanolactone (+108.0%, P<0.01), without affecting elevated PAF-AH activity in the plasma. Moreover, metformin increased DDAH activity in the renal cortex (+38.24%, P<0.01). Additionally metformin administration caused the increase in PON1 activity in the liver (+29.2%, P<0.01) accompanied by the reduction in the lipid peroxidation (-59.8%, P<0.001). Similarly, in non-diabetic treated rats the increase in liver PON1 activity was observed toward both paraoxon (+80.19%, P<0.001) and phenyl acetate (+29.3%, P<0.05), respectively. The present study has demonstrated that insulin-sensitizer metformin is important for preserving antioxidant HDL function in diabetes. Metformin might also exert its effect against diabetic complications by improving DDAH activity in the kidney and increasing PON1 activity in the liver.

Topics: 1-Alkyl-2-acetylglycerophosphocholine Esterase; Amidohydrolases; Animals; Aryldialkylphosphatase; Body Weight; Diabetes Mellitus, Experimental; Hypoglycemic Agents; Lipid Peroxidation; Male; Metformin; Organ Size; Oxidative Stress; Rats; Rats, Wistar; Tyrosine

Chronic exposure to intermittent hypoxia (CIH) elicits plasticity of the carotid sinus and phrenic nerves via reactive oxygen species (ROS). To determine whether CIH-induced alterations in ventilation, metabolism, and heart rate are also dependent on ROS, we measured responses to acute hypoxia in conscious rats after 14 and 21 d of either CIH or normoxia (NORM), with or without concomitant administration of allopurinol (xanthine oxidase inhibitor), combined allopurinol plus losartan (angiotensin II type 1 receptor antagonist), or apocynin (NADPH oxidase inhibitor). Carotid body nitrotyrosine production was measured by immunohistochemistry. CIH produced an increase in the ventilatory response to acute hypoxia that was virtually eliminated by all three pharmacologic interventions. CIH caused a robust increase in carotid body nitrotyrosine production that was greatly attenuated by allopurinol plus losartan and by apocynin but unaffected by allopurinol. CIH caused a decrease in metabolic rate and a reduction in hypoxic bradycardia. Both of these effects were prevented by allopurinol, allopurinol plus losartan, and apocynin.

Topics: Acetophenones; Allopurinol; Analysis of Variance; Animals; Anti-Arrhythmia Agents; Antioxidants; Body Weight; Carbon Dioxide; Carotid Sinus; Catecholamines; Chemoreceptor Cells; Free Radical Scavengers; Heart Rate; Hypoxia; Losartan; Male; Oxidative Stress; Oxygen Consumption; Plethysmography; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Regression Analysis; Respiration; Tidal Volume; Time Factors; Tyrosine

Leukocyte infiltration, improved levels of intercellular adhesion molecule 1 (ICAM-1), and oxidative stress in the colon are the principal factors in inflammatory bowel disease. The goal of the current study was to explore the effects of adelmidrol, an analog of the anti-inflammatory fatty acid amide signaling molecule palmitoylethanolamide, in mice subjected to experimental colitis. Additionally, to clarify whether the protective action of adelmidrol is dependent on the activation of peroxisome proliferator-activated receptors (PPARs), we investigated the effects of a PPARγ antagonist, GW9662, on adelmidrol action. Adelmidrol (10 mg/kg daily, o.s.) was tested in a murine experimental model of colitis induced by intracolonic administration of dinitrobenzene sulfonic acid. Nuclear factor-κB translocation, cyclooxygenase-2, and phosphoextracellular signal-regulated kinase, as well as tumor necrosis factor-α and interleukin-1β, were significantly increased in colon tissues after dinitrobenzene sulfonic acid administration. Immunohistochemical staining for ICAM-1, P-selectin, nitrotyrosine, and poly(ADP)ribose showed a positive staining in the inflamed colon. Treatment with adelmidrol decreased diarrhea, body weight loss, and myeloperoxidase activity. Adelmidrol treatment, moreover, reduced nuclear factor-κB translocation, cyclooxygenase-2, and phosphoextracellular signal-regulated kinase expression; proinflammatory cytokine release; and the incidence of nitrotyrosine and poly(ADP)ribose in the colon. It also decreased the upregulation of ICAM-1 and P-selectin. Adelmidrol treatment produced a reduction of Bax and an intensification of Bcl-2 expression. This study clearly demonstrates that adelmidrol exerts important anti-inflammatory effects that are partly dependent on PPARγ, suggesting that this molecule may represent a new pharmacologic approach for inflammatory bowel disease treatment.

Topics: Amides; Animals; Anti-Inflammatory Agents; Apoptosis; Body Weight; Colitis; Cyclooxygenase 2; Cytokines; Dicarboxylic Acids; Dinitrofluorobenzene; Ethanolamines; Extracellular Signal-Regulated MAP Kinases; Inflammatory Bowel Diseases; Intercellular Adhesion Molecule-1; Lipid Peroxidation; Male; Mice; NF-kappa B; P-Selectin; Palmitic Acids; Peroxidase; Phosphorylation; PPAR alpha; PPAR gamma; Receptor, Cannabinoid, CB2; Signal Transduction; Tyrosine

Objective To study the effects of Chinese medicinal compound Jinmaitong(JMT) on the expressions of nitrotyrosine (NT) and nerve growth factor (NGF) in dorsal root ganglia of diabetic rats. Methods Experimental rat diabetic models were established by the intraperitoneal injection of streptozotocin. Rat models were then randomly divided into four groups including normal control group (Con group),diabetes mellitus group (DM group),Jinmaitong group(JMT group)(treated with JMT similar to the fifteen-fold dose of adult recommended dosage),and taurine group(Tau group)(treated with Taurine similar to the fifteen-fold dose of adult recommended dosage),with 10 rats in each group. The Con and DM groups were treated with distilled water at a daily dose of 1 ml/100 g. All rats were given intragastric administration for 16 weeks and then killed. Body weight and blood glucose were detected before and at the 4th,8th,12th,and 16th week after treatment. The pain threshold to mechanical stimulation with von Frey filament were carried out before death. The expressions of NT and NGF in dorsal root ganglion were detected by immunohistochemistry and Western blot analysis,respectively. Results Immunohistochemistry showed that the average optical density (AOD) of NT expression in DM group were significantly higher than those in control group (P=0.000),and the AOD of NGF was significantly lower than the control group (P=0.006).The AOD of NT(P=0.000,P=0.000) in both treatment groups decreased significantly and the AOD of NGF(P=0.000, P=0.004)significantly increased compared with DM group. The AOD of NT in JMT group was significantly lower than Tau group (P=0.004). Western blot analysis showed that the protein level of NT in DM group was significantly higher than that in control group (P=0.000),and the protein level of NGF was significantly lower than that in control group (P=0.000). Compared with the DM group,the protein level of NT in both treatment groups significantly decreased (P=0.001,P=0.000),and the protein level of NGF increased significantly (P=0.000,P=0.001). Conclusion Traditional Chinese medicine JMT can obviously up-regulate the expressions of NGF and reduce the NT levels in dorsal root ganglia of diabetic rats.

Topics: Animals; Blood Glucose; Body Weight; Diabetes Mellitus, Experimental; Drugs, Chinese Herbal; Ganglia, Spinal; Immunohistochemistry; Nerve Growth Factor; Pain Threshold; Random Allocation; Rats; Tyrosine

Liver fibrosis and cirrhosis are leading causes of morbidity and mortality, with majority of preventable cases attributed to excessive alcohol consumption, viral hepatitis, or non-alcoholic fatty liver disease. We previously reported the hepatoprotective effect of Glycine propionyl-l-carnitine (GPLC) against the fulminant hepatic failure (FHF) induced by d-Galactosamine (D-GalN). In this study we evaluated the protective effect of GPLC against D-GalN induced chronic liver damage.. Animals received D-GalN twice a week for 12 weeks at a dose of 250 mg/kg body weight (BW). GPLC was given daily for 12 weeks as co-treatment along with D-GalN at a dose of 35 mg/kg BW.. D-GalN injection resulted in a considerable decrease in body weight, hepatocellular disintegration, necrosis and lipid peroxidation as evident from altered levels of SOD, CAT and MDA while GPLC significantly restored the reduced body weight and ameliorated hepatocellular damage and lipid peroxidation. D-GalN administration resulted in DNA damage as evident from TUNEL positive cells in disease control rats while; GPLC significantly alleviated the genotoxic effects of D-GalN. Further histopathological analysis revealed significant tissue and cellular damage, and increased collagen content in D-GalN challenged rats. GPLC however ameliorated the damage as evident from normal cellular and morphological architecture in GPLC co-treated rats. Hydroxyproline and nitrotyrosine (NTY) levels marked a significant decrease in GPLC co-treated rats relative to disease control. GPLC significantly blocked D-GalN induced pro-inflammatory cytokine (TNF-α, IL-6) production and at the same time inhibited the expression of α-smooth muscle actin (α-SMA), collagen-I (COL-I) and transforming growth factor-β (TGF-β) significantly.. Our results demonstrate significant protective activity of GPLC in chronic liver damage and other complications related to it. This study is a novel study to demonstrate the hepatoprotective effect of GPLC in chronic liver damage.

Topics: Actins; Animals; Body Weight; Carnitine; Chronic Disease; Collagen Type I; Galactosamine; Gene Expression Regulation; Glycine; Hydroxyproline; In Situ Nick-End Labeling; Interleukin-6; Liver; Liver Cirrhosis; Male; Oxidative Stress; Rats, Wistar; RNA, Messenger; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Tyrosine

Chronic renal fibrosis is the final common pathway of end stage renal disease caused by glomerular or tubular pathologies. Genetic background has a strong influence on the progression of chronic renal fibrosis. We recently found that Rowett black hooded rats were resistant to renal fibrosis. We aimed to investigate the role of sustained inflammation and oxidative/nitrative stress in renal fibrosis progression using this new model. Our previous data suggested the involvement of podocytes, thus we investigated renal fibrosis initiated by doxorubicin-induced (5 mg/kg) podocyte damage. Doxorubicin induced progressive glomerular sclerosis followed by increasing proteinuria and reduced bodyweight gain in fibrosis-sensitive, Charles Dawley rats during an 8-week long observation period. In comparison, the fibrosis-resistant, Rowett black hooded rats had longer survival, milder proteinuria and reduced tubular damage as assessed by neutrophil gelatinase-associated lipocalin (NGAL) excretion, reduced loss of the slit diaphragm protein, nephrin, less glomerulosclerosis, tubulointerstitial fibrosis and matrix deposition assessed by periodic acid-Schiff, Picro-Sirius-red staining and fibronectin immunostaining. Less fibrosis was associated with reduced profibrotic transforming growth factor-beta, (TGF-β1) connective tissue growth factor (CTGF), and collagen type I alpha 1 (COL-1a1) mRNA levels. Milder inflammation demonstrated by histology was confirmed by less monocyte chemotactic protein 1 (MCP-1) mRNA. As a consequence of less inflammation, less oxidative and nitrative stress was obvious by less neutrophil cytosolic factor 1 (p47phox) and NADPH oxidase-2 (p91phox) mRNA. Reduced oxidative enzyme expression was accompanied by less lipid peroxidation as demonstrated by 4-hydroxynonenal (HNE) and less protein nitrosylation demonstrated by nitrotyrosine (NT) immunohistochemistry and quantified by Western blot. Our results demonstrate that mediators of fibrosis, inflammation and oxidative/nitrative stress were suppressed in doxorubicin nephropathy in fibrosis-resistant Rowett black hooded rats underlying the importance of these pathomechanisms in the progression of renal fibrosis initiated by glomerular podocyte damage.

Topics: Aldehydes; Animals; Body Weight; Chemokine CCL2; Connective Tissue Growth Factor; Disease Progression; Disease Resistance; Dose-Response Relationship, Drug; Doxorubicin; Fibrosis; Kidney; Male; Membrane Proteins; Oxidative Stress; Proteinuria; Rats; Reactive Nitrogen Species; Species Specificity; Transforming Growth Factor beta1; Tyrosine

Chronic kidney disease (CKD) patients have a poor prognosis due to cardiovascular disease. Anemia and endothelial dysfunction are important risk factors for cardiovascular events in CKD patients, and treatment with erythropoiesis-stimulating agent (ESA) has been reported to improve the quality of life in CKD patients. In this study, we evaluated the effect of anemia correcting dose of epoetin beta pegol (continuous erythropoietin receptor activator; C.E.R.A.) on endothelial function in 5/6 nephrectomized rats (Nx rats). C.E.R.A. was subcutaneously administered once a fortnight, 5 times in total, from 1 week after nephrectomy. Twenty-four hours after last administration, endothelial function was evaluated by measuring flow-mediated dilation (FMD) in the femoral arteries of anesthetized Nx rats by ultrasound system. Femoral arteries were harvested for western blot analysis. C.E.R.A. significantly increased FMD of Nx rats. Endothelium-independent vasodilation induced by nitroglycerin injection was not influenced by C.E.R.A treatment. Nox4 expression and nitrotyrosine accumulation were significantly decreased, and phosphorylation of eNOS was significantly enhanced in the femoral arteries of C.E.R.A.-treated rats. C.E.R.A. normalized hemoglobin levels but did not affect body weight, systolic blood pressure, heart rate, urinary protein excretion and plasma creatinine. These results indicate that C.E.R.A. prevented endothelial dysfunction in Nx rats, possibly through reduction of local oxidative stress and enhancement of eNOS phosphorylation in the arteries. This study provides the first evidence that C.E.R.A. prevented endothelial dysfunction in CKD model rats under conditions of amelioration of anemia.

Topics: Animals; Blood Pressure; Body Weight; Creatinine; Disease Models, Animal; Endothelium, Vascular; Erythropoietin; Femoral Artery; Heart Rate; Hemoglobins; Kidney Function Tests; Male; NADPH Oxidase 4; NADPH Oxidases; Nephrectomy; Nitric Oxide Synthase Type III; Nitroglycerin; Polyethylene Glycols; Proteinuria; Rats; Renal Insufficiency, Chronic; Tyrosine; Ultrasonography; Vasodilation

The critical influence of the white adipose tissue (WAT) on metabolism is well-appreciated in obesity, but adipose tissue dysfunction as a mechanism underlying age-associated metabolic dysfunction requires elucidation. To explore this possibility, we assessed metabolism and measures of epididymal (e)WAT mitochondria and artery function in young (6.1 ± 0.4 months) and old (29.6 ± 0.2 months) B6D2F1 mice. There were no group differences in average daily oxygen consumption, fasted blood glucose or plasma free fatty acids, but fasted plasma insulin and the homeostatic model assessment of insulin resistance (HOMA-IR%) were higher in the old (∼50-85%, P < 0.05). Tissue mass (P < 0.05) and adipocyte area were lower (∼60%) (P < 0.01) and fibrosis was greater (sevenfold, P < 0.01) in eWAT with older age. The old also exhibited greater liver triglycerides (∼60%, P < 0.05). The mitochondrial respiratory oxygen flux after the addition of glutamate and malate (GM), adenosine diphosphate (d), succinate (S) and octanoyl carnitine (O) were one- to twofold higher in eWAT of old mice (P < 0.05). Despite no change in the respiratory control ratio, substrate control ratios of GMOd/GMd and GMOSd/GMd were ∼30-40% lower in old mice (P < 0.05) and were concomitant with increased nitrotyrosine (P < 0.05) and reduced expression of brown adipose markers (P < 0.05). Ageing reduced vascularity (∼50%, P < 0.01), angiogenic capacity (twofold, P < 0.05) and expression of vascular endothelial growth factor (∼50%, P < 0.05) in eWAT. Finally, endothelium-dependent dilation was lower (P < 0.01) in isolated arteries from eWAT arteries of the old mice. Thus, metabolic dysfunction with advancing age occurs in concert with dysfunction in the adipose tissue characterized by both mitochondrial and arterial dysfunction.

Topics: Adipose Tissue; Aging; Animals; Arteries; Body Weight; Carnitine; Glutamic Acid; Malates; Male; Mice; Mitochondria; Neovascularization, Physiologic; Oxidative Stress; Oxygen Consumption; Succinic Acid; Triglycerides; Tyrosine; Vasodilation

The present study examined whether hemin could prevent the development of high-fat diet-induced insulin resistance in the liver and skeletal muscle using a hyperinsulinemic-euglycemic clamp. A four-week high-fat feeding to mice increased the body weight, fat mass, and plasma levels of insulin and lipid, which were reduced by hemin. High-fat diet reduced whole body glucose uptake, which were increased by hemin. Insulin-stimulated hepatic glucose production (HGP) was increased by high-fat diet, but hemin had no significant effect on HGP. Skeletal muscle glucose uptake was reduced by high-fat diet, and hemin normalized the glucose uptake. High-fat diet increased triglyceride levels and mRNA levels of lipogenic enzymes, and decreased mRNA levels of enzymes involved in lipid β-oxidation, which was reversed by hemin. Phosphorylated AMP-activated protein kinase levels were increased in the skeletal muscle of high fat-fed hemin-injected mice. High-fat diet reduced mRNA levels of antioxidant enzymes and increased mRNA levels of inflammatory cytokines and nitrotyrosine levels, which was normalized by hemin in the skeletal muscle. However, hemin had no significant effect on these factors in the liver. These results suggest that hemin prevents the development of high-fat diet-induced insulin resistance by increased insulin sensitivity in the skeletal muscle.

Topics: Adipose Tissue; AMP-Activated Protein Kinases; Animals; Body Weight; Cytokines; Depression, Chemical; Diet, High-Fat; Gene Expression; Glucose; Glucose Clamp Technique; Glutathione Peroxidase; Glutathione Peroxidase GPX1; Heme Oxygenase (Decyclizing); Hemin; Hyperinsulinism; Hyperlipidemias; Insulin Resistance; Liver; Male; Mice, Inbred C57BL; Muscle, Skeletal; Superoxide Dismutase; Triglycerides; Tyrosine

Atherosclerosis and vascular calcification are major contributors to cardiovascular morbidity and mortality among chronic kidney disease patients. The mevalonate pathway may play a role in this vascular pathology. Farnesyltransferase inhibitors such as R115777 block one branch of mevalonate pathway. We studied the effects of farnesyltransferase inhibitor R115777 on vascular disease in apolipoprotein E deficient mice with chronic renal failure and on mineral deposition in vitro.. Female uremic and non-uremic apolipoprotein E deficient mice were randomly assigned to four groups and treated with either farnesyltransferase inhibitor R115777 or vehicle. Farnesyltransferase inhibitor R115777 inhibited protein prenylation in mice with chronic renal failure. It decreased aortic atheromatous lesion area and calcification in these animals, and reduced vascular nitrotyrosine expression and total collagen as well as collagen type I content. Proteomic analysis revealed that farnesyltransferase inhibitor corrected the chronic renal failure-associated increase in serum apolipoprotein IV and α globin, and the chronic renal failure-associated decrease in serum fetuin A. Farnesyltransferase inhibitor further inhibited type I collagen synthesis and reduced mineral deposition in vascular smooth muscle cells in vitro, probably involving Ras-Raf pathway.. We show for the first time that farnesyltransferase inhibition slows vascular disease progression in chronic renal failure by both indirect systemic and direct local actions. This beneficial effect was mediated via a reduction in oxidative stress and favorable changes in vasoprotective peptides.

Topics: Animals; Aorta; Apolipoproteins E; Apoptosis; Atherosclerosis; Blood Proteins; Body Weight; Collagen Type I; Enzyme Inhibitors; Farnesyltranstransferase; Female; Kidney Failure, Chronic; Liver; Macrophages; Mevalonic Acid; Mice; Mice, Knockout; Muscle, Smooth, Vascular; Prenylation; Quinolones; Random Allocation; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Tyrosine; Uremia; Vascular Calcification

Evidence for an important role for Na(+)/H(+) exchangers in diabetic complications is emerging. The aim of this study was to evaluate whether Na(+)/H(+) exchanger 1 inhibition reverses experimental peripheral diabetic neuropathy. Control and streptozotocin-diabetic rats were treated with the specific Na(+)/H(+) exchanger 1 inhibitor cariporide for 4 wk after 12 wk without treatment. Neuropathy end points included sciatic motor and sensory nerve conduction velocities, endoneurial nutritive blood flow, vascular reactivity of epineurial arterioles, thermal nociception, tactile allodynia, and intraepidermal nerve fiber density. Advanced glycation end product and markers of oxidative stress, including nitrated protein levels in sciatic nerve, were evaluated by Western blot. Rats with 12-wk duration of diabetes developed motor and sensory nerve conduction deficits, thermal hypoalgesia, tactile allodynia, and intraepidermal nerve fiber loss. All these changes, including impairment of nerve blood flow and vascular reactivity of epineurial arterioles, were partially reversed by 4 wk of cariporide treatment. Na(+)/H(+) exchanger 1 inhibition was also associated with reduction of diabetes-induced accumulation of advanced glycation endproduct, oxidative stress, and nitrated proteins in sciatic nerve. In conclusion, these findings support an important role for Na(+)/H(+) exchanger 1 in functional, structural, and biochemical manifestations of peripheral diabetic neuropathy and provide the rationale for development of Na(+)/H(+) exchanger 1 inhibitors for treatment of diabetic vascular and neural complications.

Topics: Aldehydes; Animals; Arterioles; Behavior, Animal; Blood Glucose; Blotting, Western; Body Weight; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Diabetic Neuropathies; Glycation End Products, Advanced; Guanidines; Male; Nerve Fibers; Nitrates; Pain Measurement; Pyruvaldehyde; Rats; Rats, Wistar; Reduced Folate Carrier Protein; Sciatic Nerve; Skin; Sulfones; Superoxides; Tyrosine

Oxidative stress is one of the main contributors in the pathogenesis of diabetic retinopathy. The aim of this study is to investigate the effects of SS31 which is a mitochondria-targeted antioxidant peptide on the retinas of streptozotocin (STZ)-induced diabetic rats. Two weeks after induction of diabetes, SS31 (3 mg/kg) or the same volume of normal saline (N.S) was injected subcutaneously into the back of diabetic rats every day. Four months later, the integrity of inner blood retinal barrier (iBRB) was measured by Evans blue perfusion. The expression and distribution of claudin-5, occludin, acrolein, 8-OHdG and nitrotyrosine in the rat retinas were detected by immunofluorescent staining. Retinal ultrastructures were observed by transmission electron microscopy. The protein level of VEGFR2, Trx-2, Bcl-2, Bax, caspase-3, p53, and NF-κB in the rat retinas were assayed by western blot. Four months after subcutaneous injection, the diabetic rats treated with SS31 had better structures of retinal ganglion cells, thinner capillary basement membrane, less iBRB leakage, more uniform staining of claudin-5 and occludin in the retinal vessels, lower levels of acrolein, 8-OHdG, nitrotyrosine, Bax, caspase-3, p53, and NF-κB, and higher levels of Trx-2 and Bcl-2 in the retinas than those treated with N.S. In conclusion, SS31 could protect the retinal structures and inhibit the breakdown of iBRB by reducing oxidative damage, increasing Trx-2 and Bcl-2 expression, and decreasing p53, NF-κB, Bax, caspase-3, and VEGFR2 expression in the retinas of diabetic rats. SS31 could be a potential new treatment for diabetic retinopathy and other oxidative stress-related diseases.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acrolein; Animals; Antioxidants; Blood Glucose; Blood-Retinal Barrier; Body Weight; Capillaries; Claudin-5; Deoxyguanosine; Diabetes Mellitus, Experimental; Endoplasmic Reticulum; Male; Mitochondria; Occludin; Oligopeptides; Oxidative Stress; Protective Agents; Protein Transport; Rats; Rats, Sprague-Dawley; Retina; Signal Transduction; Tyrosine; Vascular Endothelial Growth Factor Receptor-2

Little is known about the anti-colitis effect of tryptophan or its metabolites. Here, the protective effect and its mechanism of tryptophan administration on dextran sulfate sodium -induced colitis in mice was studied.. Twenty C57black6 female mice were equally divided into the control group, and treatment group. The control group received a standard CE-2 diet, while the tryptophan group received a CE-2 diet containing 0.5% l-tryptophan. After one week on this diet, all mice were orally administered a solution of 3.5% dextran sulfate sodium for 12 days to induce colitis. Changes in body weight and bloody stool frequency were monitored during dextran sulfate sodium administration. At 12 days post initial dextran sulfate sodium administration, all mice were sacrificed and the histology of their colonic tissue was examined. The nitrotyrosine levels in colonic tissues in both groups, and nitrate and nitrite levels in the urine of the control group, the tryptophan group and the group of mice without dextran sulfate sodium administration was measured.. The tryptophan group showed significantly attenuated body weight loss, bloody stool frequency and ameliorated histological changes of colitis. While tryptophan treatment significantly reduced nitrotyrosine level in the colonic tissues, there was no significant reduction in urine nitrate and nitrite levels compared with the (dextran sulfate sodium-induced) control group.. Tryptophan treatment ameliorated dextran sulfate sodium-induced colitis in this study. One of the anti-colitis mechanisms of tryptophan treatment is attributable to an anti-nitration effect, and may not be via the suppression of nitric oxide generation.

Topics: Animal Feed; Animals; Antidepressive Agents, Second-Generation; Body Weight; Colitis, Ulcerative; Dextran Sulfate; Disease Models, Animal; Female; Intestinal Mucosa; Mice; Mice, Inbred C57BL; Nitric Oxide; Oxidative Stress; Random Allocation; Tryptophan; Tyrosine

In the present study, we investigated the effects of 7 and 14 days of re-loading following 14-day muscle unweighting (hindlimb suspension, HS) on iron transport, non-heme iron levels and oxidative damage in the gastrocnemius muscle of young (6 months) and old (32 months) male Fischer 344×Brown Norway rats. Our results demonstrated that old rats had lower muscle mass, higher levels of total non-heme iron and oxidative damage in skeletal muscle in comparison with young rats. Non-heme iron concentrations and total non-heme iron amounts were 3.4- and 2.3-fold higher in aged rats as compared with their young counterparts, respectively. Seven and 14 days of re-loading was associated with higher muscle weights in young animals as compared with age-matched HS rats, but there was no difference in muscle weights among aged HS, 7 and 14 days of re-loading rats, indicating that aged rats may have a lower adaptability to muscle disuse and a lower capacity to recover from muscle atrophy. Protein levels of cellular iron transporters, such as divalent metal transport-1 (DMT1), transferrin receptor-1 (TfR1), Zip14, and ferroportin (FPN), and their mRNA abundance were determined. TfR1 protein and mRNA levels were significantly lower in aged muscle. Seven and 14 days of re-loading were associated with higher TfR1 mRNA and protein levels in young animals in comparison with their age-matched HS counterparts, but there was no difference between cohorts in aged animals, suggesting adaptive responses in the old to cope with iron deregulation. The extremely low expression of FPN in skeletal muscle might lead to inefficient iron export in the presence of iron overload and play a critical role in age-related iron accumulation in skeletal muscle. Moreover, oxidative stress was much greater in the muscles of the older animals measured as 4-hydroxy-2-nonhenal (HNE)-modified proteins and 8-oxo-7,8-dihydroguanosine levels. These markers remained fairly constant with either HS or re-loading in young rats. In old rats, HNE-modified proteins and 8-oxo-7,8-dihydroguanosine levels were markedly higher in HS and were lower after 7 days of recovery. However, no difference was observed following 14 days of recovery between control and re-loading animals. In conclusion, advanced age is associated with disruption of muscle iron metabolism which is further perturbed by disuse and persists over a longer time period.

Topics: Animals; Body Weight; Cation Transport Proteins; DNA; GPI-Linked Proteins; Hemochromatosis Protein; Hindlimb Suspension; Homeostasis; Iron; Lipid Peroxidation; Male; Membrane Proteins; Muscle, Skeletal; Muscular Atrophy; Oxidation-Reduction; Oxidative Stress; Rats; Rats, Inbred F344; Receptors, Transferrin; Recovery of Function; RNA; Tyrosine

Post-translational oxidative protein modifications which are more marked during aging and/or high-calorie (HC) diets affect protein function and metabolism. Protein function and metabolism are different according to the type of muscle proteins. Oxidative muscle protein modifications may thus be associated with age-related sarcopenia, and HC may be implicated in the development of sarcopenia by emphasizing protein modifications. Understanding the role of protein modifications in the process of sarcopenia and metabolism associated with a high fat diet may be elucidated by investigations with skeletal muscle protein subfractionations. To study this hypothesis, carbonylated protein (CP) and 3-nitrotyrosine (3-NT) levels were measured in mixed, sarcoplasmic, myofibrillar and mitochondrial protein fractions of quadriceps in rats aged 6months (A) and 25months (O) fed a normal calorie (NC) or HC diet for 3months (AN, AH, ON, OH n=7-8). Muscle weight was lower in the older rats (AN: 0.79±0.03g, ON: 0.43±0.12g, P<0.05), but no HC effect was observed. CP did not differ between groups while 3-NT accumulated significantly in ON compared with AN, especially in mitochondria (2.4±0.5, 1.3±0.1, 1.9±0.4, 2.9±1.2 -fold in mixed, sarcoplasmic, myofibrillar and mitochondrial fractions respectively, P<0.05). 3-NT in mixed protein was negatively correlated with muscle mass (r(2)=-0.812). 3-NT accumulation during HC was observed only in specific proteins of mitochondria (100kDa) (1.0±0.6, 1.7±0.9, 3.3±1.4 and 7.0±2.5 -fold in AN, AH, ON and OH, respectively, P<0.05). Hence cumulative 3-NT in skeletal muscle protein appears associated with the development of age-related muscle loss. Mitochondrial proteins are more prone to nitration during aging and nutritional stress.

Topics: Aging; Animals; Biomarkers; Body Weight; Energy Intake; Hindlimb; Mitochondria, Muscle; Mitochondrial Proteins; Muscle Proteins; Muscle, Skeletal; Myofibrils; Organ Size; Protein Carbonylation; Quadriceps Muscle; Rats; Rats, Wistar; Sarcopenia; Tyrosine

Chronic hypoxia increases the expression of inducible nitric oxide synthase (iNOS) mRNA and protein levels in fetal guinea pig heart ventricles. Excessive generation of nitric oxide (NO) can induce nitrosative stress leading to the formation of peroxynitrite, which can upregulate the expression of matrix metalloproteinases (MMPs). This study tested the hypothesis that maternal hypoxia increases fetal cardiac MMP9 and collagen through peroxynitrite generation in fetal hearts.. In heart ventricles, levels of malondialdehyde, 3-nitrotyrosine (3-NT), MMP9, and collagen were increased in hypoxic (HPX) vs. normoxic (NMX) fetal guinea pigs.. Thus, maternal hypoxia induces oxidative-nitrosative stress and alters protein expression of the extracellular matrix (ECM) through upregulation of the iNOS pathway in fetal heart ventricles. This identifies iNOS-derived NO as an important stimulus for initiating the adverse effects of peroxynitrite in HPX fetal hearts.. Pregnant guinea pigs were exposed to normoxia (room air) or hypoxia (10.5% O(2), 14 d) before term (term ≈ 65 d) and administered water, L-N6-(1-iminoethyl)-lysine (LNIL), an iNOS inhibitor, or N-acetylcysteine (NAC), an antioxidant.

Topics: Acetylcysteine; Animals; Body Weight; Collagen; Female; Fetal Heart; Guinea Pigs; Hypoxia; Lipid Peroxidation; Matrix Metalloproteinase 9; Nitric Oxide Synthase Type II; Organ Size; Peroxynitrous Acid; Pregnancy; Tyrosine

To observe the effect of Jinmaitong capsule (JMT), a compound traditional Chinese medicine, on expressions of inducible nitric oxide synthase (iNOS) and nitro tyrosine (NT) protein in streptozocin-induced diabetic (STZ-DM) rats.. Intraperitoneal injection of streptozocin in rats to establish a model. STZ-DM rats were randomly divided into the model control group (distilled water), the small-dose JMT group (JMT at dose of 0.45 g x kg(-1) x d(-1)), the medium-dose JMT group (JMT at dose of 0.88 g x kg(-1) x d(-1)), the large-dose JMT group (JMT at dose of 1.75 g x kg(-1) x d(-1)) and Vitamin C group (VC at dose of 0.05 g x kg(-1) x d(-1)). Ten normal rats with matching weight and age were selected as the normal control group (distilled water). After intragastric administration for 16 weeks, the expressions of iNOS and NT in sciatic nerve were detected by the immunohistochemistry method.. The expression levels of iNOS and NT protein in diabetic rats were higher than those in normal rats (P<0.05, P<0.01). Compared with the model group, the levels of iNOS and NT protein in JMT and VC groups were significantly decreased (P<0.05, P<0.01). Particularly, the medium-dose JMT group showed a better effect than the VC group (P<0.05, P<0.01).. JMT could down-regulate the expressions of iNOS and NT protein of sciatic nerve in diabetic rats.

Topics: Animals; Blood Glucose; Body Weight; Capsules; Diabetes Mellitus, Experimental; Diabetic Neuropathies; Drugs, Chinese Herbal; Enzyme Activation; Male; Nitric Oxide Synthase Type II; Rats; Rats, Sprague-Dawley; Sciatic Nerve; Tyrosine

Chronic inflammation accompanied by arginine deficiency, immune dysfunction, and excess nitric oxide (NO) production is a clinical condition found in patients with peritonitis. A previous study showed that the nonselective NOS inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) may facilitate the metabolism of the immune nutrient arginine without altering NO homeostasis in rats with sub-acute peritonitis. Here, we investigated the effects of L-NAME on the immunocytic subpopulation distribution and response.. Male Wistar rats with cecal puncture-induced peritonitis were administered parenteral nutrition solutions supplemented with 0 (CPP group), 5 (LNA group), 25 (MNA group) or 50 (HNA group) mg · kg(-1) · day(-1) of L-NAME for 7 days. Parenteral-fed sham-operated rats (TPN group) and orally-fed healthy rats (R group) were included as controls.. The TPN group had significantly increased spleen weights and levels of plasma nitrite/nitrate (NOx), circulating white blood cells (WBC), and splenocytic T cells, as well as significantly decreased levels of cytotoxic T- and B-leukocytes and B-splenocytes compared to the R group. The CPP group had significantly decreased levels of plasma NOx and concanavalin (Con) A-stimulated interferon (IFN)-γ and interleukin (IL)-2 production by leukocytes and significantly increased production of Con A-stimulated tumor necrosis factor (TNF)-α and lipopolysaccharide (LPS)-stimulated IFN-γ in the leukocytes. In addition, the LNA and MNA groups had significantly decreased spontaneous IL-6 and Con A-stimulated TNF-α and IFN-γ production by the leukocytes while the HNA group had significantly increased LPS-stimulated TNF-α and Con A-stimulated IFN-γ and IL-2 production by the splenocytes compared to the CPP group.. Low-dose L-NAME infusion may suppress proinflammatory and T-helper-1 (Th1) response in leukocytes, and high-dose infusion may activate the proinflammatory response in splenic macrophages and Th1 response in T-splenocytes in rats with sub-acute peritonitis.

Topics: Acute Disease; Animals; Arginine; Body Weight; Cell Proliferation; Cytokines; Dose-Response Relationship, Drug; Humans; Immunomodulation; Infusions, Parenteral; Leukocyte Count; Leukocytes; Male; Nitrates; Nitric Oxide Synthase; Nitrites; Organ Size; Parenteral Nutrition; Peritonitis; Rats; Rats, Wistar; Spleen; Tyrosine

We recently showed that combination therapy with losartan and pioglitazone provided synergistic effects compared with monotherapy in improving lesions of renal structure and function in Sprague-Dawley rats fed with a high-fat, high-sodium diet and 20% sucrose solution. This study was designed to explore the underlying mechanisms of additive renoprotection provided by combination therapy. Losartan, pioglitazone, and their combination were orally administered for 8 weeks. The increased level of renal malondialdehyde and expression of nicotinamide adenine dinucleotide phosphate oxidase subunit p47(phox) and nitrotyrosine as well as the decreased total superoxide dismutase activity and copper, zinc-superoxide dismutase expression were tangible evidence for the presence of oxidative and nitrative stress in the kidney of model rats. Treatment with both drugs, individually and in combination, improved these abnormal changes. Combination therapy showed synergistic effects in reducing malondialdehyde level, p47(phox), and nitrotyrosine expression to almost the normal level compared with monotherapy. All these results suggest that the additive renoprotection provided by combination therapy might be attributed to a further reduction of oxidative and nitrative stress.

Topics: Animals; Body Weight; Diet, High-Fat; Dietary Sucrose; Drug Synergism; Drug Therapy, Combination; Gene Expression Regulation, Enzymologic; Immunohistochemistry; Kidney; Losartan; Male; Malondialdehyde; NADPH Oxidases; Nitrosation; Oxidation-Reduction; Oxidative Stress; Pioglitazone; Rats; Rats, Sprague-Dawley; RNA, Messenger; Sodium, Dietary; Superoxide Dismutase; Thiazolidinediones; Tyrosine

Accumulation of pro-nerve growth factor (NGF), the pro form of NGF, has been detected in neurodegenerative diseases. However, the role of proNGF in the diabetic retina and the molecular mechanisms by which proNGF causes retinal neurodegeneration remain unknown. The aim of this study was to elucidate the role of proNGF in neuroglial activation and to examine the neuroprotective effects of epicatechin, a selective inhibitor of tyrosine nitration, in an experimental rat model of diabetes.. Expression of proNGF and its receptors was examined in retinas from streptozotocin-induced diabetic rats, and in retinal Müller and retinal ganglion cells (RGCs). RGC death was assessed by TUNEL and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assays in diabetic retinas and cell culture. Nitrotyrosine was determined using Slot-blot. Activation of the tyrosine kinase A (TrkA) receptor and p38 mitogen-activated protein kinase (p38MAPK) was assessed by western blot.. Diabetes-induced peroxynitrite impaired phosphorylation of TrkA-Y490 via tyrosine nitration, activated glial cells and increased expression of proNGF and its receptor, p75 neurotrophin receptor (p75(NTR)), in vivo and in Müller cells. These effects were associated with activation of p38MAPK, cleaved poly-(ADP-ribose) polymerase and RGC death. Treatment of diabetic animals with epicatechin (100 mg kg(-1) day(-1), orally) blocked these effects and restored neuronal survival. Co-cultures of RGCs with conditioned medium of activated Müller cells significantly reduced RGC viability (44%). Silencing expression of p75(NTR) by use of small interfering RNA protected against high glucose- and proNGF-induced apoptosis in RGC cultures.. Diabetes-induced peroxynitrite stimulates p75(NTR) and proNGF expression in Müller cells. It also impairs TrkA receptor phosphorylation and activates the p75(NTR) apoptotic pathway in RGCs, leading to neuronal cell death. These effects were blocked by epicatechin, a safe dietary supplement, suggesting its potential therapeutic use in diabetic patients.

Topics: Animals; Blood Glucose; Blotting, Western; Body Weight; Catechin; Cell Line; In Situ Nick-End Labeling; Male; Nerve Degeneration; Nerve Growth Factor; Rats; Rats, Sprague-Dawley; Receptor, Nerve Growth Factor; Retina; Retinal Ganglion Cells; RNA, Small Interfering; Tyrosine

Asymmetric dimethylarginine (ADMA) is an endogenous modulator of endothelial function and oxidative stress, and increased levels of this molecule have been reported in some metabolic disorders and cardiovascular diseases. The aim of this work was to analyze the time course of dimethylarginine compounds and oxidative stress levels and the relationship between these and cardiovascular function in fructose-hypertensive rats.. 90 male Sprague-Dawley rats were randomized into 2 groups, fed for 3 months with standard (C) chow supplemented or not with fructose (F, 60%). After sacrifice at different weeks (W), the aorta and plasma were harvested to assess the vascular and biochemical parameters. Our work showed that the plasma levels of ADMA in the fructose-fed rats increased after 2 weeks of the diet (1.6 ± 0.3 μM vs. 1.2 ± 0.3 μM, p < 0.05) with no changes in plasma levels of either SDMA or L-arginine and after an increase in glycemia. Levels of vascular oxidative stress, estimated in aortic segments using an oxidative fluorescence technique, were higher in the F group (W2: 1.14 ± 0.2% vs. 0.33 ± 0.02%, p < 0.01). An increase in expression levels of nitrotyrosine (3-fold) and iNOS (2-fold) were noted in the fructose-fed rats. After 1 month, this was associated with a significant increase in NAD(P)H oxidase activity. Concerning vascular function, a 15% decrease in maximal endothelium-dependent relaxation was found in the aorta of the F group. Our work showed that the presence of exogenous L-MMA, an inhibitor of NO synthase, was associated with a significant reduction in endothelium-dependent relaxation in isolated aorta rings of the C group; this effect was not observed in the vessels of fructose-fed rats.. Our findings suggest that the elevated levels of ADMA observed could in part be secondary to the early development of oxidative stress associated with the development of hypertension.

Topics: Animals; Aorta; Arginine; Blood Glucose; Blood Pressure; Body Weight; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Inhibitors; Fructose; Heart Rate; Hypertension; Male; Metabolic Syndrome; NADPH Oxidases; Nitric Oxide Synthase Type II; Oxidative Stress; Rats; Rats, Sprague-Dawley; Time Factors; Tyrosine; Vasodilation; Vasodilator Agents

Olprinone is a specific phosphodiesterase (PDE)-III inhibitor, which has been found to have anti-inflammatory effects in addition to its main inotropic and peripheral vasodilatory effects. In the present study we investigated the effects of olprinone (0.2mg/kg, i.p.) on the development of zymosan-induced multiple organ failure in mice. Treatment with olprinone attenuated the peritoneal exudation and the migration of polymorphonuclear cells caused by zymosan. Olprinone also attenuated the lung, liver and pancreatic injury, renal dysfunction as well as the increased lung and intestine myeloperoxidase (MPO) activity caused by zymosan. Immunohistochemical analysis for inducible nitric oxide synthase (iNOS), nitrotyrosine, poly(ADP-ribose) (PAR), tumor necrosis factor-α (TNF-α) and interleuchin-1β (IL-1β) revealed positive staining in pancreatic and intestinal tissue obtained from zymosan-injected mice. The degree of staining for nitrotyrosine, iNOS, PAR, TNF-α and IL-1β was markedly reduced in tissue sections obtained from zymosan-injected mice, which had received olprinone. In addition, administration of zymosan caused a severe illness in the mice characterized by significant loss of body weight and a 60% of mortality at the end of observation period (7 days). Treatment with olprinone significantly reduced the development of systemic toxicity, loss in body weight and mortality, caused by zymosan. This study provides evidence that olprinone attenuates the degree of zymosan-induced shock in mice.

Topics: Alanine Transaminase; Alkaline Phosphatase; Amylases; Animals; Apoptosis; Ascitic Fluid; Aspartate Aminotransferases; bcl-2-Associated X Protein; Bicarbonates; Bilirubin; Blood Gas Analysis; Body Weight; Cell Count; Creatinine; Fas Ligand Protein; Hydrogen-Ion Concentration; Imidazoles; Intercellular Adhesion Molecule-1; Interleukin-1beta; Kidney; Lipase; Liver; Lung; Male; Mice; Mice, Inbred Strains; Multiple Organ Failure; Nitrates; Nitrites; P-Selectin; Pancreas; Phosphodiesterase 3 Inhibitors; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Proto-Oncogene Proteins c-bcl-2; Pyridones; Survival Analysis; Tumor Necrosis Factor-alpha; Tyrosine; Zymosan

Glutamine is the most abundant amino acid in the bloodstream. It is important in nucleotide synthesis, is anti-catabolic, has anti-oxidant properties via metabolism to glutathione, may enhance immune responsiveness and possesses immunoregulatory functions. Moreover, it reduces atrophy of intestinal mucosa in animals on total parenteral nutrition and prevents intestinal mucosal injury accompanying small bowel transplantation, chemotherapy and radiation. In the present study, we investigated the effects of glutamine on development of non-septic shock caused by zymosan. Mice received either zymosan (500 mg/kg, administered i.p., as a suspension in saline) or vehicle (saline). Glutamine (1.5 mg/kg i.p.) was administered 1 and 6h after zymosan administration. Organ failure and systemic inflammation in mice were assessed 18 h after administration of zymosan and/or glutamine. Glutamine-treatment reduced the peritoneal exudation and the migration of polymorphonuclear cells caused by zymosan-injection and also attenuated the pancreatic and gut injury. Inflammatory and apoptotic parameters were evaluated to better investigate the effects of the glutamine-administration. So, by immunohistochemical analysis and in vitro assays, we have clearly showed that glutamine reduces: 1) the histological damage in pancreas and gut; 2) the inducible nitric oxide synthase expression; 3) nitrotyrosine and poly (ADP-ribose) formation; 4) TNF-α and IL-1β tissue and plasma levels; 5) FasL localization; and 6) alteration of the balance between Bax and Bcl-2. In addition, at the end of the observation period (7 days), zymosan causes severe illness in the mice characterized by a systemic toxicity, significant loss of body weight and mortality. Glutamine-treatment significantly reduced all these parameters.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Body Weight; Cytokines; Enzyme Activation; Fas Ligand Protein; Gene Expression Regulation; Glutamine; Inflammation; Lipid Peroxidation; Male; Mice; Multiple Organ Failure; Neutrophil Infiltration; Nitric Oxide; Peritoneal Cavity; Poly Adenosine Diphosphate Ribose; Poly(ADP-ribose) Polymerases; Protein Transport; Proto-Oncogene Proteins c-bcl-2; Tyrosine; Zymosan

With the consideration of the multifactorial etiology of diabetic peripheral neuropathy, an ideal drug or drug combination should target at least several key pathogenetic mechanisms. The flavonoid baicalein (5,6,7-trihydroxyflavone) has been reported to counteract sorbitol accumulation, activation of 12/15-lipoxygenase, oxidative-nitrosative stress, inflammation, and impaired signaling in models of chronic disease. This study evaluated baicalein on diabetic peripheral neuropathy. Control and streptozotocin-diabetic C57Bl6/J mice were maintained with or without baicalein treatment (30 mg kg(-1) d(-1), i.p., for 4 weeks after 12 weeks without treatment). Neuropathy was evaluated by sciatic motor and hind-limb digital sensory nerve conduction velocities, thermal algesia (Hargreaves test), tactile response threshold (flexible von Frey filament test), and intraepidermal nerve fiber density (fluorescent immunohistochemistry with confocal microscopy). Sciatic nerve and spinal cord 12/15-lipoxygenase and total and phosphorylated p38 mitogen-activated protein kinase expression and nitrated protein levels were evaluated by Western blot analysis, 12(S)hydroxyeicosatetraenoic acid concentration (a measure of 12/15-lipoxygenase activity) by ELISA, and glucose and sorbitol pathway intermediate concentrations by enzymatic spectrofluorometric assays. Baicalein did not affect diabetic hyperglycemia, and alleviated nerve conduction deficit and small sensory nerve fiber dysfunction, but not intraepidermal nerve fiber loss. It counteracted diabetes-associated p38 mitogen-activated protein kinase phosphorylation, oxidative-nitrosative stress, and 12/15-lipoxygenase overexpression and activation, but not glucose or sorbitol pathway intermediate accumulation. In conclusion, baicalein targets several mechanisms implicated in diabetic peripheral neuropathy. The findings provide rationale for studying hydroxyflavones with an improved pharmacological profile as potential treatments for diabetic neuropathy and other diabetic complications.

Topics: Animals; Antioxidants; Arachidonate 12-Lipoxygenase; Arachidonate 15-Lipoxygenase; Blood Glucose; Body Weight; Diabetes Mellitus, Experimental; Diabetic Neuropathies; Disease Models, Animal; Flavanones; Gene Expression Regulation; Hyperalgesia; Mice; Mice, Inbred C57BL; Neural Conduction; Oligonucleotides, Antisense; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Reaction Time; Receptors, Eicosanoid; Sciatic Nerve; Spinal Cord; Statistics, Nonparametric; Tyrosine

The extracellular signal-regulated kinase (ERK) cascade has long been known to be central to the activation of cellular processes such as proliferation, differentiation, and oncogenic transformation. The mitogen-activated protein (MAP) serine/threonine family of protein kinases, of which ERK is a member, is activated by a mechanism that includes protein kinase cascades. Mitogen-activated protein kinases (MAPKs) are well-conserved enzymes connecting cell surface receptors to intracellular regulatory targets; they are activated in response to a wide variety of stimuli. The aim of this study was to investigate the effects of PD98059, a highly selective inhibitor of MAP/ERK kinase1 (MEK1) activation, on the development of lung inflammation and fibrosis. Lung injury was induced by intratracheal instillation of bleomycin (1 mg/kg), and PD98059 (10 mg/kg, 10% dimethyl sulfoxide, i.p.) was administrated 1 h after bleomycin instillation and daily for 7 days. PD98059 treatment shows therapeutic effects on pulmonary damage, decreasing many inflammatory and apoptotic parameters, such as (1) cytokine production; (2) IkBα degradation and NF-kB nuclear translocation; (3) iNOS expression; (4) nitrotyrosine and PAR localization; and (5) the degree of apoptosis, as evaluated by Bax and Bcl-2 balance, FAS ligand expression, and terminal deoxynucleotidyl transferase dUTP nick-end labeling staining. In particular, to assess whether PD98059 treatment influences MAPKs pathway, we have also investigated the expression of activated ERK and JNK after bleomycin-induced pulmonary fibrosis, showing that the inhibition of the cascade reduces the inflammatory processes that lead to the appearance of the fibrosis. Taken together, all our results clearly show that PD98059 reduces the lung injury and inflammation due to the intratracheal bleomycin administration in mice.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Bleomycin; Body Weight; Butadienes; Fas Ligand Protein; Flavonoids; I-kappa B Proteins; Instillation, Drug; Interleukin-1beta; JNK Mitogen-Activated Protein Kinases; Lung; Male; MAP Kinase Kinase 1; Mice; Mice, Inbred Strains; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Neutrophils; NF-KappaB Inhibitor alpha; Nitric Oxide Synthase Type II; Nitriles; Phosphorylation; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-bcl-2; Pulmonary Edema; Pulmonary Fibrosis; Transcription Factor RelA; Tumor Necrosis Factor-alpha; Tyrosine

Endothelial dysfunction is a hallmark of hypertension and vascular oxidative stress can contribute to endothelial dysfunction and hypertension development. Resveratrol is an antioxidant polyphenol which improves endothelium dependent relaxation, the mechanisms of which are unknown. Also, the role of resveratrol in hypertension remains to be established. The purpose of this study was to investigate the mechanisms of resveratrol induced improvement of endothelial function and establish its role in hypertension. SHR and WKY rats, 3-4 weeks old, were treated with resveratrol in drinking water for 10 weeks, untreated SHR and WKY rats served as controls. At the end of the treatment, control SHR exhibited increased blood pressure, oxidative stress and attenuated endothelium dependent relaxation in comparison to WKY rats. The impaired endothelium function in SHR was associated with lower nitrite/nitrate levels, elevated nitrotyrosine content and eNOS uncoupling. Resveratrol treatment attenuated hypertension development in SHR as indicated by lower blood pressure in resveratrol treated SHR (SHR-R) compared to control SHR. SHR-R also exhibited reduced H(2)O(2) content and elevated superoxide dismutase activity. Resveratrol treatment normalized endothelium dependent vasorelaxation in SHR. In parallel, resveratrol restored nitrite/nitrate levels and normalized nitrotyrosine content in SHR. SHR exhibited increased l-arginine dependent superoxide production which was blocked by NOS inhibitor l-NNA, suggesting eNOS uncoupling. eNOS uncoupling was prevented by resveratrol treatment. In conclusion, early treatment with resveratrol lowers oxidative stress, preserves endothelial function and attenuates development of hypertension in SHR. More importantly, prevention of eNOS uncoupling and NO scavenging could represent novel mechanisms for resveratrol-mediated antihypertensive effects.

Topics: Animals; Antioxidants; Blood Pressure; Body Weight; Drinking; Eating; Endothelium, Vascular; Heart; Hypertension; Male; Nitric Oxide; Nitric Oxide Synthase Type III; Organ Size; Oxidative Stress; Rats; Rats, Inbred SHR; Resveratrol; Stilbenes; Tyrosine

We have previously shown that treating streptozotocin-induced diabetic rats, an animal model of type 1 diabetes, with Ilepatril (an inhibitor of neutral endopeptidase and angiotensin converting enzyme (ACE)) improves vascular and neural functions. In this study we sought to determine the effect of Ilepatril treatment of high fat fed/low dose streptozotocin-diabetic rats, a model for type 2 diabetes, on vascular and neural complications. Following 8 weeks on a high fat diet rats were treated with 30 mg/kg streptozotocin (i.p.) and after 4 additional weeks a group of these rats was treated for 12 weeks with Ilepatril followed by analysis of neural and vascular functions. Included in these studies were age-matched control rats and rats fed a high fat diet and treated with or without Ilepatril. Diabetic and diet induced obese rats have characteristics of insulin resistance, slowing of nerve conduction velocity, thermal hypoalgesia, reduction in intraepidermal nerve fiber density in the hindpaw and impairment in vascular relaxation to acetylcholine and calcitonin gene-related peptide in epineurial arterioles of the sciatic nerve. Treatment with Ilepatril was efficacious in improving all of these endpoints although improvement of insulin resistance in diabetic rats was minimal. These studies suggest that dual inhibition of angiotensin converting enzyme and neutral endopeptidase activity of type 2 diabetic rats is an effective approach for treatment of diabetic neural and vascular complications.

Topics: Adipose Tissue; Animals; Arterioles; Blood Glucose; Blood Vessels; Body Weight; Carbohydrate Metabolism; Diabetes Complications; Diabetes Mellitus, Experimental; Diet, High-Fat; Dose-Response Relationship, Drug; Enzyme Inhibitors; Glucose Tolerance Test; Glutathione; Heterocyclic Compounds, 3-Ring; Insulin; Lens, Crystalline; Leptin; Male; Muscle, Skeletal; Nerve Fibers; Nervous System; Nociception; Organ Size; Rats; Rats, Sprague-Dawley; Sciatic Nerve; Superoxides; Thiobarbiturates; Tyrosine; Vasodilation

Diabetic nephropathy is associated with cardiovascular morbidity. Angiotensin-converting enzyme (ACE) inhibitors provide imperfect renoprotection in advanced type 2 diabetes, and cardiovascular risk remains elevated. Endothelin (ET)-1 has a role in renal and cardiac dysfunction in diabetes. Here, we assessed whether combination therapy with an ACE inhibitor and ET(A) receptor antagonist provided reno- and cardioprotection in rats with overt type 2 diabetes. Four groups of Zucker diabetic fatty (ZDF) rats were treated orally from 4 (when proteinuric) to 8 mo with vehicle, ramipril (1 mg/kg), sitaxsentan (60 mg/kg), and ramipril plus sitaxsentan. Lean rats served as controls. Combined therapy ameliorated proteinuria and glomerulosclerosis mostly as a result of the action of ramipril. Simultaneous blockade of ANG II and ET-1 pathways normalized renal monocyte chemoattractant protein-1 and interstitial inflammation. Cardiomyocyte loss, volume enlargement, and capillary rarefaction were prominent abnormalities of ZDF myocardium. Myocyte volume was reduced by ramipril and sitaxsentan, which also ameliorated heart capillary density. Drug combination restored myocardial structure and reestablished an adequate capillary network in the presence of increased cardiac expression of VEGF/VEGFR-1, and significant reduction of oxidative stress. In conclusion, in type 2 diabetes concomitant blockade of ANG II synthesis and ET-1 biological activity through an ET(A) receptor antagonist led to substantial albeit not complete renoprotection, almost due to the ACE inhibitor. The drug combination also showed cardioprotective properties, which however, were mainly dependent on the contribution of the ET(A) receptor antagonist through the action of VEGF.

Topics: Angiotensin-Converting Enzyme Inhibitors; Animals; Blood Glucose; Body Weight; Cell Count; Collagen Type III; Diabetes Mellitus, Type 2; Endothelin A Receptor Antagonists; Endothelin-1; Heart; Hemodynamics; Immunohistochemistry; Kidney; Kidney Function Tests; Male; Muscle Cells; Myocardium; Rats; Rats, Zucker; Real-Time Polymerase Chain Reaction; Receptors, Vascular Endothelial Growth Factor; Survival; Tyrosine; Vascular Endothelial Growth Factor A

Kurozu has been reported to ameliorate colitis in mice and to have an anti-oxidative effect. However, the active components and mechanism of action remain unknown. Here, as a first step to identify the active components, we chromatographically fractionated Kurozu and investigated the anti-colitis activity of the fractions, focusing on anti-nitration activity.. Kurozu was divided into 4 molecular-weight fractions (fraction I, >4,000 daltons; II, 2,000~4,000 daltons; III, 800~2,000 daltons; IV, <800 daltons). Forty C57black6 mice were divided into 5 groups as follows: the control group received standard CE-2 diet, and Groups I~IV received CE-2 diet containing Kurozu fractions I~IV, respectively. Dextran sulfate sodium was administered to the mice for 12 days to induce colitis. Body weight and bloody stool frequency were monitored as indices of severity of colitis after administration of dextran sulfate sodium, and at 12 days, all mice were sacrificed for examination of colonic pathology and nitrotyrosine production in the colon tissues.. Colitis was markedly ameliorated in Group III, followed by Group II, while Group IV showed little difference from the control. The colonic nitrotyrosine level in Group III was significantly reduced compared with the control.. The major protective components in Kurozu appear to have molecular weights in the range of 800~4,000 daltons, and their action appears to be related, at least in part, to anti-oxidative and anti-nitration effects.

Topics: Acetic Acid; Animals; Body Weight; Colitis; Colon; Dextran Sulfate; Female; Mice; Mice, Inbred C57BL; Molecular Weight; Occult Blood; Oryza; Oxidative Stress; Tyrosine

To determine if the circulating nitrotyrosine level significantly correlates with parameters measuring cognitive abilities.. One-hundred and twelve community-living subjects (ranging in age from 27 to 98 years) were evaluated for cognitive abilities [Mini Mental State Examination (MMSE) score] and circulating free nitrotyrosine plasma level, as well as for several variables that might influence cognitive abilities (age, education) and nitrotyrosine level (body mass index, haematological parameters, cardiovascular and inflammatory indices).. In the sub-group of cognitively impaired subjects (score at MMSE < 23.9), but not in that of cognitively not impaired subjects, a significant inverse correlation exists between nitrotyrosine level and MMSE score (r = -0.378; P < 0.02).. The finding, if confirmed by longitudinal studies, could play a role in the management of the subjects with Mild Cognitive Impairment, the clinical condition considered as a transitional state between the changes of cognitive ability in normal aging and dementia.

Topics: Adult; Aged; Aged, 80 and over; Blood Cell Count; Body Weight; Cognition Disorders; Enzyme-Linked Immunosorbent Assay; Female; Hemoglobins; Humans; Male; Mental Status Schedule; Middle Aged; Neuropsychological Tests; Residence Characteristics; Sex Factors; Tyrosine

Oxidative stress resulting in excessive generation of ROS is a compelling initiator of DNA damage along with damage to various cellular proteins and other macromolecules. Poly(ADP-ribose) polymerase (PARP) activation in response to DNA damage, stirs an energy-consuming cellular metabolic cycle; culminating into cell death. The present study was designed to determine the effect of combining an antioxidant, Melatonin and a PARP inhibitor, Nicotinamide on the hallmark deficits developing in diabetic neuropathy (DN). Streptozotocin (STZ, 55 mg/kg, i.p.) was administered to induce diabetes. Six weeks post diabetes induction, two week treatment with Melatonin (3 and 10 mg/kg) and Nicotinamide (100 and 300 mg/kg) either alone or in combination was given. Effect of these interventions on the functional, behavioral and biochemical changes caused by hyperglycemia were studied in treated animals. Melatonin and Nicotinamide alone as well as in combination ameliorated the functional deficits along with improvement in pain parameters. The combination also demonstrated an essential reversal of biochemical alterations. Nitrotyrosine and Poly ADP Ribose (PAR) immunopositivity was significantly decreased in sciatic nerve micro-sections of treatment group. The results of this study advocate that simultaneous inhibition of oxidative stress-PARP activation cascade may prove useful for the pharmacotherapy of DN.

Topics: Animals; Antioxidants; Blood Glucose; Body Weight; Diabetic Neuropathies; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Interactions; Hyperalgesia; Lipid Peroxidation; Male; Melatonin; Neural Conduction; Niacinamide; Pain Measurement; Pain Threshold; Poly(ADP-ribose) Polymerases; Rats; Rats, Sprague-Dawley; Regional Blood Flow; Sciatic Nerve; Streptozocin; Tyrosine; Vitamin B Complex

Inducible nitric oxide synthase (iNOS) is involved in obesity-induced insulin resistance. Since aging is accompanied by increased iNOS expression, the effect of iNOS gene deletion on aging-associated insulin resistance was investigated in 7-month-old (adult) and 22-month-old (old) iNOS knockout and wild-type mice using the hyperinsulinemic-euglycemic clamp. While body weight and fat mass were increased, muscle mass was reduced with aging in wild-type mice. However, body composition was not changed with aging in iNOS knockout mice due to increased locomotor activity. NO metabolites in plasma, and protein levels of iNOS and nitrotyrosine in skeletal muscle increased with aging in wild-type mice. Deletion of iNOS gene attenuated NO metabolites and nitrotyrosine with aging in iNOS knockout mice. Glucose uptake in whole body and skeletal muscle was reduced with aging in both wild-type and iNOS knockout mice and there was no difference between two groups. Plasma level of tumor necrosis factor-alpha and gene expression of proinflammatory cytokines in peripheral tissues were increased with aging in both groups, and that was more heightened in iNOS knockout mice. These results suggest that lack of iNOS does not prevent aging-associated insulin resistance in mice and heightened production of proinflammatory cytokines may be involved.

Topics: Adipose Tissue; Aging; Animals; Body Composition; Body Weight; Cytokines; Gene Deletion; Glucose Clamp Technique; Hyperinsulinism; Inflammation; Insulin Resistance; Mice; Mice, Inbred C57BL; Mice, Knockout; Motor Activity; Nitric Oxide Synthase Type II; Oxygen Consumption; Polymerase Chain Reaction; Tumor Necrosis Factor-alpha; Tyrosine

Recent basic research and clinical data have provided new insights into the role of glycoxidative and nitrosative stresses (both oxidative stress) in diabetic complications, such as diabetic nephropathy, suggesting a different and innovative approach to antioxidant therapy. In streptozotocin-induced diabetic rat kidney, the present study investigated the effects of the synthetic pyridoindole antioxidant stobadine (STB) on renal total antioxidant potential (AOP) and protein oxidation parameters such as protein carbonyl content (PCC), advanced oxidation protein products (AOPPs) and nitrotyrosine (NT), a marker specific for protein modification by peroxynitrite.. Wistar Albino rats were divided into two groups: normal and streptozotocin-induced diabetic rats. Each group of the animals was further divided into two groups: untreated and treated with stobadine (24.7 mg/kg) during 16 weeks daily by oral gavage.. The renal tissue AOP and the levels of AOPPs, PCC and NT were increased in diabetic rats compared with the untreated control animals. Furthermore, stobadine treatment significantly decreased protein carbonylation and AOPPs but not NT.. These findings indicate that STB is an antioxidant factor which can improve glycoxidative stress markers in kidney, while it has no effect on protein nitrosylation.

Topics: Animals; Antioxidants; Blood Glucose; Body Weight; Carbolines; Diabetes Mellitus, Experimental; Glycation End Products, Advanced; Kidney; Male; Oxidative Stress; Protein Carbonylation; Rats; Rats, Wistar; Tyrosine

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

Thee present study analysed the quantification of rutin in raw buckwheat extract (RBE) and germinated buckwheat extract (GBE) by high performance liquid chromatography (HPLC), and examined changes in body weight, systolic blood pressure (SBP) and nitrotyrosine (a marker for peroxynitrite formation) immunoreactivity in aortic endothelial cells in spontaneously hypertensive rats (SHRs) and normotensive Wistar-Kyoto (WKY) rats after treatment with RBE and GBE for 5 weeks. In the HPLC study, RBE and GBE contained a mean content of rutin of 1.52 +/- 0.21 and 2.92 +/- 0.88 mg/g, respectively. In the 600 mg/kg GBE-treated group, SBP was lower than that in the 600 mg/kg RBE-treated group. The treatment with RBE and/or GBE significantly reduced oxidative damage in aortic endothelial cells by lowering nitrotyrosine immunoreactivity. These results suggest that GBE has an antihypertensive effect and may protect arterial endothelial cells from oxidative stress.

Topics: Animals; Antihypertensive Agents; Aorta; Blood Pressure; Body Weight; Chromatography, High Pressure Liquid; Endothelial Cells; Fagopyrum; Molecular Structure; Oxidative Stress; Plant Extracts; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Rutin; Tyrosine

It is not known whether there are mechanisms linking adipose tissue mass and increased oxidative stress in obesity. This study investigated associations between decreasing general and abdominal fat depots and oxidative stress during weight loss. Subjects were severely obese women who were measured serially at baseline and at 1, 6 (n = 30), and 24 months (n = 18) after bariatric surgery. Total fat mass (FAT) and volumes of visceral (VAT) and subcutaneous abdominal adipose tissue (SAT) were related to plasma concentrations of derivatives of reactive oxidative metabolites (dROMS), a measure of lipid peroxides and oxidative stress. After intervention, BMI significantly decreased, from 47.7 +/- 0.8 kg/m(2) to 43.3 +/- 0.8 kg/m(2) (1 month), 35.2 +/- 0.8 kg/m(2) (6 months), and 30.2 +/- 1.2 kg/m(2) (24 months). Plasma dROMS also significantly deceased over time. At baseline, VAT (r = 0.46), FAT (r = 0.42), and BMI (r = 0.37) correlated with 6-month decreases in dROMS. Similarly, at 1 month, VAT (r = 0.43) and FAT (r = 0.41) correlated with 6-month decreases in dROMS. Multiple regression analysis showed that relationships between VAT and dROMS were significant after adjusting for FAT mass. Increased plasma dROMS at baseline were correlated with decreased concentrations of high-density lipoprotein (HDL) at 1 and 6 months after surgery (r = -0.38 and -0.42). This study found longitudinal associations between general, and more specifically intra-abdominal adiposity, and systemic lipid peroxides, suggesting that adipose tissue mass contributes to oxidative stress.

Topics: Adipose Tissue; Adult; Bariatric Surgery; Body Mass Index; Body Weight; Female; Humans; Intra-Abdominal Fat; Lipid Peroxides; Lipoproteins, HDL; Longitudinal Studies; Middle Aged; Obesity; Oxidative Stress; Predictive Value of Tests; Reactive Oxygen Species; Subcutaneous Fat; Tyrosine; Weight Loss

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

We sought to directly compare the effects of type 1 and type 2 diabetes on postischemic neovascularization and evaluate the mechanisms underlying differences between these groups. We tested the hypothesis that type 2 diabetic mice have a greater reduction in endothelial nitric oxide synthase (eNOS) expression, a greater increase in oxidative stress, and reduced arteriogenesis and angiogenesis, resulting in less complete blood flow recovery than type 1 diabetic mice after induction of hind limb ischemia.. Hind limb ischemia was generated by femoral artery excision in streptozotocin-treated mice (model of type 1 diabetes), in Lepr(db/db) mice (model of type 2 diabetes), and in control (C57BL/6) mice. Dependent variables included eNOS expression and markers of arteriogenesis, angiogenesis, and oxidative stress.. Postischemia recovery of hind limb perfusion was significantly less in type 2 than in type 1 diabetic mice; however, neither group demonstrated a significant increase in collateral artery diameter or collateral artery angioscore in the ischemic hind limb. The capillary/myofiber ratio in the gastrocnemius muscle decreased in response to ischemia in control or type 1 diabetic mice but remained the same in type 2 diabetic mice. Gastrocnemius muscle eNOS expression was lower in type 1 and 2 diabetic mice than in control mice. This expression decreased after induction of ischemia in type 2 but not in type 1 diabetic mice. The percentage of endothelial progenitor cells (EPC) in the peripheral blood failed to increase in either diabetic group after induction of ischemia, whereas this variable significantly increased in the control group in response to ischemia. EPC eNOS expression decreased after induction of ischemia in type 1 but not in type 2 diabetic mice. EPC nitrotyrosine accumulation increased after induction of ischemia in type 2 but not in type 1 diabetic mice. EPC migration in response to vascular endothelial growth factor was reduced in type 1 and type 2 diabetic mice vs control mice. EPC incorporation into tubular structures was less effective in type 2 diabetic mice. Extensive fatty infiltration was present in ischemic muscle of type 2 but not in type 1 diabetic mice.. Type 2 diabetic mice displayed a significantly less effective response to hind limb ischemia than type 1 diabetic mice.

Topics: Animals; Blood Glucose; Body Weight; Chemotaxis; Collateral Circulation; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Diabetes Mellitus, Type 2; Endothelial Cells; Hindlimb; Ischemia; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Muscle, Skeletal; Neovascularization, Physiologic; Nitric Oxide Synthase Type III; Oxidative Stress; Receptors, Leptin; Recovery of Function; Regional Blood Flow; Stem Cells; Time Factors; Tyrosine; Vascular Endothelial Growth Factor A

Metabolic syndrome (MetS) is a group of cardiovascular risk factors, including visceral obesity, glucose intolerance, hypertension, and dyslipidemia. Increased oxidative and nitrative stress and inflammation and decreased endothelial function occur in an animal model of metabolic syndrome, SHR/NDmcr-cp (SHR/cp) rats. The present study investigated the effects of coenzyme Q10 (CoQ10), one of the important antioxidants, on the abnormal oxidative condition and characteristic components of metabolic syndrome in SHR/cp rats by maintaining them on a diet supplemented with 0.07% - 0.7% CoQ10 for 26 weeks. We determined serum levels of oxidatively modified low-density lipoprotein (Ox-LDL) and 8-hydroxy-2'-deoxyguanosine (8-OHdG) as oxidative stress markers, 3-nitrotyrosine as a nitrative stress marker, 3-chlorotyrosine as a marker of myeloperoxidase (MPO)-catalyzed oxidation and high-sensitivity C-reactive protein (hsCRP) as an inflammatory marker. The administration of CoQ10 significantly attenuated the increase of oxidative and nitrative stress markers and inflammatory markers in a dose-dependent manner. CoQ10 prevented the elevated serum insulin levels, although it did not affect the elevated glucose level and dyslipidemia. CoQ10 also reduced elevated blood pressure, but did not affect body weight gain. In addition, CoQ10 improved endothelial dysfunction in the mesenteric arteries. These findings suggest that the antioxidant properties of CoQ10 can be effective for ameliorating cardiovascular risk in MetS.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Blood Glucose; Body Weight; Deoxyguanosine; Disease Models, Animal; Inflammation; Insulin; Lipids; Lipoproteins, LDL; Metabolic Syndrome; Oxidative Stress; Peroxidase; Rats; Rats, Inbred SHR; Tyrosine; Ubiquinone

This study was designed to examine the effects of a high-fat, high-sucrose (HFHS) diet on vascular and metabolic actions of insulin. Male rats were randomized to receive an HFHS or regular chow diet for 4 wk. In a first series of experiments, the rats had pulsed Doppler flow probes and intravascular catheters implanted to measure blood pressure, heart rate, and regional blood flows. Insulin sensitivity and vascular responses to insulin were assessed during a euglycemic hyperinsulinemic clamp performed in conscious rats. In a second series of experiments, new groups of rats were used to examine skeletal muscle glucose transport activity and to determine in vitro vascular reactivity, endothelial nitric oxide synthase (eNOS) protein expression in muscle and vascular tissues and endothelin content, nitrotyrosine formation, and NAD(P)H oxidase protein expression in vascular tissues. The HFHS-fed rats displayed insulin resistance, hyperinsulinemia, hypertriglyceridemia, hyperlipidemia, elevated blood pressure, and impaired insulin-mediated renal and skeletal muscle vasodilator responses. A reduction in endothelium-dependent vasorelaxation, accompanied by a decreased eNOS protein expression in muscles and blood vessel endothelium, and increased vascular endothelin-1 protein content were also noted in HFHS-fed rats compared with control rats. Furthermore, the HFHS diet induced a reduced insulin-stimulated glucose transport activity in muscles and increased levels of NAD(P)H oxidase protein and nitrotyrosine formation in vascular tissues. These findings support the importance of eNOS protein in linking metabolic and vascular disease and indicate the ability of a Westernized diet to induce endothelial dysfunction and to alter metabolic and vascular homeostasis.

Topics: Animals; Blood Pressure; Blotting, Western; Body Weight; Deoxyglucose; Diet; Dietary Fats; Endothelin-1; Endothelium, Vascular; Fatty Acids, Nonesterified; Fluorescent Antibody Technique; Glucose Clamp Technique; Heart Rate; Insulin; Insulin Resistance; Male; Obesity; Organ Size; Rats; Rats, Sprague-Dawley; Sucrose; Triglycerides; Tyrosine; Vascular Diseases; Vascular Resistance

Endothelial dysfunction is a characteristic feature during the renal damage induced by mild hyperuricemia. The mechanism by which uric acid reduces the bioavailability of intrarenal nitric oxide is not known. We tested the hypothesis that oxidative stress might contribute to the endothelial dysfunction and glomerular hemodynamic changes that occur with hyperuricemia. Hyperuricemia was induced in Sprague-Dawley rats by administration of the uricase inhibitor, oxonic acid (750 mg/kg per day). The superoxide scavenger, tempol (15 mg/kg per day), or placebo was administered simultaneously with the oxonic acid. All groups were evaluated throughout a 5-wk period. Kidneys were fixed by perfusion and afferent arteriole morphology, and tubulointerstitial 3-nitrotyrosine, 4-hydroxynonenal, NOX-4 subunit of renal NADPH-oxidase, and angiotensin II were quantified. Hyperuricemia induced intrarenal oxidative stress, increased expression of NOX-4 and angiotensin II, and decreased nitric oxide bioavailability, systemic hypertension, renal vasoconstriction, and afferent arteriolopathy. Tempol treatment reversed the systemic and renal alterations induced by hyperuricemia despite equivalent hyperuricemia. Moreover, because tempol prevented the development of preglomerular damage and decreased blood pressure, glomerular pressure was maintained at normal values as well. Mild hyperuricemia induced by uricase inhibition causes intrarenal oxidative stress, which contributes to the development of the systemic hypertension and the renal abnormalities induced by increased uric acid. Scavenging of the superoxide anion in this setting attenuates the adverse effects induced by hyperuricemia.

Topics: Aldehydes; Angiotensin II; Animals; Antioxidants; Arterioles; Body Weight; Cyclic N-Oxides; Disease Models, Animal; Glomerular Filtration Rate; Hypertension, Renal; Hyperuricemia; Kidney Glomerulus; Male; NADPH Oxidase 4; NADPH Oxidases; Oxidative Stress; Oxonic Acid; Rats; Rats, Sprague-Dawley; Renal Circulation; Spin Labels; Superoxides; Tyrosine

Obstructive sleep apnea is characterized by upper airway collapse, leading to intermittent hypoxia (IH). It has been postulated that IH-induced oxidative stress may contribute to several chronic diseases associated with obstructive sleep apnea. We hypothesize that IH induces systemic oxidative stress by upregulating NADPH oxidase, a superoxide-generating enzyme. NADPH oxidase is regulated by a cytosolic p47(phox) subunit, which becomes phosphorylated during enzyme activation. Male C57BL/6J mice were exposed to IH with an inspired O(2) fraction nadir of 5% 60 times/h during the 12-h light phase (9 AM-9 PM) for 1 or 4 wk. In the aorta and heart, IH did not affect lipid peroxidation [malondialdehyde (MDA) level], nitrotyrosine level, or p47(phox) expression and phosphorylation. In contrast, in the liver, exposure to IH for 1 wk resulted in a trend to an increase in MDA levels, whereas IH for 4 wk resulted in a 38% increase in MDA levels accompanied by upregulation of p47(phox) expression and phosphorylation. Administration of an NADPH oxidase inhibitor, apocynin, during IH exposure attenuated IH-induced increases in hepatic MDA. In p47(phox)-deficient mice, MDA levels were higher at baseline and, unexpectedly, decreased during IH. In conclusion, oxidative stress levels and pathways under IH conditions are organ and duration specific.

Topics: Acetophenones; Animals; Body Weight; Eating; Enzyme Inhibitors; Erythrocytes; Glutathione; Glutathione Disulfide; Hypoxia; Lipid Peroxidation; Lipoproteins, LDL; Liver; Male; Malondialdehyde; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocardium; NADPH Oxidases; Oxidative Stress; Phosphorylation; Superoxide Dismutase; Tyrosine

Oxidative stress has been postulated to be involved in the development of diabetic nephropathy. In the present study, we evaluated the effect of taurine, an endogenous antioxidant, on diabetic nephropathy by mixing it with the daily drinking water (1%w/v) of streptozotocin-induced diabetic rats from the beginning of the fourth month after the induction of diabetes, during which the urinary protein excretion in untreated diabetic rats showed significant increase in comparison with nondiabetic rats. The taurine administration significantly suppressed further increase in urinary protein excretion in diabetic rats, accompanied by the reduction of mesangial extracellular matrix expansion and TGF-beta expression in the renal glomerulus. Immunohistochemical study showed that taurine administration suppressed the intensified stainings to the three different types of oxidative stress markers, such as 8-hydroxyl-2'-deoxyguanosine (8-OHdG), pentosidine, and nitrotyrosine observed in the renal tissues of untreated diabetic rats. These findings suggest that taurine has the ability to suppress the progression of diabetic nephropathy at least in part by its antioxidant property. Since this beneficial effect of taurine was obtained even if its administration was started after the time point when urinary protein excretion already became apparently higher than that of age-matched nondiabetic animals, taurine administration was potentially expected to be applied in clinical field to retard the development of nephropathy in diagnosed diabetic patients.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Biomarkers; Blood Glucose; Body Weight; Deoxyguanosine; Diabetic Nephropathies; Disease Progression; Glycated Hemoglobin; Immunohistochemistry; Male; Oxidative Stress; Proteinuria; Rats; Rats, Sprague-Dawley; Taurine; Tyrosine

To investigate whether the micronutrients that were shown to reduce the risk of development of age-related macular degeneration in the Age-Related Eye Disease Study (AREDS) can have the same effect on the development of diabetic retinopathy in rats, and to understand the possible mechanisms.. Streptozotocin-induced diabetic rats received a powdered diet with or without supplemental micronutrients (ascorbic acid, vitamin E, beta-carotene, zinc, and copper). The retina was used after the rats had diabetes for 12 months to detect vascular histopathology and to measure the biochemical parameters and messenger RNA levels of the genes involved in oxidative and nitrative stress.. The AREDS-based micronutrients prevented a diabetes-induced increase in the number of retinal acellular capillaries. In the same rats, micronutrients inhibited increases in retinal oxidatively modified DNA and nitrotyrosine and decreases in manganese superoxide dismutase. Diabetes-induced alterations in the messenger RNA expression of mitochondrial electron transport complex III (coenzyme Q cytochrome-c reductase) and inducible nitric oxide synthase were also prevented.. Age-Related Eye Disease Study-based micronutrients inhibit the development of diabetic retinopathy in rodents by inhibiting oxidative and nitrative stress.. Micronutrients that slow down the onset and progression of age-related macular degeneration have the potential to inhibit the development of diabetic retinopathy.

Topics: Animals; Ascorbic Acid; beta Carotene; Body Weight; Copper; Deoxyadenosines; Diabetes Mellitus, Experimental; Diabetic Retinopathy; Diet; Eating; Electron Transport Complex III; Glycated Hemoglobin; Immunoenzyme Techniques; Male; Micronutrients; Nitric Oxide Synthase Type II; Oxidative Stress; Polymerase Chain Reaction; Rats; Rats, Inbred Lew; Retinal Vessels; RNA, Messenger; Superoxide Dismutase; Tyrosine; Vitamin E; Zinc Oxide

Coffee is a rich source of antioxidative polyphenols, but epidemiological studies and interventional trials have failed to demonstrate any clear beneficial effects of coffee consumption on hypertension. The interaction between hydroxyhydroquinone (HHQ) and 5-caffeoylquinic acid (CQA) was examined, in an attempt to understand the controversial effects of coffee on hypertension.. Male Wistar Kyoto (WKY) rats or spontaneously hypertensive rats (SHRs, 14 weeks old) were divided into the following four groups; those on a control diet, 0.005% HHQ diet, 0.5% CQA diet, and HHQ plus CQA diet. The rats were fed the above diets for 8 weeks, and the tail arterial blood pressure was monitored in conscious rats at 2-week intervals. Urinary nitric oxide (NO) metabolites and hydrogen peroxide (H(2)O(2)) excretion were measured 8 weeks after the start of the experiment. Endothelium-dependent and -independent vasorelaxant responses and immunohistochemical staining for nitrotyrosine were examined in aortas.. HHQ inhibited the CQA-induced improvement in hypertension, urinary NO metabolites or H(2)O(2) excretion, endothelial dysfunction, and nitrotyrosine deposits in aortas in SHR. However, the administration of HHQ alone had little effect on either strain.. Based on the content ratio of HHQ and chlorogenic acids in coffee, HHQ interfered with the CQA-induced improvement in blood pressure and endothelial function in SHR. The results explain, at least in part, the conflicting action of coffee drinking on hypertension and vascular reactivity.

Topics: Acetylcholine; Animals; Antihypertensive Agents; Blood Pressure; Body Weight; Chlorogenic Acid; Disease Models, Animal; Endothelium, Vascular; Heart Rate; Hydrogen Peroxide; Hydroquinones; Hypertension; Immunohistochemistry; Male; Nitric Oxide; Nitroprusside; Quinic Acid; Rats; Rats, Inbred SHR; Rats, Inbred WKY; Time Factors; Tyrosine; Vasodilation; Vasodilator Agents

In liver cirrhosis, elevated levels of NO and ROS (reactive oxygen species) might greatly favour the generation of peroxynitrite. Peroxynitrite is a highly reactive oxidant and it can potentially alter the vascular reactivity and the function of different organs. In the present study, we evaluated whether peroxynitrite levels are related to the progression of renal vascular and excretory dysfunction during experimental cirrhosis induced by chronic BDL (bile-duct ligation) in rats. Experiments were performed at 7, 15 and 21 days after BDL in rats and in rats 21 days post-BDL chronically treated with L-NAME (N(G)-nitro-L-arginine methyl ester). Sodium balance, BP (blood pressure), basal RPP (renal perfusion pressure) and the renal vascular response to PHE (phenylephrine) and ACh (acetylcholine) in isolated perfused kidneys were measured. NO levels were calculated as 24-h urinary excretion of nitrites, ROS as TBARS (thiobarbituric acid-reacting substances), and peroxynitrite formation as the renal expression of nitrotyrosine. BDL rats had progressive sodium retention, and decreased BP, RPP and renal vascular responses to PHE and ACh in the time following BDL. They also had increasing levels of NO and ROS, and renal nitrotyrosine accumulation,especially in the medulla. All of these changes were either prevented or significantly decreased by chronic L-NAME administration. In conclusion, these results suggest that the increasing levels of peroxynitrite might contribute to the altered renal vascular response and sodium retention in the development of the experimental biliary cirrhosis. Moreover, the beneficial effects of decreasing NO synthesis are, at least in part, mediated by anti-peroxinitrite-related effects.

Topics: Animals; Body Weight; Chronic Disease; Kidney; Liver Cirrhosis, Experimental; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Organ Size; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Renal Circulation; Spleen; Tyrosine

Methotrexate (MTX), a structural analogue of folic acid, is widely used as a chemotherapeutic agent for leukemia and other malignancies. One of the major toxic effects of MTX is intestinal injury and enterocolitis .The mechanism of gastrointestinal toxicity of methotrexate has not been investigated completely. Therefore cancer chemotherapy has to be accompanied by symptomatic therapy such as antibiotics and anti-diarrheal drugs. It is important to investigate the mechanism by which methotrexate induces intestinal damage in order to perform cancer chemotherapy effectively by preventing the side effects. This study aimed at investigating whether nitrosative stress plays a role in methotrexate induced small intestinal damage using a rat model. Adult male rats were administered methotrexate at the dose of 7 mg/kg body weight intraperitoneally for 3 consecutive days and sacrificed 12 or 24 h after the final dose of methotrexate. Vehicle treated rats served as control. The intestinal tissue was used for light microscopic studies and markers of nitrosative stress including tissue nitrite level and nitrotyrosine. Myeloperoxidase (MPO) activity, a marker of neutrophil infiltration was also measured in intestinal homogenates. The villi were damaged at 12 h and the damage progressed and became severe at 24 h after the final dose of MTX. Biochemically, tissue nitrate was elevated fivefold at 12 h and fourfold at 24 h after the final dose of MTX as compared with control. Nitrotyrosine, measured immunohistochemically was detected in all the parts of the small intestine. Duodenum stained the most for nitrotyrosine, followed by ileum and then jejunum. The staining for nitrotyrosine was more intense at 24 h as compared with 12 h after the final dose of methotrexate. There was marked neutrophil infiltration as evidenced by increase in MPO activity in the small intestines. In conclusion, the results of the present study reveal that nitrosative stress may play a critical role in methotrexate induced small intestinal damage. Intervention studies using nitric oxide synthase inhibitors is being carried out in order to confirm the role of nitrosative stress in methotrexate induced small intestinal damage.

Topics: Animals; Antimetabolites, Antineoplastic; Body Weight; Duodenum; Ileum; Immunohistochemistry; Injections, Intraperitoneal; Intestine, Small; Jejunum; Male; Methotrexate; Neutrophil Infiltration; Neutrophils; Nitrites; Oxidative Stress; Peroxidase; Rats; Rats, Wistar; Reactive Nitrogen Species; Time Factors; Tyrosine

We investigated the effect of rosiglitazone (RSG), a high-affinity ligand for the peroxisome proliferator-activated receptor gamma which mediates insulin-sensitizing actions, on the lipid profile and oxidative status in streptozotocin (STZ)-induced Type 2 diabetes mellitus (DM) rats. Wistar albino male rats were randomly divided into an untreated control group (C), a C + RSG group which was treated with RSG (4 mg kg(-1)) two times a day by gavage, a diabetic group (D) that was treated with a single intraperitoneal injection of STZ (45 mgkg(-1)), D + RSG group which were treated with RSG two times a day by gavage, respectively. Lipid profiles, HbA(1c) and blood glucose levels in the circulation and malondialdehyde (MDA) and 3-nitrotyrosine (3-NT) levels in left ventricular muscle were measured. Treatment of D rats with RSG resulted in a time-dependent decrease in blood glucose. We found that the lipid profile and HbA(1c) levels in D + RSG group reached the C rat values at the end of the treatment period. There was a statistically significant difference between the C + RSG and C groups in 3-NT levels. In group D, 3-NT and MDA levels were found to be increased when compared with C, C + RSG and D + RSG groups. In the D + RSG group, MDA levels were found to be decreased when compared with C and C + RSG. Our study suggests that the treatment of D rats with RSG for 8 weeks may decrease the oxidative/nitrosative stress in left ventricular tissue of rats. Thus in diabetes-related vascular diseases, RSG treatment may be cardioprotective.

Topics: Animals; Blood Glucose; Body Weight; Diabetes Mellitus, Experimental; Glycated Hemoglobin; Heart Ventricles; Lipid Metabolism; Male; Malondialdehyde; Oxidative Stress; Rats; Rats, Wistar; Rosiglitazone; Streptozocin; Thiazolidinediones; Tyrosine

Uncoupling protein 2 (UCP2) is an important regulator of intracellular reactive oxygen species (ROS) production. We determined the effects of calorie restriction (CR) on the dynamic aspects of mitochondrial ROS production, UCP2, and the nitric oxide (NO)-cGMP pathway in the cardiovascular tissues of type II diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Some rats were on restricted diets (30% reduction from free intake) from age 29 to 42 weeks. Blood glucose, hemoglobin A1c, plasma levels of free fatty acid, triacylglycerol, and plasminogen activator inhibitor-1 in OLETF rats were significantly higher than those in nondiabetic control [Long-Evans Tokushima Otsuka (LETO)] rats at 29 weeks. Mitochondrial ROS production and UCP2 expression significantly increased in the heart and aorta of OLETF rats compared with those in LETO rats. A fibrogenic growth factor, transforming growth factor (TGF)-beta1 in the coronary vessels, endothelial nitric-oxide synthase, and aortic nitrotyrosine were increased in OLETF rats at 42 weeks. In contrast, an index of the NO-cGMP pathway, phosphorylated vasodilator-stimulated phosphoprotein, and superoxide dismutase activity in the aorta were significantly diminished. The relationship between UCP2 and ROS production in the cardiovascular function of diabetic rats being fed a calorie-restricted diet is unknown. These abnormalities in OLETF rats were reversed to normal levels by CR. CR significantly improved the NO-cGMP pathway via normalizing ROS generation in OLETF rats. A decrease in UCP2 expression by CR may be a compensatory mechanism to counteract decreased intracellular oxidative stress. The data suggest that CR may prevent cardiovascular tissues from oxidative stress provoked by diabetes mellitus.

Topics: Animals; Body Weight; Caloric Restriction; Cardiovascular Diseases; Diabetes Mellitus, Type 2; Glucose Transporter Type 4; Ion Channels; Lipid Peroxidation; Male; Mitochondria; Mitochondrial Proteins; Nitric Oxide Synthase Type III; Organ Size; Plasminogen Activator Inhibitor 1; Rats; Rats, Long-Evans; Reactive Oxygen Species; Risk Factors; Tyrosine; Uncoupling Protein 2

Whereas an important role of free radicals and oxidants in peripheral diabetic neuropathy is well established, the contribution of nitrosative stress and, in particular, of the highly reactive oxidant peroxynitrite, has not been properly explored. Our previous findings implicate peroxynitrite in diabetes-associated motor and sensory nerve conduction deficits and peripheral nerve energy deficiency and poly(ADP-ribose) polymerase activation associated with Type 1 diabetes. In this study the role of nitrosative stress in diabetic sensory neuropathy is evaluated. The peroxynitrite decomposition catalyst Fe(III) tetrakis-2-(N-triethylene glycol monomethyl ether)pyridyl porphyrin (FP15) was administered to control and streptozotocin (STZ)-diabetic mice at the dose of 5 mg kg(-1) day(-1) (FP15), for 3 weeks after initial 3 weeks without treatment. Mice with 6-week duration of diabetes developed clearly manifest thermal hypoalgesia (paw withdrawal, tail-flick, and hot plate tests), mechanical hypoalgesia (tail pressure Randall-Sellito test), tactile allodynia (flexible von Frey filament test), and approximately 38% loss of intraepidermal nerve fibers. They also had increased nitrotyrosine and poly(ADP-ribose) immunofluorescence in the sciatic nerve, grey matter of spinal cord, and dorsal root ganglion neurons. FP15 treatment was associated with alleviation of thermal and mechanical hypoalgesia. Tactile response threshold tended to increase in response to peroxynitrite decomposition catalyst treatment, but still remained approximately 59% lower compared with non-diabetic controls. Intraepidermal nerve fiber density was 25% higher in FP15-treated than in untreated diabetic rats, but the difference between two groups did not achieve statistical significance (p=0.054). Nitrotyrosine and poly(ADP-ribose) immunofluorescence in sciatic nerve, spinal cord, and dorsal root ganglion neurons of peroxynitrite decomposition catalyst-treated diabetic mice were markedly reduced. In conclusion, nitrosative stress plays an important role in sensory neuropathy associated with Type 1 diabetes. The findings provide rationale for further studies of peroxynitrite decomposition catalysts in a long-term diabetic model.

Topics: Animals; Behavior, Animal; Body Weight; Diabetes Mellitus, Experimental; Diabetic Neuropathies; Fluorescent Antibody Technique; Hot Temperature; Hyperalgesia; Immunohistochemistry; Male; Metalloporphyrins; Mice; Mice, Inbred C57BL; Nerve Fibers; Neurons; Pain Threshold; Peroxynitrous Acid; Poly Adenosine Diphosphate Ribose; Reaction Time; Sciatic Nerve; Spinal Nerve Roots; Streptozocin; Tyrosine

Insulin resistance is present in almost all patients with nonalcoholic steatohepatitis (NAFLD), and mitochondrial dysfunction likely plays a critical role in the progression of fatty liver into nonalcoholic steatohepatitis. Rosiglitazone, a selective ligand of peroxisome proliferator-activated receptor gamma (PPARgamma), is an insulin sensitizer drug that has been used in a number of insulin-resistant conditions, including NAFLD. The aim of this study was to analyze the effects of rosiglitazone on the liver histology and mitochondrial function in a model of NAFLD. All studies were carried out in wild-type and leptin-deficient (ob/ob) C57BL/6J mice. Ob/ob mice were treated with 1 mg/kg/day, and activity of mitochondrial respiratory chain (MRC), beta-oxidation, lipid peroxidation, glutathione content in mitochondria, and 3-tyrosine-nitrated proteins in mitochondria were measured. In addition, histological and ultrastructural changes induced by rosiglitazone were also noted. Rosiglitazone treatment increased liver steatosis, particularly microvesicular steatosis. In these animals, mitochondria were markedly swollen with cristae peripherally placed. In ob/ob mice, this drug increased PPARgamma protein expression and lipid peroxide content in liver tissue and decreased glutathione concentration in mitochondria. Rosiglitazone suppressed the activity of complex I of the MRC in ob/ob mice, but did not affect beta-oxidation. 3-Tyrosine nitrated mitochondrial proteins, significantly increased in ob/ob mice, were not modified by rosiglitazone treatment.. Treatment of ob/ob mice with rosiglitazone did not reverse histological lesions of NAFLD or improve MRC activity. On the contrary, rosiglitazone reduced activity of complex I and increased oxidative stress and liver steatosis.

Topics: Alanine Transaminase; Animals; Aspartate Aminotransferases; Body Weight; Fatty Liver; Lipid Peroxidation; Liver; Male; Mice; Mice, Inbred C57BL; Mice, Obese; Microscopy, Electron; Mitochondria, Liver; PPAR gamma; Rosiglitazone; Thiazolidinediones; Tyrosine

3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors are known to modulate carcinogenesis. In this study, we investigated whether a lipophilic HMG-CoA reductase inhibitor pitavastatin suppresses inflammation-related mouse colon carcinogenesis. Male CD-1 (ICR) mice were initiated with a single intraperitoneal injection of azoxymethane (AOM, 10 mg/kg body weight) and promoted by 2% (w/v) dextran sodium sulfate (DSS) in drinking water for 7 days. The experimental diets containing pitavastatin at 2 dose levels (1 and 10 ppm) were fed to male CD-1 (ICR) mice for 17 weeks, staring 1 week after the cessation of DSS exposure. The effects of dietary pitavastatin on colonic tumor development were assessed at Weeks 5, 10 and 20. Feeding with pitavastatin at both doses significantly inhibited the multiplicity of colonic adenocarcinoma at Week 20. Furthermore, the treatment significantly lowered the positive rates of proliferating cell nuclear antigen and increased the apoptotic index in the colonic epithelial malignancies. The treatment also reduced nitrotyrosine-positivity in the colonic mucosa. Our findings thus show that pitavastatin is effective in inhibiting colitis-related colon carcinogenesis through modulation of mucosal inflammation, oxidative/nitrosative stress, and cell proliferation.

Topics: Animals; Body Weight; Cholesterol; Colonic Neoplasms; DNA, Single-Stranded; Immunohistochemistry; Inflammation; Male; Mice; Molecular Structure; Organ Size; Proliferating Cell Nuclear Antigen; Quinolines; Time Factors; Triglycerides; Tyrosine

Accelerated formation of advanced glycation/lipoxidation and endproducts (AGEs/ALEs) has been implicated in the pathogenesis of various diabetic complications. Several natural and synthetic compounds have been proposed and tested as inhibitors of AGE/ALE formation. We have previously reported the therapeutic effects of several new AGE/ALE inhibitors on the prevention of nephropathy and dyslipidemia in streptozotocin (STZ)-induced diabetic rats. In this study, we investigated the effects of various concentrations of a compound, LR-90, on the progression of renal disease and its effects on AGE and receptor for AGE (RAGE) protein expression on the kidneys of diabetic STZ-rats. Diabetic male Sprague-Dawley rats were treated with or without LR-90 (0, 5, 20, 25, and 50 mg/l of drinking water). After 32 weeks, body weight, glycemic status, renal function, and plasma lipids were measured. Kidney histopathology and AGE/ALE accumulation and RAGE protein expression in tissues were also determined. In vitro studies were also performed to determine the possible mechanism of action of LR-90 in inhibiting AGE formation and AGE-protein cross-linking. LR-90 protected the diabetic kidneys by inhibiting the increase in urinary albumin-to-creatinine ratio and ameliorated hyperlipidemia in diabetic rats in a concentration-dependent fashion without any effects on hyperglycemia. LR-90 treatment also reduced kidney AGE/ALE accumulation and RAGE protein expression in a concentration-dependent manner. In vitro, LR-90 exhibited general antioxidant properties by inhibiting metal-catalyzed reactions and reactive oxygen species (OH radical) and reactive carbonyl species (methlyglyoxal, glyoxal) generations without any effect on pyridoxal 5' phosphate. The compound also prevents AGE-protein cross-linking reactions. These findings demonstrate the bioefficacy of LR-90 in treating nephropathy and hyperlipidemia in diabetic animals by inhibiting AGE accumulation, RAGE protein expression, and protein oxidation in the diabetic kidney. Additionally, our study suggests that LR-90 may be useful also to delay the onset and progression of diabetic atherosclerosis as the compound can inhibit the expression of RAGE and inflammation-related pathology, as well as prevent lipid peroxidation reactions.

Topics: Animals; Ascorbic Acid; Body Weight; Butyrates; Cholesterol; Creatinine; Diabetes Mellitus, Experimental; Diabetic Nephropathies; Dyslipidemias; Glycation End Products, Advanced; Glycosylation; Hydroxyl Radical; Hypoglycemic Agents; Kidney; Lipid Metabolism; Lipoproteins, LDL; Male; Molecular Structure; Oxidation-Reduction; Pyridoxal Phosphate; Rats; Rats, Sprague-Dawley; Receptor for Advanced Glycation End Products; Receptors, Immunologic; Triglycerides; Tyrosine

Glycation, oxidation, and nitration of endogenous proteins occur spontaneously and these modifications are also present in foods. Increased levels of these chemical changes are associated with chronic renal failure; however, little is known about acute kidney failure. We measured these modifications of plasma protein and related free adducts in plasma following bilateral nephrectomy and bilateral ureteral obstruction. Advanced glycation end-product (AGE) residues of plasma protein were increased 3 h post-surgery, and thereafter slowly decreased in all groups, reflecting changes in plasma protein synthesis and transcapillary flow post-surgery. Ureteral ligation increased oxidation and nitration adduct residues. There were, however, marked increases in AGE, dityrosine, or 3-nitrotyrosine free adducts in both nephrectomized and ureter-ligated rats compared to rats that had undergone sham operations. There were lower modified adduct concentrations in the ureter-ligated compared to the nephrectomized rats, reflecting residual glomerular filtration and tubular removal. There was no increase in glycated, oxidized, and nitrated proteins. Glyoxal and methylglyoxal were also increased in both renal failure models. Our study shows that the acute loss of renal function and urinary excretion leads to the accumulation of AGE, oxidation, and nitration free adducts in the plasma.

Topics: Animals; Body Weight; Disease Models, Animal; Glycation End Products, Advanced; Glyoxal; Kidney Failure, Chronic; Ligation; Male; Nephrectomy; Oxidative Stress; Pyruvaldehyde; Rats; Rats, Wistar; Tyrosine

Evidence for important roles of the highly reactive oxidant peroxynitrite in diabetic complications is emerging. We evaluated the role of peroxynitrite in early peripheral neuropathy and vascular dysfunction in STZ-diabetic rats. In the first dose-finding study, control and STZ-diabetic rats were maintained with or without the potent peroxynitrite decomposition catalyst Fe(III)tetrakis-2-(N-triethylene glycol monomethyl ether) pyridyl porphyrin (FP15) at 3, 5, or 10 mg.kg(-1).day(-1) in the drinking water for 4 wk after an initial 2 wk without treatment for assessment of early neuropathy. In the second study with similar experimental design, control and STZ-diabetic rats were maintained with or without FP15, 5 mg.kg(-1).day(-1), for vascular studies. Rats with 6-wk duration of diabetes developed motor and sensory nerve conduction velocity deficits, mechanical hyperalgesia, and tactile allodynia in the absence of small sensory nerve fiber degeneration. They also had increased nitrotyrosine and poly(ADP-ribose) immunofluorescence in the sciatic nerve and dorsal root ganglia. All these variables were dose-dependently corrected by FP15, with minimal differences between the 5 and 10 mg.kg(-1).day(-1) doses. FP15, 5 mg.kg(-1).day(-1), also corrected endoneurial nutritive blood flow and nitrotyrosine, but not superoxide, fluorescence in aorta and epineurial arterioles. Diabetes-induced decreases in acetylcholine-mediated relaxation by epineurial arterioles and coronary and mesenteric arteries, as well as bradykinin-induced relaxation by coronary and mesenteric arteries, were alleviated by FP15 treatment. The findings reveal the important role of nitrosative stress in early neuropathy and vasculopathy and provide the rationale for further studies of peroxynitrite decomposition catalysts in long-term diabetic models.

Topics: Animals; Aorta; Arterioles; Blood Glucose; Body Weight; Coronary Vessels; Diabetes Mellitus, Experimental; Diabetic Angiopathies; Diabetic Neuropathies; Ganglia, Spinal; Hyperalgesia; Hypoglycemic Agents; Male; Mesenteric Arteries; Metalloporphyrins; Neural Conduction; Peroxynitrous Acid; Poly Adenosine Diphosphate Ribose; Rats; Rats, Wistar; Regional Blood Flow; Sciatic Nerve; Superoxides; Tyrosine; Vasodilator Agents

We have previously established a nonobese diabetes mouse model characterized by moderate hyperglycemic levels, like those usually occurring in human type 2 diabetes. As oxidative stress is considered a major mechanism of progressive beta-cell damage in diabetes, we tested in this model the protective effects of a new low-molecular-weight antioxidant, namely, bis(1-hydroxy-2,2,6,6-tetramethyl-4-piperidinyl)decandioate dihydrochloride (IAC).. Diabetes was induced in C57Bl/6J mice by streptozotocin (STZ) and nicotinamide (NA) administration. Two weeks later, STZ-NA mice were treated for 5 weeks with different doses of IAC (15 or 30 mg/kg per day intraperitoneally) and monitored for glycemia, insulinemia, glucose tolerance, and pancreatic insulin content.. Streptozotocin-NA mice showed moderate hyperglycemia, hypoinsulinemia, glucose intolerance, growth impairment, and markedly reduced pancreatic insulin content (22% of controls). IAC-treated STZ-NA mice showed clear-cut reduction of hyperglycemia and attenuation of glucose intolerance, associated to higher residual pancreatic insulin content with respect to untreated diabetic animals. Plasma nitrotyrosine levels (an index of oxidative stress), enhanced 3-fold in diabetic mice, were significantly reduced by IAC treatment. Significant correlations were found between plasma nitrotyrosine values and either blood glucose levels or pancreatic insulin content.. In the STZ-NA diabetic mouse model, the new antioxidant, IAC, improves diabetic metabolic alterations, likely by counteracting beta-cell dysfunction and loss associated with oxidative stress.

Topics: Animals; Antioxidants; Blood Glucose; Body Weight; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 2; Dose-Response Relationship, Drug; Esters; Fatty Acids, Nonesterified; Glucose Intolerance; Glucose Tolerance Test; Hyperglycemia; Hypoglycemic Agents; Insulin; Male; Mice; Mice, Inbred C57BL; Molecular Weight; Niacinamide; Oxidative Stress; Pancreas; Piperidines; Streptozocin; Time Factors; Tyrosine

Both increased aldose reductase (AR) activity and oxidative/nitrosative stress have been implicated in the pathogenesis of diabetic nephropathy, but the relation between the two factors remains a subject of debate. This study evaluated the effects of AR inhibition on nitrosative stress and poly(ADP-ribose) polymerase (PARP) activation in diabetic rat kidney and high-glucose-exposed human mesangial cells. In animal experiments, control (C) and streptozotocin-diabetic (D) rats were treated with/without the AR inhibitor fidarestat (F, 16 mg kg(-1) day(-1)) for 6 weeks starting from induction of diabetes. Glucose, sorbitol, and fructose concentrations were significantly increased in the renal cortex of D vs C (p < 0.01 for all three comparisons), and sorbitol pathway intermediate, but not glucose, accumulation, was completely prevented in D + F. F at least partially prevented diabetes-induced increase in kidney weight as well as nitrotyrosine (NT, a marker of peroxynitrite-induced injury and nitrosative stress), and poly(ADP-ribose) (a marker of PARP activation) accumulation, assessed by both immunohistochemistry and Western blot analysis, in glomerular and tubular compartments of the renal cortex. In vitro studies revealed the presence of both AR and PARP-1 in human mesangial cells, and none of these two variables were affected by high glucose or F treatment. Nitrosylated and poly(ADP-ribosyl)ated proteins (Western blot analysis) accumulated in cells cultured in 30 mM D-glucose (vs 5.55 mM glucose, p < 0.01), but not in cells cultured in 30 mM L-glucose or 30 mM D-glucose plus 10 microM F. AR inhibition counteracts nitrosative stress and PARP activation in the diabetic renal cortex and high-glucose-exposed human mesangial cells. These findings reveal new beneficial properties of the AR inhibitor F and provide the rationale for detailed studies of F on diabetic nephropathy.

Topics: Aldehyde Reductase; Animals; Body Weight; Diabetes Mellitus, Experimental; Enzyme Activation; Glucose; Immunohistochemistry; Kidney; Male; Mesangial Cells; Nitrogen; Oxidation-Reduction; Poly(ADP-ribose) Polymerases; Rats; Rats, Wistar; Tyrosine

Poly(ADP-ribose) polymerase (PARP) activation is emerging as a fundamental mechanism in the pathogenesis of diabetes complications including diabetic neuropathy. This study evaluated the role of PARP in diabetic sensory neuropathy. The experiments were performed in control and streptozotocin-induced diabetic rats treated with or without the PARP inhibitor 1,5-isoquinolinediol (ISO; 3 mg x kg(-1) x day(-1) i.p.) for 2 weeks after 2 weeks without treatment. Diabetic rats developed thermal hyperalgesia (assessed by paw-withdrawal and tail-flick tests), mechanical hyperalgesia (von Frey anesthesiometer/rigid filaments and Randall-Sellito tests), tactile allodynia (flexible von Frey filaments), and increased flinching behavior in phases 1 and 2 of the 2% formalin pain test. They also had clearly manifest increase in nitrotyrosine and poly(ADP-ribose) immunoreactivities in the sciatic nerve and increased superoxide formation (hydroxyethidine method) and nitrotyrosine immunoreactivity in vasa nervorum. ISO treatment alleviated abnormal sensory responses, including thermal and mechanical hyperalgesia and tactile allodynia as well as exaggerated formalin flinching behavior in diabetic rats, without affecting the aforementioned variables in the control group. Poly(ADP-ribose) and, to a lesser extent, nitrotyrosine abundance in sciatic nerve, as well as superoxide and nitrotyrosine formation in vasa nervorum, were markedly reduced by ISO therapy. Apoptosis in dorsal root ganglion neurons (transferase-mediated dUTP nick-end labeling assay) was not detected in any of the groups. In conclusion, PARP activation contributes to early diabetic sensory neuropathy by mechanisms that may include oxidative stress but not neuronal apoptosis.

Topics: Animals; Apoptosis; Arthropathy, Neurogenic; Blood Glucose; Body Weight; Diabetes Mellitus, Experimental; Diabetic Neuropathies; Ganglia, Spinal; Hyperalgesia; Immunohistochemistry; Isoquinolines; Male; Neural Conduction; Neurons; Oxidative Stress; Poly(ADP-ribose) Polymerase Inhibitors; Quinolines; Rats; Rats, Wistar; Streptozocin; Tyrosine

Renal cyclooxygenase (COX)-2 expression and arachidonic acid-stimulated prostaglandin release are increased in streptozotocin-diabetic rats and are reduced by tempol treatment, indicating a role for superoxide. Generation of nitric oxide (NO) and its product with superoxide, peroxynitrite, is also increased in diabetes and can induce COX-2. To investigate a role of NO, rats were treated with L-nitroarginine methyl ester (L-NAME; 100 mg/kg/day) to inhibit NO synthase (NOS) for 14-18 days. In isolated perfused kidneys from diabetic rats, prostaglandin release and vasoconstrictor responses to arachidonic acid were increased and renal cortical expression of COX-2 was 2-fold that of control rats. Treatment of diabetic rats with L-NAME reduced arachidonic acid-stimulated release of prostaglandins and the expression of COX-2. L-NAME increased vasoconstrictor responses to AA in diabetic and non-diabetic rats but abolished the differences between the two. These results, coupled with those using tempol, suggest that NO or its product with superoxide may contribute to the induction of renal COX-2 in the diabetic rat.

Topics: Angiotensin II; Animals; Arachidonic Acid; Blood Pressure; Blotting, Western; Body Weight; Cyclooxygenase 1; Cyclooxygenase 2; Diabetes Mellitus, Experimental; Enzyme Inhibitors; In Vitro Techniques; Kidney; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Perfusion; Pressure; Prostaglandin-Endoperoxide Synthases; Prostaglandins; Rats; Rats, Wistar; Tyrosine

Although antioxidants are used to treat an overdose of the analgaesic/antipyretic drug APAP (acetaminophen), roles of antioxidant enzymes in APAP-induced hepatotoxicity remain controversial. Our objective was to determine impacts of knockout of SOD1 (superoxide dismutase; Cu,Zn-SOD) alone or in combination with selenium-dependent GPX1 (glutathione peroxidase-1) on APAP-induced hepatotoxicity. All SOD1-null (SOD1-/-) and SOD1- and GPX1-double-knockout mice survived an intraperitoneal injection of 600 mg of APAP per kg of body mass, whereas 75% of WT (wild-type) and GPX1-null mice died within 20 h. Survival time of SOD1-/- mice injected with 1200 mg of APAP per kg of body mass was longer than that of the WT mice (934 compared with 315 min, P<0.05). The APAP-treated SOD1-/- mice had less (P<0.05) plasma ALT (alanine aminotransferase) activity increase and attenuated (P<0.05) hepatic glutathione depletion than the WT mice. The protection conferred by SOD1 deletion was associated with a block of the APAP-mediated hepatic protein nitration and a 50% reduction (P<0.05) in activity of a key APAP metabolism enzyme CYP2E1 (cytochrome P450 2E1) in liver. The SOD1 deletion also caused moderate shifts in the APAP metabolism profiles. In conclusion, deletion of SOD1 alone or in combination with GPX1 greatly enhanced mouse resistance to APAP overdose. Our results suggest a possible pro-oxidant role for the physiological level of SOD1 activity in APAP-mediated hepatotoxicity.

Topics: Acetaminophen; Alanine Transaminase; Animals; Benzoquinones; Biotransformation; Body Weight; Cytochrome P-450 CYP2E1; Cytochrome P-450 CYP2E1 Inhibitors; Drug Resistance; Enzyme Induction; Glutathione; Glutathione Peroxidase; Glutathione Peroxidase GPX1; Imines; Liver Failure; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Oxidation-Reduction; Oxidative Stress; Peroxynitrous Acid; Superoxide Dismutase; Tyrosine

Corosolic acid (CRA), a constituent of banaba leaves, has been reported to have anti-inflammatory and hypoglycemic activities. The aim of this study was to determine the effects of CRA on metabolic risk factors including obesity, hypertension, hyperinsulinemia, hyperglycemia, and hyperlipidemia together with oxidative stress and inflammation, all of which are characteristic of the SHR/NDmcr-cp (cp/cp) (SHR-cp) rat, an animal model of metabolic syndrome. Six-week-old male SHR-cp rats were fed a high fat diet containing 0.072% CRA for 14 weeks. Treatment with CRA lowered blood pressure, which was elevated in control animals, by 10% after 8 weeks, and serum free fatty acids by 21% after 2 weeks. CRA treatment resulted in decreases in the levels of the oxidative stress markers thiobarbituric acid-reactive substances and 8-hydroxydeoxyguanosine by 27% and 59%, respectively, after 2 weeks. CRA treatment also reduced the levels of myeloperoxidase markers, 3-nitrotyrosine and 3-chlorotyrosine by 38% and 39%, respectively, after 10 weeks, and tended to decrease the levels of high sensitivity C-reactive protein, a marker of inflammation, after 6 weeks. However, CRA had no effect on weight gain or hyperglycemia. These results demonstrate that CRA can ameliorate hypertension, abnormal lipid metabolism, and oxidative stress as well as the inflammatory state in SHR-cp rats. This implies that CRA can be beneficial for preventing atherosclerosis-related diseases that are an increasing health care problem worldwide.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Animals; Blood Glucose; Blood Pressure; Body Weight; Cholesterol; Deoxyguanosine; Disease Models, Animal; Fatty Acids, Nonesterified; Hypertension; Inflammation; Insulin; Male; Metabolic Syndrome; Molecular Structure; Musa; Oxidative Stress; Phytotherapy; Plant Extracts; Rats; Rats, Inbred SHR; Thiobarbituric Acid Reactive Substances; Triglycerides; Triterpenes; Tyrosine

Oxidative stress is an important susceptibility factor for dilated cardiomyopathy. We have investigated the effects of bisoprolol, a beta1-selective adrenoceptor blocker, on oxidative stress and the development of cardiac dysfunction in a model of dilated cardiomyopathy. Male TO-2 and control hamsters at 8 weeks of age were treated with bisoprolol (5 mg/kg per day) or vehicle for 4 weeks. Treatment with bisoprolol prevented the progression of cardiac dysfunction in TO-2 hamsters. This drug did not affect the increase in NADPH oxidase activity but prevented the reduction in activity and expression of mitochondrial manganese-dependent superoxide dismutase as well as the increases in the concentrations of interleukin-1beta and tumor necrosis factor-alpha in the left ventricle of TO-2 hamsters. Attenuation of the development of cardiac dysfunction by bisoprolol may thus result in part from normalization of the associated increases in the levels of oxidative stress and pro-inflammatory cytokines in the left ventricle.

Topics: Aldehydes; Animals; Antioxidants; Bisoprolol; Blood Pressure; Body Weight; Cardiomyopathy, Dilated; Cricetinae; Disease Models, Animal; Echocardiography; Fibrosis; Glutathione; Heart Failure; Heart Rate; Interleukin-1; Isoenzymes; Male; NADPH Oxidases; Organ Size; Oxidative Stress; Superoxide Dismutase; Tumor Necrosis Factor-alpha; Tyrosine; Ventricular Function, Left

Chronic, inappropriate (relative to dietary Na+) elevations in circulating aldosterone, such as occur in congestive heart failure, are accompanied by a proinflammatory vascular phenotype involving the coronary and systemic vasculature. An immunostimulatory state with activated peripheral blood mononuclear cells (PBMCs) precedes this phenotype and is induced by a fall in cytosolic free [Mg2+]i and subsequent Ca2+ loading of these cells and transduced by oxidative/nitrosative stress.. We sought to further validate this hypothesis in rats with aldosterone/1%NaCl treatment (ALDOST) by using several interventions as cotreatment: a Mg2+-supplemented diet; amlodipine, a CCB; and N-acetylcysteine, an antioxidant. Blood samples were obtained at weeks 1 to 4 of ALDOST to monitor [Mg2+]i, [Ca2+]I, and H2O2 production in PBMCs. Coronal ventricular sections were examined for invading inflammatory cells and 3-nitrotyrosine labeling, a marker of oxidative/nitrosative stress. In response to ALDOST and compared with untreated controls, we found an early and persistent reduction in [Mg2+]i with a subsequent rise in [Ca2+]i and H2O2 production, each of which was either attenuated or abrogated by the Mg2+-supplemented diet and by N-acetylcysteine, whereas amlodipine prevented Ca2+ loading and an altered redox state. Cotreatment with these interventions either markedly attenuated or prevented the appearance of the proinflammatory coronary vascular phenotype and the presence of 3-nitrotyrosine in invading inflammatory cells.. We suggest that the immunostimulatory state that appears during aldosteronism and leads to a proinflammatory coronary vascular phenotype is induced by a fall in [Mg2+]i with Ca2+ loading of PBMCs and is transduced by H2O2 production in these cells.

Topics: Acetylcysteine; Amlodipine; Animals; Biomarkers; Body Weight; Calcium; Dietary Supplements; Hydrogen Peroxide; Hyperaldosteronism; Leukocytes, Mononuclear; Magnesium; Male; Oxidation-Reduction; Oxidative Stress; Phenotype; Rats; Rats, Sprague-Dawley; Sodium Chloride, Dietary; Tyrosine; Vasculitis

Currently, obesity is considered a systemic inflammation; however, the effects of obesity on the vulnerability of dopaminergic neurons to oxidative stress are not fully defined. We evaluated the effects of high-fat diet-induced obesity (HF DIO) on neurotoxicity in mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). Eight weeks after a HF or matched normal diet, a severe decrease in the levels of striatal dopamine and of nigral microtubule-associated protein 2, manganese superoxide dismutase, and tyrosine hydroxylase was observed in obese mice treated with subtoxic doses of MPTP (20 mg/kg) compared with the matched lean group. In addition, the levels of nitrate/nitrite and thiobarbituric acid-malondialdehyde adducts in the substantia nigra of obese mice were reciprocally elevated or suppressed by MPTP. Interestingly, striatal nNOS phosphorylation and dopamine turnover were elevated in obese mice after MPTP treatment, but were not observed in lean mice. The nitrotyrosine immunoreactivity for evaluation of nigral nitrogenous stress in obese mice with MPTP was higher than that in matched lean mice. At higher doses of MPTP (60 mg/kg), the mortality was higher in obese mice than in lean mice. These results suggest that DIO may increase the vulnerability of dopaminergic neurons to MPTP via increased levels of reactive oxygen and nitrogen species, and the role of nNOS phosphorylation in the MPTP toxicities and dopamine homeostasis should be further evaluated.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animal Feed; Animals; Antioxidants; Astrocytes; Blotting, Western; Body Weight; Chromatography, High Pressure Liquid; Dietary Fats; Disease Susceptibility; Dopamine; Dopamine Agents; Glial Fibrillary Acidic Protein; Immunoblotting; Immunohistochemistry; Lipid Peroxidation; Male; Mice; Mice, Inbred C57BL; Mice, Obese; MPTP Poisoning; Neurons; Neurotoxicity Syndromes; Nitrates; Nitric Oxide; Nitrites; Obesity; Oxidative Stress; Phosphorylation; Substantia Nigra; Superoxide Dismutase; Thiobarbituric Acid Reactive Substances; Time Factors; Tyrosine

Although diabetes is a major risk factor for vascular diseases, e.g., hypertension and atherosclerosis, mechanisms that underlie the "risky" aspects of diabetes remain obscure. The current study is intended to examine the notion that diabetic endothelial dysfunction stems from a heightened state of oxidative stress induced by an imbalance between vascular production and scavenging of reactive oxygen/nitrogen species. Goto-Kakizaki (GK) rats were used as a genetic animal model for non-obese type II diabetes. Nitric oxide (NO) bioavailability and O2- generation in aortic tissues of GK rats were assessed using the Griess reaction and a lucigenin-chemiluminescence-based technique, respectively. Organ chamber-based isometric tension studies revealed that aortas from GK rats had impaired relaxation responses to acetylcholine whereas a rightward shift in the dose-response curve was noticed in the endothelium-independent vasorelaxation exerted by the NO donor sodium nitroprusside. An enhancement in superoxide (O2-) production and a diminuation in NO bioavailability were evident in aortic tissues of GK diabetic rats. Immunoblotting and high-performance liquid chromatography (HPLC)-based techniques revealed, respectively, that the above inverse relationship between O2- and NO was associated with a marked increase in the protein expression of nitric oxide synthase (eNOS) and a decrease in the level of its cofactor tetrahydrobiopterin (BH4) in diabetic aortas. Endothelial denudation by rubbing or the addition of pharmacological inhibitors of eNOS (e.g. N(omega)-nitro-L-arginine methyl ester (L-NAME)), and NAD(P)H oxidase (e.g. diphenyleneiodonium, apocynin) strikingly reduced the diabetes-induced enhancement in vascular O2- production. Aortic contents of key markers of oxidative stress (isoprostane F2alpha III, protein-bound carbonyls, nitrosylated protein) in connection with the protein expression of superoxide generating enzyme NAD(P)H oxidase (e.g. p47phox, pg91phox), a major source of reactive oxygen species in vascular tissue, were elevated as a function of diabetes. In contrast, the process involves in the vascular inactivation of reactive oxygen species exemplified by the activity of CuZnSOD was reduced in this diseased state. Our studies suggest that diabetes produces a cascade of events involving production of reactive oxygen species from the NADPH oxidase leading to oxidation of BH4 and uncoupling of NOS. This promotes the oxidative inactivation of NO with

Topics: Animals; Aorta, Thoracic; Biopterins; Body Weight; Diabetes Mellitus, Type 2; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelium, Vascular; Enzyme Inhibitors; Female; Humans; In Vitro Techniques; Isoprostanes; Male; NADPH Oxidases; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Nitroprusside; Oxidation-Reduction; Protein Subunits; Proteins; Rats; Rats, Inbred Strains; Rats, Wistar; Superoxide Dismutase; Superoxides; Tyrosine; Vasodilation; Vasodilator Agents

Iron deficiency is associated with multiple health problems, including the cardiovascular system. However, the mechanism of action of iron-deficiency-induced cardiovascular damage is unclear. The aim of the present study was to examine the effect of dietary iron deficiency on cardiac ultrastructure, mitochondrial cytochrome c release, NOS (nitric oxide synthase) and several stress-related protein molecules, including protein nitrotyrosine, the p47phox subunit of NADPH oxidase, caveolin-1 and RhoA. Male weanling rats were fed with either control or iron-deficient diets for 12 weeks. Cardiac ultrastructure was examined by transmission electron microscopy. Western blot analysis was used to evaluate cytochrome c, endothelial and inducible NOS, NADPH oxidase, caveolin-1 and RhoA. Protein nitrotyrosine formation was measured by ELISA. Rats fed an iron-deficient diet exhibited increased heart weight and size compared with the control group. Heart width, length and ventricular free wall thickness were similar between the two groups. However, the left ventricular dimension and chamber volume were significantly enhanced in the iron-deficient group compared with controls. Ultrastructural examination revealed mitochondrial swelling and abnormal sarcomere structure in iron-deficient ventricular tissues. Cytochrome c release was significantly enhanced in iron-deficient rats. Protein expression of eNOS (endothelial NOS) and iNOS (inducible NOS), and protein nitrotyrosine formation were significantly elevated in cardiac tissue or mitochondrial extraction from the iron-deficient group. Significantly up-regulated NADPH oxidase, caveolin-1 and RhoA expression were also detected in ventricular tissue of the iron-deficient group. Taken together, these results suggest that dietary iron deficiency may have induced cardiac hypertrophy characterized by aberrant mitochondrial and irregular sarcomere organization, which was accompanied by increased reactive nitrogen species and RhoA expression.

Topics: Animals; Body Weight; Cardiomegaly; Cytochromes c; Heart Ventricles; Heat-Shock Proteins; Iron Deficiencies; Male; Microscopy, Electron; Mitochondria, Heart; Myocardium; Myocytes, Cardiac; Nitric Oxide Synthase; Organ Size; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Tyrosine

Cardiovascular disease is the most frequent cause of mortality in chronic renal failure (CRF). Therefore, it is important to identify appropriate treatment measures. The antioxidant N-acetylcysteine (NAC) has been shown to reduce cardiovascular events in hemodialysis patients. Here we examine a possible direct effect of NAC supplementation on uremia-enhanced atherosclerosis in apolipoprotein E-deficient (apoE(-/-)) mice.. Uremia was induced surgically in 8-week-old female apoE(-/-) mice. Two weeks after creation of CRF mice were randomized to receive either NAC (daily oral gavage with 200 mg/kg for 8 weeks) or placebo. They were compared to a control group of sham-operated apoE(-/-) mice receiving placebo. After 8 weeks of treatment, the mice were sacrificed, and the cross-section surface area of atherosclerotic plaques was measured in aortic root and descending aorta.. At 10 weeks following surgery, atherosclerotic lesions were significantly larger in uremic apoE(-/-) mice than in nonuremic controls. This accelerated atherosclerosis was associated with an increase in aortic nitrotyrosine expression and collagen plaque content. NAC treatment inhibited the progression of atherosclerotic lesions and plaque collagen content compared with placebo treatment. In addition, plaques from NAC-treated uremic animals showed a significant decrease in nitrotyrosine expression whereas the degree of macrophage infiltration was comparable in both uremic groups. There was no difference in mean arterial blood pressure between the three groups.. We show for the first time that the antioxidant NAC is capable of reducing atheroma progression, in an animal model of uremia-enhanced atherosclerosis, probably via a decrease in oxidative stress.

Topics: Acetylcysteine; Animals; Antioxidants; Aorta; Apolipoproteins E; Arteriosclerosis; Body Weight; Collagen; Female; Mice; Mice, Knockout; Tyrosine; Uremia

It is generally assumed that inflammatory bowel disease (IBD)-related carcinogenesis occurs as a result of chronic inflammation. We previously developed a novel colitis-related mouse colon carcinogenesis model initiated with azoxymethane (AOM) and followed by dextran sodium sulfate (DSS). In the present study we investigated whether a cyclooxygenase (COX)-2 inhibitor nimesulide and ligands for peroxisome proliferator-activated receptors (PPARs), troglitazone (a PPARgamma ligand) and bezafibrate (a PPARalpha ligand) inhibit colitis-related colon carcinogenesis using our model to evaluate the efficacy of these drugs in prevention of IBD-related colon carcinogenesis.. Female CD-1 (ICR) mice were given a single intraperitoneal administration of AOM (10 mg/kg body weight) and followed by one-week oral exposure of 2% (w/v) DSS in drinking water, and then maintained on the basal diets mixed with or without nimesulide (0.04%, w/w), troglitazone (0.05%, w/w), and bezafibrate (0.05%, w/w) for 14 weeks. The inhibitory effects of dietary administration of these compounds were determined by histopathological and immunohistochemical analyses.. Feeding with nimesulide and troglitazone significantly inhibited both the incidence and multiplicity of colonic adenocarcinoma induced by AOM/DSS in mice. Bezafibrate feeding significantly reduced the incidence of colonic adenocarcinoma, but did not significantly lower the multiplicity. Feeding with nimesulide and troglitazone decreased the proliferating cell nuclear antigen (PCNA)-labeling index and expression of beta-catenin, COX-2, inducible nitric oxide synthase (iNOS) and nitrotyrosine. The treatments increased the apoptosis index in the colonic adenocarcinoma. Feeding with bezafibrate also affected these parameters except for beta-catenin expression in the colonic malignancy.. Dietary administration of nimesulide, troglitazone and bezafibrate effectively suppressed the development of colonic epithelial malignancy induced by AOM/DSS in female ICR mice. The results suggest that COX-2 inhibitor and PPAR ligands could serve as an effective agent against colitis-related colon cancer development.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Bezafibrate; Body Weight; Chromans; Colitis; Colonic Neoplasms; Cyclooxygenase 2; Cyclooxygenase Inhibitors; Female; Hypolipidemic Agents; Immunohistochemistry; Inflammation; Injections, Intraperitoneal; Ligands; Male; Mice; Mice, Inbred ICR; Nitric Oxide Synthase Type II; Organ Size; Peroxisome Proliferator-Activated Receptors; Sulfonamides; Temperature; Thiazolidinediones; Troglitazone; Tyrosine; Vasodilator Agents

Recent studies suggest that adipose tissue hormone, leptin, is involved in the pathogenesis of arterial hypertension. However, the mechanism of hypertensive effect of leptin is incompletely understood. We investigated whether antioxidant treatment could prevent leptin-induced hypertension. Hyperleptinemia was induced in male Wistar rats by administration of exogenous leptin (0.25 mg/kg twice daily s.c. for 7 days) and separate groups were simultaneously treated with superoxide scavenger, tempol, or NAD(P)H oxidase inhibitor, apocynin (2 mM in the drinking water). After 7 days, systolic blood pressure was 20.6% higher in leptin-treated than in control animals. Both tempol and apocynin prevented leptin-induced increase in blood pressure. Plasma concentration and urinary excretion of 8-isoprostanes increased in leptin-treated rats by 66.9% and 67.7%, respectively. The level of lipid peroxidation products, malonyldialdehyde + 4-hydroxyalkenals (MDA+4-HNE), was 60.3% higher in the renal cortex and 48.1% higher in the renal medulla of leptin-treated animals. Aconitase activity decreased in these regions of the kidney following leptin administration by 44.8% and 45.1%, respectively. Leptin increased nitrotyrosine concentration in plasma and renal tissue. Urinary excretion of nitric oxide metabolites (NO(x)) was 57.4% lower and cyclic GMP excretion was 32.0% lower in leptin-treated than in control group. Leptin decreased absolute and fractional sodium excretion by 44.5% and 44.7%, respectively. Co-treatment with either tempol or apocynin normalized 8-isoprostanes, MDA+4-HNE, aconitase activity, nitrotyrosine, as well as urinary excretion of NO(x), cGMP and sodium in rats receiving leptin. These results indicate that oxidative stress-induced NO deficiency is involved in the pathogenesis of leptin-induced hypertension.

Topics: Acetophenones; Aconitate Hydratase; Aldehydes; Animals; Antioxidants; Blood Pressure; Body Weight; Creatine; Cyclic GMP; Cyclic N-Oxides; Drinking; Eating; Hypertension; Isoprostanes; Kidney; Leptin; Male; Malondialdehyde; Natriuresis; Nitric Oxide; Rats; Rats, Wistar; Reactive Nitrogen Species; Sodium; Spin Labels; Tyrosine

Reduced activity of the mitochondrial respiratory chain--particularly complex I--may be implicated in the etiology of both Parkinson's disease and progressive supranuclear palsy, although these neurodegenerative diseases differ substantially as to their distinctive pattern of neuronal cell loss and the predominance of cerebral alpha-synuclein or tau protein pathology. To determine experimentally whether chronic generalized complex I inhibition has an effect on the distribution of alpha-synuclein or tau, we infused rats systemically with the plant-derived isoflavonoid rotenone. Rotenone-treated rats with a pronounced metabolic impairment had reduced locomotor activity, dystonic limb posture and postural instability. They lost neurons in the substantia nigra and in the striatum. Spherical deposits of alpha-synuclein were observed in a few cells, but cells with abnormal cytoplasmic accumulations of tau immunoreactivity were significantly more numerous in the striatum of severely lesioned rats. Abnormally high levels of tau immunoreactivity were found in the cytoplasm of neurons, oligodendrocytes and astrocytes. Ultrastructurally, tau-immunoreactive material consisted of straight 15-nm filaments decorated by antibodies against phosphorylated tau. Many tau+ cell bodies also stained positive for thioflavin S, nitrotyrosine and ubiquitin. Some cells with abnormal tau immunoreactivity contained activated caspase 3. Our data suggest that chronic respiratory chain dysfunction might trigger a form of neurodegeneration in which accumulation of hyperphosphorylated tau protein predominates over deposits of alpha-synuclein.

Topics: alpha-Synuclein; Amyloid beta-Peptides; Analysis of Variance; Animals; Antineoplastic Combined Chemotherapy Protocols; Behavior, Animal; Benzothiazoles; Body Weight; Caspase 3; Caspases; Cell Death; Cerebral Cortex; Cytarabine; Diagnostic Imaging; Dopamine and cAMP-Regulated Phosphoprotein 32; Doxorubicin; Dystonia; Electron Transport Complex III; Enzyme Activation; Glial Fibrillary Acidic Protein; Immunohistochemistry; Locomotion; Male; Microscopy, Electron, Transmission; Mitochondria; Neurons; Phosphopyruvate Hydratase; Phosphorylation; Posture; Psychomotor Performance; Rats; Rats, Inbred Lew; Rotenone; tau Proteins; Tauopathies; Thiazoles; Time Factors; Tyrosine; Tyrosine 3-Monooxygenase; Ubiquitin; Uncoupling Agents

Obesity has emerged as a major cause of diabetes, cardiovascular disease, and renal insufficiency worldwide. Obese Zucker rats exhibit hyperphagia, obesity, insulin resistance, hyperlipidemia, and glomerulosclerosis and are frequently used as a model to study hereditary form of metabolic syndrome. Nitric oxide plays a major role in preservation of renal function and structure. The present study was designed to test the hypothesis that renal disease in this model may be associated with down-regulation of endothelial (eNOS) and neuromal NO synthases (nNOS) in the kidney. The study further sought to explore expressions of caveolin-1, phospho AKt, and calmodulin, which regulate activities of constituitive NOS isoforms, as well as soluble guanylate cyclase (sGC), which is involved in NO signaling.. Twenty-two-week-old male obese and lean Zucker rats were studied. Body weight, serum lipids, urine albumin excretion, and renal tissue abundance of the above proteins were determined.. Serum glucose and arterial pressure were unchanged, whereas urinary NO metabolite (NO(chi)) excretion and renal tissue nitrotyrosine abundance were markedly reduced (denoting depressed NO production) in the obese versus lean Zucker rats. This was accompanied by significant glomerulosclerosis, tubulointerstitial damage, renal immune cell infiltration, marked down-regulations of renal tissue eNOS and nNOS, mild reduction of caveolin-1, and unchanged calmodulin, phospho-AKt, and sGC.. Hereditary obesity can result in down-regulations of kidney eNOS and nNOS, marked reduction of NO production, and glomerulosclerosis prior to the onset of frank diabetes and hypertension.

Topics: Animals; Body Weight; Calmodulin; Caveolin 1; Down-Regulation; Guanylate Cyclase; Immunohistochemistry; Kidney; Male; Nitrates; Nitric Oxide Synthase Type I; Nitric Oxide Synthase Type III; Nitrites; Obesity; Phosphorylation; Proto-Oncogene Proteins c-akt; Rats; Rats, Zucker; Receptors, Cytoplasmic and Nuclear; Soluble Guanylyl Cyclase; Tyrosine

It has been shown that reactive oxygen species are involved in chronic puromycin aminonucleoside (PAN) induced nephrotic syndrome (NS) and that a 20% soy protein diet reduces renal damage in this experimental model. The purpose of the present work was to investigate if a 20% soy protein diet is able to modulate kidney nitrotyrosine formation and the activity of renal antioxidant enzymes (catalase, glutathione peroxidase, Cu,Zn- or Mn-superoxide dismutase) which could explain, at least in part, the protective effect of the soy protein diet in rats with chronic NS induced by PAN. Four groups of rats were studied: (1) Control rats fed 20% casein diet, (2) Nephrotic rats fed 20% casein diet, (3) Control rats fed 20% soy protein diet, and (4) Nephrotic rats fed 20% soy protein diet. Chronic NS was induced by repeated injections of PAN and rats were sacrificed at week nine. The soy protein diet ameliorated proteinuria, hypercholesterolemia, and the increase in serum creatinine and blood urea nitrogen observed in nephrotic rats fed 20% casein diet. Kidney nitrotyrosine formation increased in nephrotic rats fed 20% casein diet and this increase was ameliorated in nephrotic rats fed 20% soy protein diet. However, the soy protein diet was unable to modulate the antioxidant enzymes activities in control and nephrotic rats fed 20% soy protein diet. Food intake was similar in the two diet groups. The protective effect of a 20% soy protein diet on renal damage in chronic nephropathy induced by PAN was associated with the amelioration in the renal nitrotyrosine formation but not with the modulation of antioxidant enzymes.

Topics: Animals; Antimetabolites; Antioxidants; Blood Urea Nitrogen; Body Weight; Catalase; Cholesterol; Creatinine; Diet; Eating; Glutathione Peroxidase; Glycine max; Immunohistochemistry; Kidney; Kidney Failure, Chronic; Kidney Function Tests; Lipids; Male; Proteinuria; Puromycin Aminonucleoside; Rats; Rats, Wistar; Soybean Proteins; Superoxide Dismutase; Tyrosine

Aging and estrogen deficiency increase the risk for developing cardiovascular disease (CVD). Oxidative stress has also been implicated in the pathophysiology of CVD and in ischemia-reperfusion (I/R) injury. We tested the hypothesis that chronic in vivo estrogen treatment or superoxide inhibition with the SOD mimetic EUK-8 improves cardiac functional recovery after I/R in the aged female rat. Sprague-Dawley rats (12-14 mo) were used as follows: intact (n = 6), ovariectomized + placebo (OVX, n = 6), OVX + EUK-8 (EUK-8, 3 mg/kg, n = 6), and OVX + estrogen (1.5 mg/pellet, 60 days release, n = 6). Perfused isolated hearts were subjected to global ischemia (25 min) followed by reperfusion (40 min). Functional recovery after I/R and myocardial protein expression of NADPH oxidase (p22, p67, and gp91(phox)), inducible nitric oxide synthase (NOS), endothelial NOS, and SOD1, as well as nitrotyrosine levels (as a marker for peroxynitrite), were assessed. Compared with OVX, EUK-8 and estrogen markedly improved functional recovery after I/R, which was associated with a decrease in NADPH oxidase expression and nitrotyrosine staining. However, estrogen increased inducible NOS expression, whereas EUK-8 had little effect. There were no significant changes in endothelial NOS and SOD1 expression among the groups. These results indicate that EUK-8 and estrogen improved cardiac recovery after I/R. Given the controversy surrounding hormone replacement therapy, EUK-8 may be an alternative to estrogen in protecting those at risk for myocardial ischemia in the aging population.

Topics: Aging; Animals; Body Weight; Cardiotonic Agents; Estrogens; Ethylenediamines; Female; Heart; Heart Ventricles; Myocardial Ischemia; Myocardial Reperfusion Injury; NADPH Oxidases; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitric Oxide Synthase Type III; Organ Size; Organometallic Compounds; Ovariectomy; Rats; Rats, Sprague-Dawley; Recovery of Function; Superoxide Dismutase; Tyrosine; Uterus

Oxidative and nitrosative stress have been involved in gentamicin-induced nephrotoxicity. The purpose of this work was to study the effect of S-allylmercaptocysteine, a garlic derived compound, on gentamicin-induced oxidative and nitrosative stress and nephrotoxicity. In addition, the in vitro reactive oxygen species scavenging properties of S-allylmercaptocysteine were studied.. S-allylmercaptocysteine was able to scavenge hydroxyl radicals and singlet oxygen in vitro. In rats treated with gentamicin (70 mg/Kg body weight, subcutaneously, every 12 h, for 4 days), renal oxidative stress was made evident by the increase in protein carbonyl content and 4-hydroxy-2-nonenal, and the nitrosative stress was made evident by the increase in 3-nitrotyrosine. In addition, gentamicin-induced nephrotoxicity was evident by the: (1) decrease in creatinine clearance and in activity of circulating glutathione peroxidase, and (2) increase in urinary excretion of N-acetyl-beta-D-glucosaminidase, and (3) necrosis of proximal tubular cells. Gentamicin-induced oxidative and nitrosative stress and nephrotoxicity were attenuated by S-allylmercaptocysteine treatment (100 mg/Kg body weight, intragastrically, 24 h before the first dose of gentamicin and 50 mg/Kg body weight, intragastrically, every 12 h, for 4 days along gentamicin-treatment).. In conclusion, S-allylmercaptocysteine is able to scavenge hydroxyl radicals and singlet oxygen in vitro and to ameliorate the gentamicin-induced nephrotoxicity and oxidative and nitrosative stress in vivo.

Topics: Aldehydes; Animals; Biomarkers; Body Weight; Carboxylic Acids; Cysteine; Dinitrophenols; Energy Intake; Free Radical Scavengers; Gentamicins; Hydroxyl Radical; Immunohistochemistry; Kidney Cortex; Kidney Diseases; Kidney Glomerulus; Kidney Tubules, Proximal; Male; Oxidation-Reduction; Oxidative Stress; Proteins; Rats; Rats, Wistar; Reactive Oxygen Species; Tyrosine; Urine

Oxidative stress is involved in cisplatin-nephrotoxicity. However, it has not completely established if reactive nitrogen species and nitrosative stress are involved in this experimental model. The purpose of this work was to study the role of peroxynitrite, a reactive nitrogen specie, in cisplatin-nephrotoxicity using the compound 5,10,15,20-tetrakis (4-sulfonatophenyl) porphyrinato iron (III) (FeTPPS), a soluble complex able to metabolize peroxynitrite.. In rats treated with cisplatin (a single intraperitoneal dose of 7.5 mg/kg body weight), renal nitrosative stress was made evident by the increase in 3-nitrotyrosine on day 3. In addition, cisplatin-induced nephrotoxicity was evident by the histological damage of proximal tubular cells and by the increase in (a) serum creatinine, (b) blood urea nitrogen, and (c) urinary excretion of N-acetyl-beta-D-glucosaminidase and total protein. Cisplatin-induced nitrosative stress and nephrotoxicity were attenuated by FeTPPS-treatment (15 mg/kg body weight, intraperitoneally, every 12 hours for 3 days).. Nitrosative stress is involved in cisplatin-induced nephrotoxicity in rats. Our data suggest that peroxynitrite is involved, at least in part, in cisplatin-induced nephrotoxicity and protein nitration.

Topics: Animals; Body Weight; Catalysis; Cisplatin; Glomerular Mesangium; Kidney Diseases; Kidney Tubules; Male; Metalloporphyrins; Peroxynitrous Acid; Rats; Rats, Wistar; Reactive Nitrogen Species; Tyrosine; Urine

Chronic consumption of a high-fat, refined-carbohydrate (HFS) diet causes hypertension. In an earlier study, we found increased nitric oxide (NO) inactivation by reactive oxygen species (ROS) and functional NO deficiency in this model. Given the critical role of NO in renal sodium handling, we hypothesized that diet-induced hypertension may be associated with salt sensitivity. Female Fischer rats were fed an HFS or a standard low-fat, complex-carbohydrate (LFCC) rat chow diet starting at 2 mo of age for 2 yr. Arterial blood pressure, renal neuronal NO synthase (nNOS), endothelial NO synthase (eNOS), and inducible NO synthase (iNOS) protein and nitrotyrosine abundance (a marker of NO inactivation by ROS), and urinary NO metabolite excretion were measured. To assess salt sensitivity, the blood pressure response to a high-salt (4%) diet for 1 wk was determined. After 2 yr, renal nNOS and urinary NO metabolite excretion were significantly depressed, whereas arterial pressure, eNOS, iNOS, and nitrotyrosine were elevated in the HFS group but remained virtually unchanged in the LFCC group. Consumption of the high-salt diet resulted in a significant rise in arterial pressure in the HFS, but not in the LFCC, group. Thus chronic consumption of an HFS diet results in hypertension and salt sensitivity, which may be in part due to a combination of ROS-mediated NO inactivation and depressed renal nNOS protein expression.

Topics: Animals; Blood Pressure; Blotting, Western; Body Weight; Diet; Dietary Carbohydrates; Dietary Fats; Female; Isoenzymes; Kidney; Nitric Oxide; Nitric Oxide Synthase; Rats; Rats, Inbred F344; Sodium Chloride; Tyrosine

The purpose of these experiments was to determine the specific role of reactive oxygen species (ROS) in the blood-retinal barrier (BRB) breakdown that characterizes the early stages of vascular dysfunction in diabetes. Based on our data showing that high glucose increases nitric oxide, superoxide, and nitrotyrosine formation in retinal endothelial cells, we hypothesized that excess formation of ROS causes BRB breakdown in diabetes. Because ROS are known to induce increases in expression of the well-known endothelial mitogen and permeability factor vascular endothelial growth factor (VEGF) we also examined their influence on the expression of VEGF and its downstream target urokinase plasminogen activator receptor (uPAR). After 2 weeks of streptozotocin-induced diabetes, analysis of albumin leakage confirmed a prominent breakdown of the BRB. This permeability defect was correlated with significant increases in the formation of nitric oxide, lipid peroxides, and the peroxynitrite biomarker nitrotyrosine as well as with increases in the expression of VEGF and uPAR. Treatment with a nitric oxide synthase inhibitor (N-omega-nitro-L-arginine methyl ester, 50 mg/kg/day) or peroxynitrite scavenger (uric acid, 160 mg/kg/day) blocked the breakdown in the BRB and prevented the increases in formation of lipid peroxides and tyrosine nitration as well as the increases in expression of VEGF and uPAR. Taken together, these data indicate that early diabetes causes breakdown of the BRB by a mechanism involving the action of reactive nitrogen species in promoting expression of VEGF and uPAR.

Topics: Animals; Blood-Retinal Barrier; Body Weight; Diabetes Mellitus, Experimental; Endothelial Growth Factors; Enzyme Inhibitors; Gene Expression; Intercellular Signaling Peptides and Proteins; Lipid Peroxidation; Lymphokines; NG-Nitroarginine Methyl Ester; Nitrates; Nitric Oxide Synthase; Nitrites; Oxidative Stress; Peroxynitrous Acid; Rats; Rats, Sprague-Dawley; Reactive Nitrogen Species; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Retina; RNA, Messenger; Tyrosine; Uric Acid; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Gentamicin (GM) is an antibiotic whose clinical use is limited by its nephrotoxicity. Experimental evidences suggest a role of reactive oxygen species in GM-induced nephrotoxicity. In this work we explored the effect of diallyl sulfide (DAS), a garlic-derived compound with antioxidant properties, on GM-induced nephrotoxicity. Four groups of rats were studied: (1) Control, treated intragastrically with olive oil as a vehicle, (2) GM, treated subcutaneously with GM (125 mg/kg/day for 4 days), (3) DAS, treated intragastrically with DAS (50 mg/kg/day for 4 days), and (4) GM + DAS. Nephrotoxicity was made evident by: (1) the increase in creatinine and blood urea nitrogen in serum, (2) the increase in urinary excretion of N-acetyl-beta-D-glucosaminidase and total protein, and (3) necrosis of proximal tubular cells. These functional and structural alterations were prevented or ameliorated by DAS treatment. In addition, GM increased levels of renal oxidative stress markers nitrotyrosine and protein carbonyl groups which were also ameliorated by DAS in GM + DAS group. The mechanism by which DAS has a protective effect on GM-induced nephrotoxicity may be related, at least in part, to the decrease in oxidative stress in renal cortex.

Topics: Acetylglucosaminidase; Allyl Compounds; Animals; Anti-Bacterial Agents; Antioxidants; Blood Urea Nitrogen; Body Weight; Carbon; Creatinine; Gentamicins; Immunohistochemistry; Kidney; Kidney Cortex; Kidney Tubules; Male; Necrosis; Olive Oil; Oxidative Stress; Oxygen; Plant Oils; Rats; Rats, Wistar; Reactive Oxygen Species; Sulfides; Time Factors; Tyrosine

The objective of this study was to address the hepatic effects of acute alcohol consumption in obesity by simulating an alcohol binge in genetically obese fa/fa rats compared with lean Fa/? rats.. Ethanol 4 g/kg or saline was administered by gavage every 12 hours for 3 days.. Plasma alcohol levels were similar in both groups. Binge ethanol exposure caused liver injury in obese fa/fa but not in lean Fa/? rats, as assessed by alanine aminotransferase and H&E staining. Obesity impaired the antioxidant defense because basal levels of glutathione, glutamate cysteine ligase modulatory subunit, catalase, glutathione reductase, and superoxide dismutase were lower in fa/fa compared with Fa/? rats; the ethanol binge further decreased these antioxidants in fa/fa rats and also decreased glutathione peroxidase activity. Nonesterified fatty acids and lipid peroxidation were increased after ethanol treatment in fa/fa rats. Cytochrome P450 2E1 was down-regulated in fa/fa compared with Fa/? rats; however, the ethanol binge increased cytochrome P450 2E1 in both genotypes. Adenosine triphosphate decreased and uncoupling protein 2 increased in fa/fa rats treated with ethanol. 3-Nitrotyrosine protein adducts were detected only in fa/fa rats treated with ethanol, and this was accompanied by an induction of inducible nitric oxide synthase. Ethanol binge increased caspase-3 and caspase-8 activity, the expression of Fas ligand, and terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling in fa/fa rats.. These data indicate that binge drinking increases apoptosis and liver injury in obese rats more than in lean controls and suggest that the injury may involve oxidative and nitrosative damage.

Topics: Adenosine Triphosphate; Animals; Apoptosis; Body Weight; Cytochrome P-450 CYP2E1; Ethanol; Fatty Acids, Nonesterified; Glutathione; Ion Channels; Lipid Peroxidation; Liver; Male; Membrane Transport Proteins; Mitochondrial Proteins; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Obesity; Organ Size; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Zucker; Tyrosine; Uncoupling Protein 2

Oxidative stress contributes to early alcohol-induced liver injury, and superoxide (O(2)*-) production from NADPH oxidase plays a key role. However, the production of the free radical nitric oxide (NO*) by inducible nitric oxide synthase (iNOS) could also be involved.. To test this hypothesis, iNOS knockout (B6.129P2-Nos2 (tm1 Lau)) and wild-type mice were fed high-fat control or ethanol-containing diets for 4 weeks.. Mean body weight gains were not significantly different between treatment groups, and average urine ethanol concentrations were similar in wild-type and iNOS knockout mice. After 4 weeks, serum alanine aminotransferase (ALT) levels were increased significantly about 4-fold over control values (29 +/- IU/L) by enteral ethanol (113 +/- 20) in wild-type mice; this effect of ethanol was significantly blunted in iNOS knockout mice (50 +/- 9). Similar protective effects against liver damage were observed if wild-type mice were treated with the iNOS inhibitor N -(3-aminomethyl)benzyl-acetamindine (1400W). Enteral ethanol also caused severe fatty accumulation, mild inflammation, and necrosis in the liver in wild-type mice but had no effect in iNOS knockout mice. The accumulation of 4-hydroxynonenal (lipid peroxidation) and 3-nitrotyrosine (reactive nitrogen species formation) protein adducts caused by alcohol was completely blocked in iNOS knockout mice.. These data strongly support the hypothesis that iNOS is required for the pathogenesis of early alcohol-induced hepatitis by production of nitric oxide-derived pro-oxidants (e.g., peroxynitrite).

Topics: Alanine Transaminase; Aldehydes; Animals; Aspartate Aminotransferases; Body Weight; Cytochrome P-450 CYP2E1; Endotoxins; Ethanol; Free Radicals; Liver; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; RNA, Messenger; Tumor Necrosis Factor-alpha; Tyrosine

Free radicals appear to participate in the final common pathway of neuronal death in ischemia and may therefore be an adequate target for therapy. Tempol is a nitroxide antioxidant with proven protective efficacy in several animal models, including myocardial ischemia, that has not been previously tested in models of permanent cerebral ischemia. Spontaneously hypertensive rats underwent permanent middle cerebral artery occlusion (PMCAO). Following dose-response and time-window-finding experiments rats were given vehicle or tempol (50 mg/kg) subcutaneously 1 h after PMCAO (n = 10/group). Five animals in each group were evaluated with a motor scale 24 h after the infarct and were then sacrificed and the injury volume was measured. The remaining animals were examined daily with the motor scale and also with a Morris water maze test on days 26-30 after PMCAO and sacrificed on day 30. Motor scores at all time points examined were significantly better in the tempol-treated animals (P < 0.05 for all). Significantly better performance in the water maze test for performance on days 26-30 was noted in the tempol group compared with the vehicle-treated group (P < 0.05). Injury volumes at days 1 and 30 were significantly reduced in the tempol group (9.83 +/- 1.05 vs 19.94 +/- 1.43% hemispheric volume, P = 0.0009, and 13.2 +/- 2.97 vs 24.4 +/- 2.38% hemispheric volume, P = 0.02, respectively). In conclusion, treatment with tempol led to significant motor and behavioral improvement and reduced injured tissue volumes both in the short and in the long term after stroke.

Topics: Animals; Antioxidants; Behavior, Animal; Body Weight; Brain; Brain Chemistry; Brain Ischemia; Cyclic N-Oxides; Disease Models, Animal; Disease Progression; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Infarction, Middle Cerebral Artery; Male; Maze Learning; Motor Activity; Neuroprotective Agents; Rats; Rats, Inbred SHR; Spin Labels; Treatment Outcome; Tyrosine

The potential utility of dietary supplementation in order to prevent some of the oxidative and inflammatory changes occurring in the brain with age, has been studied. The cerebral cortex of 27-month-old male B6C3F1 mice had elevated levels of nitric oxide synthase 1 (EC 1.14.13.39) (nNOS) and peptide nitrotyrosine relative to cortices of younger (4-month-old) animals. After 25-month-old mice received basal diet together with 300 mg/l acetyl L-carnitine in the drinking water for 8 weeks, these levels were fully restored to those found in younger animals. A partial restoration was found when old animals received basal diet supplemented with 200 ppm melatonin in the diet. Levels of mRNA (messenger RNA) for nNOS were unchanged following these treatments implying translational regulation of nNOS activity. Behavioral indices indicative of exploratory behavior were also depressed in aged animals. Dietary supplementation with melatonin or acetyl L-carnitine partially reversed these changes. These findings suggest that dietary supplementation cannot merely arrest but indeed reverse some age-related increases in markers of oxidative and inflammatory events occurring with the cortex.

Topics: Acetylcarnitine; Aging; Animals; Body Weight; Cerebral Cortex; Down-Regulation; Encephalitis; Exploratory Behavior; Male; Melatonin; Mice; Mice, Inbred Strains; Nitric Oxide; Nitric Oxide Synthase; Oxidative Stress; RNA, Messenger; Tyrosine

Nuclear factor-kappaB (NF-kappaB)-dependent up-regulation of inflammatory cytokines and inducible nitric oxide (iNOS) occurs in inflammatory bowel disease. We investigated the effect of alpha-phenylN-tert-butylnitrone (PBN), a spin-trapping agent that inhibits NF-kappaB activity, on dextran sulfate sodium (DSS)-induced colonic mucosal injury and inflammation in mice. Acute colitis was induced with DSS in female BALB/c mice receiving 0, 0.3, 3, and 30 mg/kg i.p. PBN daily. Colonic mucosal inflammation was evaluated biochemically and histologically. Nitric oxide was evaluated as luminal nitrite/nitrite concentration by the Griess reaction and as immunoreactive nitrotyrosine in mucosal cells. Mucosal tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) were determined by immunoassay. Colonic mRNA expression for iNOS, TNF-alpha, and IFN-gamma was measured by reverse transcription-polymerase chain reaction, and NF-kappaB activation was evaluated by electrophoretic mobility shift assay. After DSS administration, mice showed increased luminal nitrite/nitrate, mucosal TNF-alpha and IFN-gamma, and mRNA for iNOS and these cytokines, in addition to decreased colonic length and increased inflammatory score, luminal hemoglobin, and colonic myeloperoxidase activity. PBN inhibited increases in luminal nitric oxide production, nitrotyrosine immunoreactivity, and mucosal TNF-alpha and IFN-gamma. Colonic iNOS, TNF-alpha, and IFN-gamma mRNA were suppressed by PBN, as was a DSS-induced increase in colonic NF-kappaB DNA-binding activity. NF-kappaB is essential to DSS-induced colitis, suggesting molecular approach targeting of NF-kappaB for treatment of inflammatory bowel disease.

Topics: Animals; Body Weight; Colitis; Colon; Cyclic N-Oxides; Dextran Sulfate; Female; Immunohistochemistry; Inflammation; Interferon-gamma; Mice; Mice, Inbred BALB C; Neutrophils; NF-kappa B; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Nitrogen Oxides; Protein Binding; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Spin Labels; Tumor Necrosis Factor-alpha; Tyrosine; Up-Regulation

Recent studies have suggested that inducible nitric oxide synthase (iNOS) plays a role in the development of asbestos-related pulmonary disorders. The pulmonary reactions of rats and hamsters upon exposure to asbestos fibers are well known to be disparate. In addition, in vitro experiments have indicated that mononuclear phagocytes from hamsters, in contrast to those from rats, lack the iNOS pathway. Therefore, the purpose of this study was to investigate whether rats and hamsters differ in lung iNOS expression in vivo upon exposure to asbestos fibers and whether differences in iNOS induction are associated with differences in the acute pulmonary inflammatory reaction. Body weight, alveolar-arterial oxygen difference, differential cell count in bronchoalveolar lavage fluid, total protein leakage, lung myeloperoxidase activity and lipidperoxidation, wet/dry ratio, iNOS mRNA and protein expression, and nitrotyrosine staining of lung tissue were determined 1 and 7 days after intratracheal instillation of asbestos fibers in CD rats and Syrian golden hamsters. Exposure of rats to asbestos fibers resulted in enhanced pulmonary iNOS expression and nitrotyrosine staining together with an acute inflammation that was characterized by an influx of neutrophils, enhanced myeloperoxidase activity and lipid peroxidation, damage of the alveolar-capillary membrane, edema formation, and impairment of gas exchange. In comparison, instillation of asbestos fibers in hamsters resulted in a significantly milder inflammatory reaction of the lung with no induction of iNOS in pulmonary cells. The data obtained provide important information to understand the underlying mechanisms of species differences in the pulmonary response upon exposure to asbestos fibers.

Topics: Animals; Asbestos, Crocidolite; Asbestosis; Body Weight; Bronchoalveolar Lavage Fluid; Cell Count; Cricetinae; Disease Models, Animal; Inhalation Exposure; Intubation, Intratracheal; Lung; Mesocricetus; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Oxygen; Peroxidase; Rats; RNA, Messenger; Species Specificity; Thiobarbituric Acid Reactive Substances; Tyrosine

Oxygen-derived free radicals are involved in the vascular response to angiotensin II (Ang II), but the role of NADPH oxidase, its subunit proteins, and their vascular localization remain controversial. Our purpose was to address the role of NADPH oxidase in the blood pressure (BP), aortic hypertrophic, and oxidant responses to Ang II by taking advantage of knockout (KO) mice that are genetically deficient in gp91(phox), an NADPH oxidase subunit protein. The baseline BP was significantly lower in KO mice than in wild-type (WT) (92+/-2 [KO] versus 101+/-1 [WT] mm Hg, P<0.01), but infusion of Ang II for 6 days caused similar increases in BP in the 2 strains (33+/-4 [KO] versus 38+/-2 [WT] mm Hg, P>0.4). Ang II increased aortic superoxide anion production 2-fold in the aorta of WT mice but did not do so in KO mice. Aortic medial area increased in WT (0.12+/-0.02 to 0.17+/-0.02 mm(2), P<0.05), but did not do so in KO mice (0.10+/-0.01 to 0.11+/-0.01 mm(2), P>0.05). Histochemistry and polymerase chain reaction demonstrated gp91(phox) localized in endothelium and adventitia of WT mice. Levels of reactive oxidant species as indicated by 3-nitrotyrosine immunoreactivity increased in these regions in WT but not in KO mouse aorta in response to Ang II. These results indicate an essential role in vivo of gp91(phox) and NADPH oxidase-derived superoxide anion in the regulation of basal BP and a pressure-independent vascular hypertrophic and oxidant stress response to Ang II.

Topics: Angiotensin II; Animals; Aorta; Blood Pressure; Blood Vessels; Body Weight; Genotype; Hypertrophy; Immunohistochemistry; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Mice, Knockout; NADPH Oxidase 2; NADPH Oxidases; Oxidative Stress; RNA, Messenger; Superoxides; Tyrosine

1. To further explore the effect of antioxidants in preventing diabetes-induced vascular and neural dysfunction we treated streptozotocin-induced diabetic rats daily with subcutaneous injections of 10 mg kg(-1) of M40403 (n=11) and compared the results obtained from 17 control rats and 14 untreated diabetic rats. M40403 is a manganese(II) complex with a bis(cyclo-hexylpyridine)-substituted macrocyclic ligand that was designed to be a selective functional mimetic of superoxide dismutase. Thus, M40403 provides a useful tool to evaluate the roles of superoxide in disease states. 2. Treatment with M40403 significantly improved diabetes-induced decrease in endoneurial blood flow, acetylcholine-mediated vascular relaxation in arterioles that provide circulation to the region of the sciatic nerve, and motor nerve conduction velocity (P<0.05). M40403 treatment also reduced the appearance of superoxide in the aorta and epineurial vessels and peroxynitrite in epineurial vessels. Treating diabetic rats with M40403 reduced the diabetes-induced increase in thiobarbituric acid reactive substances in serum but did not prevent the decrease in lens glutathione level. Treating diabetic rats with M40403 did not improve sciatic nerve Na(+)/K(+) ATPase activity or the sorbitol, fructose or myo-inositol content of the sciatic nerve. 3. These studies provide additional evidence that diabetes-induced oxidative stress and the generation of superoxide and perhaps peroxynitrite may be partially responsible for the development of diabetic vascular and neural complications.

Topics: Acetylcholine; Animals; Blood Glucose; Blood Vessels; Body Weight; Diabetes Mellitus, Experimental; Dose-Response Relationship, Drug; Fatty Acids, Nonesterified; Fructose; Inositol; Male; Manganese; Neural Conduction; Organometallic Compounds; Rats; Rats, Sprague-Dawley; Regional Blood Flow; Sciatic Nerve; Sodium-Potassium-Exchanging ATPase; Sorbitol; Superoxides; Thiobarbituric Acid Reactive Substances; Triglycerides; Tyrosine; Vasodilation; Vasodilator Agents

Previous work found that dietary l-arginine alters symptom progression in mice transgenic for Huntington's disease (HD), and that cerebral blood flow (CBF) is abnormal in early stage HD patients. Both of these findings potentially implicate nitric oxide (NO) and its converting enzyme, nitric oxide synthase (NOS), in HD. The current experiment found that both NOS enzymatic activity and neuronal NOS (nNOS) protein expression were reduced (P<0.05) in R6/2 HD transgenic mice compared to non-HD controls (CON). Conversely, inducible NOS (iNOS) protein expression was not significantly different between groups. The changes in nNOS were accompanied by changes in protein expression of calmodulin kinase II (CaMKII) (P<0.05) and calmodulin kinase IV (CaMKIV) (P<0.05). Protein expression of 3-nitrotyrosine (3-NT), a marker for the neurotoxin peroxynitrite, was slightly increased in non-drug treated HD and was accompanied by increased immunostaining of 3-NT in cells adhering to the vasculature and choroid plexus. Mice that received the broad-spectrum NOS inhibitor N(g)-nitro-L-arginine methyl ester hydrochloride (L-NAME) via their drinking water had reduced NOS enzyme activity. NOS activity varied as a function of L-NAME dose, was virtually eliminated in the 500-mg/l groups, and correlated (P<0.05) with the behavioral scores as revealed by regression and correlation analyses. High dose L-NAME (500 mg/l) accelerated symptom onset in HD transgenics. These results support the hypothesis that nNOS activity and NO production are abnormal in HD, this in the setting of a more global dysregulation of calcium protein expression. Taken collectively with earlier data from our laboratory demonstrating abnormal CBF findings in early-stage HD patients, these results suggest that abnormalities in NOS function may significantly contribute to the neurodegeneration found in HD.

Topics: Animals; Behavior, Animal; Body Weight; Brain; Calcium-Calmodulin-Dependent Protein Kinases; Disease Models, Animal; Enzyme Inhibitors; Female; Huntington Disease; Immunohistochemistry; Male; Mice; Mice, Transgenic; Motor Activity; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Tyrosine

Inflammatory bowel disease is characterised by oxidative and nitrosative stress, leukocyte infiltration, and up-regulation of intercellular adhesion molecule 1 (ICAM-1) expression in the colon. The aim of the present study was to examine the effects of M40403, a superoxide dismutase mimetic, in rats subjected to experimental colitis. Colitis was induced in rats by intracolonic instillation of trinitrobenzene sulfonic acid (TNBS). Rats experienced bloody diarrhoea and significant loss of body weight. At 4 days after TNBS administration, the colon damage was characterised by areas of mucosal necrosis. Neutrophil infiltration (indicated by myeloperoxidase activity in the mucosa) was associated with up-regulation of ICAM-1 and expression of P-selectin and high levels of malondialdehyde. Immunohistochemistry for nitrotyrosine and poly (ADP-ribose) synthetase showed an intense staining in the inflamed colon. Treatment with M40403 (5 mg/kg daily i.p.) significantly reduced the appearance of diarrhoea and the loss of body weight. This was associated with a remarkable amelioration of the disruption of the colonic architecture as well as a significant reduction of colonic myeloperoxidase activity and malondialdehyde levels. M40403 also reduced the appearance of nitrotyrosine and poly (ADP-ribose) synthetase immunoreactivity in the colon as well as reduced the up-regulation of ICAM-1 and the expression of P-selectin. The results of this study suggested that administration of a superoxide dismutase mimetic may be beneficial for treatment of inflammatory bowel disease.

Topics: Animals; Body Weight; Colitis; Colon; Cytokines; Enzyme Activation; Free Radical Scavengers; Immunohistochemistry; Intercellular Adhesion Molecule-1; Lipid Peroxidation; Male; Manganese; Organometallic Compounds; P-Selectin; Peroxidase; Poly(ADP-ribose) Polymerases; Rats; Rats, Sprague-Dawley; Superoxide Dismutase; Survival Analysis; Time Factors; Tyrosine

To investigate the effects of tempol, a membrane-permeable radical scavenger, in rats with collagen-induced arthritis (CIA).. CIA was induced in Lewis rats by intradermal injection of 100 microl of an emulsion of 100 microg of bovine type II collagen (CII) in complete Freund's adjuvant (FCA) at the base of the tail. On day 21, a second injection of CII in FCA was administered.. Lewis rats developed an erosive arthritis of the hind paws when immunized with CII in FCA. Macroscopic evidence of CIA first appeared as periarticular erythema and edema in the hind paws. The incidence of CIA was 100% by day 27 in the CII-challenged rats, and the severity of CIA progressed over a 35-day period. Radiographs revealed focal resorption of bone, with osteophyte formation in the tibiotarsal joint, and soft tissue swelling. The histopathologic features included erosion of the cartilage at the joint margins. Treatment of rats with tempol (10 mg/kg/day intraperitoneally) starting at the onset of arthritis (day 23) delayed the development of the clinical signs on days 24-35 and improved the histologic status of the knee and paw. Immunohistochemical analysis for nitrotyrosine and poly(ADP-ribose) synthetase (PARS) revealed positive staining in the inflamed joints of CII-treated rats. The degree of nitrotyrosine and PARS staining was markedly reduced in tissue sections obtained from CII-treated rats that had received tempol. Furthermore, radiographs revealed protection against bone resorption and osteophyte formation in the joints of tempol-treated rats.. This study is the first to provide evidence that tempol, a small molecule that permeates biologic membranes and scavenges reactive oxygen species, attenuates the degree of chronic inflammation and tissue damage associated with CIA in the rat.

Topics: Animals; Arthritis, Experimental; Body Weight; Cyclic N-Oxides; Disease Models, Animal; Enzyme Activation; Free Radical Scavengers; Joints; Kinetics; Male; Poly(ADP-ribose) Polymerases; Radiography; Rats; Rats, Inbred Lew; Severity of Illness Index; Spin Labels; Time Factors; Tyrosine

Positioning a silicone collar around the rabbit carotid artery induces a smooth muscle cell-rich intimal thickening. We investigated the localization of inducible nitric oxide synthase (iNOS) during thickening of the intima, the effect of iNOS inhibition on intimal thickness, and the effect of oxidized LDL (ox-LDL) on iNOS expression in the vessel wall. Collars were positioned for 18 hours or for 3, 7, or 14 days. Arterial cross sections were immunostained for iNOS, including naïve, sham-operated, and carotid arteries in which ox-LDL had been infused locally for 14 days. Furthermore, collars were connected to osmotic minipumps for local delivery (5 microL. h(-1), 14 days, n=12) of saline or the iNOS inhibitor L-N(6)-(1-iminoethyl)-lysine-HCl (L-NIL, 10 mmol/L). In the adventitia and the periadventitial granulation tissue of collared arteries, iNOS-positive macrophages and T lymphocytes were present from 18 hours onward. The media and intima were negative for iNOS. Reverse transcription-polymerase chain reaction revealed iNOS mRNA in collared but not in sham-operated arteries. Local inhibition of iNOS doubled the intimal thickness and decreased nitrotyrosine staining. Ox-LDL-treated arteries, which had a thicker intima, showed a pronounced influx of macrophages and T lymphocytes in all layers of the vessel wall, accompanied by iNOS expression in a subpopulation of these cells. Our study indicates that iNOS was not induced in intimal thickenings predominantly consisting of smooth muscle cells. However, iNOS was expressed in (peri)adventitial tissue and counteracted the progression of intimal thickening. Ox-LDL treatment was accompanied by an abundant influx of iNOS-positive macrophages and T lymphocytes in the vessel, but this could not prevent the progression of intimal thickening.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Blood Pressure; Body Weight; Carotid Arteries; Enzyme Inhibitors; Gene Expression; Immunohistochemistry; Leukosialin; Macrophages; Male; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rabbits; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sialoglycoproteins; T-Lymphocytes; Tyrosine

We have recently demonstrated that long-term consumption of a high-fat, refined-carbohydrate (HFS) diet induces hypertension (HTN) in normal rats compared with a low-fat, complex-carbohydrate (LFCC) diet. Limited evidence suggests that high-fat or high-sugar diets cause enhanced generation of reactive oxygen species (ROS). We therefore hypothesized that by inducing oxidative stress, the HFS diet may promote nitric oxide (NO) inactivation and HTN. To test this hypothesis, female Fischer rats were placed on either the HFS or the LFCC diet starting at 2 months of age. Blood pressure, urinary NO metabolites (NO(x)), and total renal NO synthase activity were monitored, and the tissue abundance of nitrotyrosine (NT), which is the stable "footprint" of NO oxidation by ROS, was determined. The HFS diet group exhibited a gradual rise in arterial blood pressure and were hypertensive by 18 months. This trend was accompanied by a marked accumulation of NT in all tested tissues, an initial rise and a subsequent fall in NO synthase activity, and a fall in urinary NO(x) excretion. The HFS diet-fed animals had a blunted blood pressure response to the NO synthase inhibitor N:(omega)-nitro-L-arginine methyl ester (L-NAME) compared with the LFCC diet group, which showed a marked hypertensive response to L-NAME. L-NAME-induced HTN was reversible with L-arginine in the LFCC diet group; however, HTN was not corrected by L-arginine supplementation in the HFS diet group. These findings point to enhanced ROS-mediated inactivation and sequestration of NO, which may contribute to the reduction of bioactive NO and HTN in the HFS diet-fed animals.

Topics: Animals; Arginine; Biomarkers; Blood Pressure; Body Weight; Dietary Carbohydrates; Dietary Fats; Enzyme Inhibitors; Female; Hypertension; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Oxidation-Reduction; Oxidative Stress; Rats; Rats, Inbred F344; Reactive Oxygen Species; Tyrosine

Inflammatory bowel disease is characterized by oxidative and nitrosative stress, leukocyte infiltration, up-regulation of the expression of intercellular adhesion molecule-1 (ICAM-1), and up-regulation of P-selectin in the colon. Here we investigate the effects of the tyrosine kinase inhibitor, Tyrphostin AG 126, in rats subjected to experimental colitis. Colitis was induced in rats by intracolonic instillation of dinitrobenzene sulfonic acid (DNBS). Rats experienced hemorrhagic diarrhea and weight loss. Four days after administration of DNBS, the mucosa of the colon exhibited large areas of necrosis. Neutrophil infiltration (determined by histology as well as an increase in myeloperoxidase activity in the mucosa) was associated with up-regulation of ICAM-1 and P-selectin, as well as high tissue levels of malondialdehyde. Immunohistochemistry for nitrotyrosine and poly(ADP-ribose) polymerase showed an intense staining in the inflamed colon. Staining with an anti-COX-2 antibody of sections of colon obtained from DNBS-treated rats showed a diffuse staining of the inflamed tissue. Furthermore, expression of inducible nitric oxide synthase was found mainly in macrophages located within the inflamed colon of DNBS-treated rats. Tyrphostin AG 126 (5 mg/kg daily ip) significantly reduced the degree of hemorrhagic diarrhea and weight loss caused by administration of DNBS. Tyrphostin AG 126 also caused a substantial reduction of (1) the phosphorylation of tyrosine residues of proteins (immunoblots of inflamed colon), (2) the degree of colonic injury, (3) the rise in myeloperoxidase activity (mucosa), (4) the increase in the tissue levels of malondialdehyde, (5) the increase in staining (immunohistochemistry) for nitrotyrosine and poly(ADP-ribose) polymerase, as well as (6) the up-regulation of ICAM-1 and P-selectin caused by DNBS in the colon. Thus, we provide the first evidence that the tyrosine kinase inhibitor Tyrphostin AG126 reduces the degree of colitis caused by DNBS.

Topics: Animals; Body Weight; Colitis; Cyclooxygenase 2; Cytokines; Enzyme Inhibitors; Isoenzymes; Lipid Peroxidation; Male; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Peroxidase; Prostaglandin-Endoperoxide Synthases; Protein-Tyrosine Kinases; Rats; Rats, Sprague-Dawley; Tyrosine; Tyrphostins

Inflammatory bowel disease is characterized by oxidative and nitrosative stress, leukocyte infiltration, and up-regulation of intercellular adhesion molecule 1 (ICAM-1) expression in the colon. The aim of the present study was to examine the effects of tempol, a membrane-permeable radical scavenger, in rats subjected to experimental colitis. Colitis was induced in rats by intracolonic instillation of dinitrobenzene sulfonic acid. Rats experienced bloody diarrhea and significant loss of body weight. At 4 days after the administration of dinitrobenzene sulfonic acid, the colon injury comprised of large areas of mucosal necrosis. Neutrophil infiltration (measured as increase in myeloperoxidase activity in the mucosa) was associated with up-regulation of ICAM-1 and expression of P-selectin and high levels of malondialdehyde (an indicator of lipid peroxidation). Immunohistochemistry for nitrotyrosine and poly (ADP-ribose) synthetase showed an intense staining in the inflamed colon. Treatment of rats with tempol (15 mg/kg daily i.p.) significantly reduced the appearance of diarrhea and the loss in body weight. This was associated with a remarkable amelioration of the disruption of the colonic architecture as well as a significant reduction in the degree of both neutrophil infiltration and lipid peroxidation in the inflamed colon. Tempol also reduced the appearance of nitrotyrosine and poly (ADP-ribose) synthetase immunoreactivity in the colon as well as the up-regulation of ICAM-1 and P-selectin. The results of this study suggest that membrane-permeable radical scavengers, such as tempol, exert beneficial effects in experimental colitis and may, hence, be useful in the treatment of inflammatory bowel disease.

Topics: Animals; Benzenesulfonates; Body Weight; Cell Membrane Permeability; Colitis; Colon; Cyclic N-Oxides; Free Radical Scavengers; Immunohistochemistry; Intestinal Mucosa; Lipid Peroxidation; Male; Neutrophil Infiltration; Organ Size; Peroxidase; Poly(ADP-ribose) Polymerases; Rats; Rats, Sprague-Dawley; Spin Labels; Spleen; Survival Rate; Tyrosine

1. The aim of the present study was to investigate the effects of cloricromene, a coumarine derivative, in rats subjected to collagen-induced arthritis. 2. Collagen-induced arthritis (CIA) was induced in Lewis rats by an intradermal injection of 100 microl of the emulsion (containing 100 microg of bovine type II collagen) (CII) and complete Freund's adjuvant (CFA) at the base of the tail. On day 21, a second injection of CII in CFA was administered. 3. Lewis rats developed an erosive hind paw arthritis when immunized with CII in CFA. Macroscopic clinical evidence of CIA first appeared as peri-articular erythema and oedema in the hind paws. The incidence of CIA was 100% by day 27 in the CII challenged rats and the severity of CIA progressed over a 35-day period with radiographic evaluation revealing focal resorption of bone together with osteophyte formation in the tibiotarsal joint and soft tissue swelling. 4. The histopathology of CIA included erosion of the cartilage at the joint margins. Treatment of rats with cloricromene (10 mg kg(-1) i.p. daily) starting at the onset of arthritis (day 23), delayed the development of the clinical signs at days 24 - 35 and improved histological status in the knee and paw. 5. Immunohistochemical analysis for iNOS, COX-2, nitrotyrosine and for poly (ADP-ribose) synthetase (PARS) revealed a positive staining in inflamed joints from collagen-treated rats. The degree of staining for iNOS, COX-2, nitrotyrosine and PARS were markedly reduced in tissue sections obtained from collagen-treated rats, which had received cloricromene. 6. Radiographic signs of protection against bone resorption and osteophyte formation were present in the joints of cloricromene-treated rat. 7. This study provides the first evidence that cloricromene, a coumarine derivative, attenuates the degree of chronic inflammation and tissue damage associated with collagen-induced arthritis in the rat.

Topics: Animals; Arthritis; Body Weight; Cattle; Chromonar; Collagen; Cyclooxygenase 2; Disease Progression; Enzyme Activation; Hindlimb; Interleukin-1; Isoenzymes; Male; Malondialdehyde; Nitric Oxide; Poly(ADP-ribose) Polymerases; Prostaglandin-Endoperoxide Synthases; Radiography; Rats; Rats, Inbred Lew; Severity of Illness Index; Time Factors; Tumor Necrosis Factor-alpha; Tyrosine

Vascular aging is mainly characterized by endothelial dysfunction. We found decreased free nitric oxide (NO) levels in aged rat aortas, in conjunction with a sevenfold higher expression and activity of endothelial NO synthase (eNOS). This is shown to be a consequence of age-associated enhanced superoxide (.O(2)(-)) production with concomitant quenching of NO by the formation of peroxynitrite leading to nitrotyrosilation of mitochondrial manganese superoxide dismutase (MnSOD), a molecular footprint of increased peroxynitrite levels, which also increased with age. Thus, vascular aging appears to be initiated by augmented.O(2)(-) release, trapping of vasorelaxant NO, and subsequent peroxynitrite formation, followed by the nitration and inhibition of MnSOD. Increased eNOS expression and activity is a compensatory, but eventually futile, mechanism to counter regulate the loss of NO. The ultrastructural distribution of 3-nitrotyrosyl suggests that mitochondrial dysfunction plays a major role in the vascular aging process.

Topics: Acetylcholine; Aging; Animals; Aorta; Body Weight; Calcimycin; Cellular Senescence; Endothelium, Vascular; Enzyme Induction; Hemodynamics; Male; Microscopy, Immunoelectron; Mitochondria; Nitrates; Nitric Oxide; Nitric Oxide Synthase; Nitroprusside; Oxidative Stress; Rats; Rats, Inbred Strains; Superoxide Dismutase; Superoxides; Tyrosine; Vasodilation

The effect of mercaptoethylguanidine (MEG), a selective inhibitor of the inducible nitric oxide synthase and peroxynitrite scavenger, was evaluated in a rat model of colonic injury. A single intracolonic administration of trinitrobenzene sulfonic acid (TNBS, 20 mg/kg) dissolved in ethanol induced a severe colitis in male rats. Rats experienced bloody diarrhea and a significant loss of body weight. At 4 days after TNBS administration, the colon damage was characterized by areas of mucosal necrosis. Activity of myeloperoxidase, a marker of neutrophil infiltration, and levels of the 6-keto-prostaglandin F1alpha, were also markedly increased, whereas colonic ATP levels were reduced into the damaged tissue. Immunohistochemistry for the inducible nitric oxide synthase and nitrotyrosine, an index of nitrosative stress, showed an intense staining in the inflamed colon. Treatment with MEG (10 mg/kg i.v. b. i.d.) significantly reduced the appearance of diarrhea and the loss of body weight. This was associated with a remarkable amelioration of the disruption of the colonic architecture and suppression of the energetic failure, as well as a significant reduction of colonic myeloperoxidase activity and 6-keto-prostaglandin F1alpha levels. MEG also reduced the appearance of iNOS and nitrotyrosine immunoreactivity in the colon. The results of this study suggested that administration of MEG may be beneficial for the treatment of inflammatory bowel diseases.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Body Weight; Colitis; Enzyme Inhibitors; Free Radical Scavengers; Guanidines; Immunohistochemistry; Male; Nitrates; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Rats; Rats, Wistar; Trinitrobenzenesulfonic Acid; Tyrosine