Reactive Oxygen Species In Rats Research Articles

A New Approach in the Treatment of Traumatic Brain Injury: The Effects of Levosimendan on Necrosis, Apoptosis, and Oxidative Stress

Traumatic brain injury (TBI) is an essential and common health problem worldwide. Levosimendan is an inotropic and vasodilator drug used to treat heart failure. Moreover, it exerts pleiotropic effects and, thus, protective effects on many organs. The present study aimed to investigate the effect of levosimendan on necrosis, apoptosis, and reactive oxygen species in rats with TBI. The study included 28 female Wistar-Albino rats weighing 200-250g. The rats were divided into 4 groups with 7 rats each as follows: Group 1: No trauma group (Control), Group 2: Traumatized, untreated group (T), Group 3: Levosimendan was administered at a dose of 12μg/kg intraperitoneally 1hour after the trauma (L1), Group 4: Levosimendan was administered at a dose of 12μg/kg intraperitoneally 2hours after the concussion (L2). After the experiment, the rats were decapitated, and the brain tissue was removed. Necrosis was assessed with Cresyl violet staining, apoptosis was assessed with immunohistochemical analysis, superoxide dismutase and catalase levels were measured with the spectrophotometric method, and malondialdehyde (MDA) levels were assessed by High-Performance Liquid Chromatography. The number of necrotic cells in the L1 and L2 groups was significantly lower than in the K and T groups (P= 0.015 and P= 0.03, respectively). Although the active caspase-3 level was signified considerably in the T, L1, and L2 groups compared to the K group, no significant difference was found among these 3 groups (P > 0.05). The results of superoxide dismutase levels were similar to those of active caspase-3. catalase level was significantly higher in the K group than in the T and L2 groups (P= 0.045). Malondialdehyde activity was considerably higher in the L1 and L2 groups compared to the K group (P= 0.023). Our results indicated that levosimendan may exert a neuroprotective effect by reducing necrosis in TBI and that levosimendan does not affect apoptosis and antioxidant levels in TBI. Comprehensive studies are needed to elucidate the effect of levosimendan on TBI fully.

Comparison of toxic effects of lead and copper and protective power of glutathione on oxidative stress parameters

IntroductionLead as an industrial pollutant can be detected at all stages of the working and living environment. Lead, based on its properties, solubility and mobility, accumulates in the soil, so that the average concentrations of oil in the soil are between 15 and 25 mg/kg (Radojević at all., 1999). Due to increased human activity, the amount of copper in the air, soil and water has increased. Glutathione (GSH) is an essential cofactor of many enzymes, such as: formaldehyde dehydrogenase, glyoxalase, prostaglandin endoperoxide isomerase, dehydrochlorinase and others. GSH is a biological redox in the metabolism of erythrocytes, it also plays a role in the transport of amino acids. Reactive forms of oxygen cause oxidative biomolecules (lipids, proteins, DNA) (Freidovich, 1999; Massaad i Klann, 2010).ObjectivesThe aim of this research was to examine the protective role of supplements GSH in conditions of chronic intoxication with sublethal doses of lead acetate and copper II sulfate.MethodsThe preparation of biomaterials for testing and making homogenates of brain tissue of albino rats of Wistar strain was performed and the activity of acid and alkaline DNase was measured spectrophotometrically (Kocić i sar., 1998).ResultsLead otherwise “as soft Lewis acid” has a pronounced affinity for interaction with “soft bases” such as S-atoms of the thiol group in antioxidants, natural biomolecules and supplements in this case in glutathione.ConclusionsIt can be said that GSH is a desirable supplement and antioxidant in the detoxification of reactive oxygen species in rats exposed to lead poisoning.DisclosureNo significant relationships.

MiR-133b inhibits myocardial ischemia-reperfusion-induced cardiomyocyte apoptosis and accumulation of reactive oxygen species in rats by targeting YES1

To investigate the effect of miR-133b on cardiomyocyte apoptosis induced by myocardial ischemia-reperfusion (I/R) and explore the mechanism. Thirty-six adult SD rats were randomized into sham-operated group, I/R group, AdmiR-NC group and AdmiR-133b group, and rat models of myocardial I/R were established in the latter 3 groups with myocardial injections of saline or recombinant adenoviruses in the left ventricle. The expression of MiR-133b was detected using RT-qPCR, and cardiac function of the rats was determined using FDP 1 HRV and BRS analysis system. Serum CK-MB and cTnI levels were determined by ELISA, myocardial injury was evaluated with HE staining, cardiomocyte apoptosis was detected by flow cytometry, and ROS content was determined using a DCFH-DA probe. In the in vitro experiment, H9C2 myocardial cells with hypoxia/reoxygenation (H/R) treatment were transfected with Mir-NC or MiR-133b mimic, and the cellular expression of MiR-133b, cell apoptosis, and ROS content were determined. Dual luciferase reporter assay was performed to verify the targeting relationship between miR-133b and YES1. The effects of pc-YES1 or miR-133b mimic transfection on YES1 expression, apoptosis, and ROS content in H9C2 cells were evaluated. Compared with those in I/R group, miR-133b expression was obviously up-regulated, LVEDP, cTnI and CK-MB levels were significantly decreased, and LVSP, +dp/dt, -dp/dt, HR and CF levels were increased in admiR-133b group (P < 0.01). The rats in admiR-133b group showed obviously reduced pathological damage, cell apoptosis and ROS content compared with those in I/ R group (P < 0.01). In H9C2 cells exposed to H/R, transfection with miR-133b mimic significantly up-regulated miR-133b expression and decreased cell apoptosis and ROS content (P < 0.01). The results of dual luciferase reporter assay suggested a direct targeting relationship between miR-133b and YES1, and MiR-133b mimic transfection significantly down-regulated YES1 protein expression in cells with H/R exposure (P < 0.01). Co-transfection with pc-YES1 reversed the effect of miR-133b overexpression on myocardial cell apoptosis and ROS accumulation. miR-133b can inhibit I/R-induced myocardial cell apoptosis and ROS accumulation by targeting YES1 to reduce myocardial I/R injury in rats.

Effect of benazepril on nuclear factor E2 related factor 2, nicotinamide adenine dinucleotide phosphate oxidase and reactive oxygen species in rats with hepatic fibrosis

Objective: To study the effect of benazepril on the expression of nuclear factor E2 related factor 2 (Nrf2), nicotinamide adenine dinucleotide phosphate oxidase 4 (NOX4) and reactive oxygen species (ROS) concentration in rats with hepatic fibrosis and to explore the possible antifibrotic mechanism of benazepril. Methods: Twenty-two healthy male Sprague-Dawley rats were randomly divided into 3 groups: control group (6 rats), model group (8 rats) and benazepril treatment group (8 rats). Two rats died during modeling and treatment in the model group and the benazepril treatment group, and a model of hepatic fibrosis induced by carbon tetrachloride (CCL(4)) was established. The rats in benazepril group were given benazepril for 8 weeks by gastric gavage. The assessment of liver tissue damage in each group was measured using conventional hematoxylin-eosin and Masson staining. The mRNA level of Nrf2, NOX4 in liver tissue was detected by RT-PCR, and serum ROS concentration was determined by colorimetry. All data were expressed in mean ± standard deviations, and were analyzed using SPSS21.0 statistical software. The data were compared using one-way analysis of variance, and the LSD-t method was used for pairwise comparison between the two groups. The correlation analysis was performed by Spearman's correlation analysis. Results: In the liver of the model group, with the aggravation of liver fibrosis the expression of Nrf2mRNA, NOX4 mRNA and ROS concentration were higher than control group [(4.01 ± 3.40), (31.78 ± 3.96), (1.82 ± 0.46) μg/ ml vs. (0.12 ± 0.11), (2.03 ± 0.31), (1.56±0.84) μg/ml, P < 0.05]. After benazepril treatment, NOX4 mRNA expression and ROS concentration were decreased than the model group [(15.93 ± 5.01), (0.78 ± 0.44) μg/ml vs. (31.78 ± 3.96), (1.82 ± 0.46) μg /ml, P < 0.05], while Nrf2 mRNA expression was higher than the model group [(6.69 ± 4.86) vs. (4.01 ± 3.40), P < 0.05]. There was a positive correlation between Nrf2 and NOX4, Nrf2 and ROS, and NOX4 and ROS (r = 0.616, 0.411, 0.802, P < 0.05). Conclusion: Benazepril may exert an anti-hepatic fibrosis effect by activating Nrf2 expression, or may inhibit the ROS-mediated oxidative stress in response to NOX4.

Impact of Superoxide Dismutase Mimetic AEOL 10150 on the Endothelin System of Fischer 344 Rats.

Endothelin-1 is a potent vasoconstrictor and mitogenic peptide involved in the regulation of vasomotor tone and maintenance of blood pressure. Oxidative stress activates the endothelin system, and is implicated in pulmonary and cardiovascular diseases including hypertension, congestive heart failure, and atherosclerosis. Superoxide dismutase mimetics designed with the aim of treating diseases that involve reactive oxygen species in their pathophysiology may exert a hypotensive effect, but effects on the endothelin system are unknown. Our objective was to determine the effect of the superoxide dismutase mimetic AEOL 10150 on the basal endothelin system in vivo. Male Fischer-344 rats were injected subcutaneously with 0, 2 or 5 mg/kg body weight of AEOL 10150 in saline. Plasma oxidative stress markers and endothelins (bigET-1, ET-1, ET-2, ET-3) as well as lung and heart endothelin/nitric oxide system gene expressions were measured using HPLC-Coularray, HPLC-Fluorescence and RT-PCR respectively. AEOL 10150 reduced (p<0.05) the circulating levels of isoprostane (-25%) and 3-nitrotyrosine (-50%) measured in plasma 2h and 24h after treatment, confirming delivery of a physiologically-relevant dose and the potent antioxidant activity of the drug. The reduction in markers of oxidative stress coincided with sustained 24h decrease (p<0.05) of plasma levels of ET-1 (-50%) and ET-3 (-10%). Expression of preproET-1 and endothelin converting enzyme-1 mRNA were not altered significantly in the lungs. However preproET-1 (not significant) and ECE-1 mRNA (p<0.05) were increased (10–25%) in the heart. Changes in the lungs included decrease (p<0.05) of mRNA for the ET-1 clearance receptor ETB and the vasoconstriction-signaling ETA receptor (-30%), and an early surge of inducible nitric oxide synthase expression followed by sustained decrease (-40% after 24 hours). The results indicate that interception of the endogenous physiological flux of reactive nitrogen species and reactive oxygen species in rats impacts the endothelin/nitric oxide system, supporting a homeostatic relationship between those systems.

Topical nutraceutical Optixcare EH ameliorates experimental ocular oxidative stress in rats.

Based on the hypothesis that oral nutraceuticals do not adequately reach all ocular tissues in the anterior segment, we evaluated the ability of a 3% concentration of the ingredients in a topical nutraceutical antioxidant formulation called Optixcare Eye Health (Optixcare EH) to ameliorate oxidative stress in rat models of age-related ocular diseases. Diabetes was induced by tail-vein injection of streptozotocin, and the development of cataracts was monitored by slit lamp. Young rats were exposed to ultraviolet (UV) light, and the reduction in lens glutathione (GSH) levels and increase in 4-hydroxynonenol (4-HNE) were measured. Oxidative stress in the neural retina was generated by exposure of dark-adapted rats to 1,000 lx of light, and oxidative stress markers were measured. Dry eye was induced in rats by twice daily (b.i.d.) subcutaneous scopolamine injections. Topical Optixcare EH was administered b.i.d. and compared in select experiments to the multifunctional antioxidant JHX-4, the topical aldose reductase inhibitor (ARI) Kinostat™, oral Ocu-GLO™, and the topical ocular comfort agents Optixcare Eye Lube, Optixcare Eye Lube + Hyaluron, and Idrop Vet Plus hyaluronic acid. In diabetic rats, topical ARI treatment prevented cataract formation while the nutraceuticals delayed their development with Optixcare EH>Ocu-GLO. In UV-exposed rats, the reduction of GSH and increase in 4-HNE in the lens were normalized in order JHX-4>Optixcare EH>Ocu-GLO. In the retina, oxidative stress markers were reduced better by oral JHX-4 compared with topical Optixcare EH. In the scopolamine-induced dry-eye rats, tear flow was maintained by Optixcare EH treatment, while none of the comfort agents examined altered tear flow. Topical administration of a 3% concentration of the ingredients in Optixcare EH reduces experimentally induced reactive oxygen species in rats exposed to several sources of ocular oxidative stress. In addition, Optixcare EH maintains tear volume in scopolamine-induced dry eye. This suggests that in the anterior segment, the ingredients in Optixcare EH may have clinical potential against ocular oxidative stress.

Role of TRPV1 and P2X receptors in the activation of lung vagal C-fiber afferents by inhaled cigarette smoke in rats

Inhaled cigarette smoke (CS) triggers airway reflexes that are thought to result from the activation of lung vagal C-fiber afferents (LVCAs) via the action of reactive oxygen species in rats. We investigated the role of transient receptor potential vanilloid 1 (TRPV1) and P2X receptors in LVCA activation. Activities of LVCAs were recorded in anesthetized and artificially ventilated rats. Airway challenge of CS produced a concentration-dependent fiber stimulation. Pretreatment with dimethylthiourea [DMTU; a scavenger of hydroxyl radical (OH)], capsazepine (CPZ; a TRPV1 receptor antagonist) and iso-pyridoxalphosphate-6-azophenyl-2',5'-disulphonate (iso-PPADS; a P2X receptor antagonist) separately reduced the fiber responses by 64, 40 and 44%, respectively, whereas pretreatment with hexamethonium (a nicotinic acetylcholine receptor antagonist) failed to alter the response. A combination of CPZ and iso-PPADS exerted a greater inhibitory effect compared with the effect of either single pretreatment. However, a combination of DMTU, CPZ and iso-PPADS did not further reduce the fiber response compared with the combined effect of CPZ and iso-PPADS. It was concluded that both TRPV1 and P2X receptors, but not nicotinic acetylcholine receptors, participate in the stimulation of LVCAs by inhaled CS, possibly through the action of OH.

Exercise training provides cardioprotection via a reduction in reactive oxygen species in rats submitted to myocardial infarction induced by isoproterenol

Exercise training has demonstrated cardioprotection effects. However, the exact mechanism behind this effect is not is clear. The present study evaluated the effects of 12 weeks of previous treadmill training on the levels of oxidative damage, antioxidant enzyme activity and injury in the myocardium of rats submitted to infarction induced by isoproterenol (ISO). Isoproterenol treatment (80 mg/kg given over 2 days in two equal doses) caused arrhythmias and 60% mortality within 24 h of the last injection in the control group (C + ISO) group when compared with the saline control group (saline). Creatine Kinase − MB levels were markedly increased in hearts from ISO-treated animals in the C + ISO group. Twelve weeks of treadmill training reduced superoxide production, lipid peroxidation levels and protein carbonylation in these animals, as well as increasing the activities and expressions of SOD and CAT. Previous training also reduced CK-MB levels and numbers of deaths by 40%, preventing the deleterious effects of ISO. Based on the data obtained in this study, it is suggested that 12-week treadmill training increases antioxidant enzymes, decreases oxidative damage and reduces the degree of infarction induced by ISO in the hearts of male rats.

Oxidative stress in a rat model of nephrosis can be quantified by electron spin resonance.

The pathogenesis of nephrotic syndrome is not clear. In this study, we used electron spin resonance (ESR) to evaluate levels of reactive oxygen species in rats with puromycin aminonucleoside (PAN)-induced nephrosis. Twenty-six Wistar rats were divided into four groups: (1) PAN treated, (2) PAN treated and alpha-tocopherol supplemented, (3) supplemented with alpha-tocopherol only, (4) control. On day 9, urinary protein excretion was measured. On day 10, all animals were sacrificed with retrograde perfusion via the aorta to obtain renal venous perfusates. The signal intensities of ascorbate radicals in the perfusates were determined by ESR. After perfusion, the kidneys were isolated and sieved to obtain glomeruli for determination of glomerular thiobarbituric acid-reactive substance (TBArs) and alpha-tocopherol. Urinary protein excretion by PAN-treated rats increased significantly on day 9 and was reduced by alpha-tocopherol supplementation. The ascorbate radical intensity and glomerular TBArs level were higher in PAN-treated than in control rats and were both suppressed to control levels by alpha-tocopherol supplementation. There were positive correlations between ascorbate radical intensity and the daily urinary protein, as well as between ascorbate radical intensity and the glomerular TBArs level. Hence, it is possible to quantify oxidative stress due to PAN nephrosis by ESR. Our findings suggest that lipid peroxidation plays an important role in the pathogenesis of proteinuria in PAN-treated rats.

Xanthine Oxido-reductase Activity in Ischemic Human and Rat Intestine

We measured time course and extent of xanthine dehydrogenase (XD) to xanthine oxidase (XO) conversion in ischemic human and rat intestine. To model normothermic no-flow ischemia, we incubated fresh biopsies for 0, 2, 4, 8 and 16 h. At [Formula: See Text] XO was less in humans than in rats [Formula: See Text] while XD was essentially the same [Formula: See Text] After 16 h incubation at 37°C, there was no appreciable XD-to-XO conversion and no change in neither XO nor XD activity in human intestine. In contrast, the rat intestine had [Formula: See Text] ratio doubled in the first 2 h and then maintained that value until [Formula: See Text] In conclusion, no XO-to-XD conversion was appreciable after 16 h no-flow normothermic ischemia in human intestine; in contrast, XO activity in rats increased sharply after the onset of ischemia. An immunohistochemical labelling study shows that, whereas [Formula: See Text] expression in liver tissue is localised in both hepatocytes and endothelial cells, in the intestine that expression is mostly localised in epithelial cells. We conclude that XO may be considered as a major source of reactive oxygen species in rats but not in humans.

4680 Reversibility of kupffer cells dysfunction in rats with obstructive jaundic: phagocytic function and production of reactive oxygen species evaluated by flow cytometry.

Background: The function of the reticuloendothelial system is impaired in obstructive jaundice. The effects of obstructive jaundice on Kupffer cells and the influence of internal and external drainage are unclear. Aims: To study the effect of internal and external drainage on Kupffer cell phagocytosis and production of reactive oxygen species in rats with obstructive jaundice. Materials and Methods: Fifty male Sprague-Dawley rats weighing 300-350 g were randomly assigned to 4 groups: sham operation (SH, n = 11), obstructive jaundice by bile duct ligation and division (OJ, n = 12), internal drainage by choledochoduodenostomy (ID, n = 13) and external drainage by exteriorizing a biliary drainage tube at the nape (ED, n = 14) on day 7 after bile duct ligation. Kupffer cells were isolated by flow cytometric sorting 7 days later. The phagocytic function of the Kupffer cells was measured by flow cytometry using fluorescent microspheres (1.72 μm). The production of reactive oxygen species (ROS) was assayed by flow cytometry using dihydrorhodamine 123 (DHR) with phorbol myristate acetate (PMA) as an activator. Results: The phagocytic function was suppressed in OJ rats (20.8±7.4%) compared with SH rats (41.3±23.2%) (P=0.007). After drainage procedures, the depressed Kupffer cell function was reversed by ID (37.0±15.2%) (P=0.036), but not by ED (21.6±7.0%) (P=1.000). The release of ROS was higher in the OJ rats (1.35±0.36) than in the SH rats (0.98±0.09) (P=0.042). After drainage, the ROS production returned to normal levels in the ID rats (0.97±0.11) (P=0.032), but remained high in the ED rats (1.35±0.46) (P=1.000). Conclusion: Kupffer cell functions, including phagocytosis and release of reactive oxygen species, are impaired in obstructive jaundice. Internal biliary drainage, but not external drainage, causes recovery of Kupffer cell functions.

Effect of dietary antioxidants on puromycin aminonucleoside nephrotic syndrome

Several studies indicate the pathophysiological importance of reactive oxygen species in rats with nephrotic syndrome induced by puromycin aminonucleoside, an experimental model of the human minimal change disease. The role of reactive oxygen species in these rats was further evaluated, examining the effect of dietary deficiency and supplementation of antioxidants (vitamin E and selenium) on biochemical and renal ultrastructural alterations induced by puromycin aminonucleoside. Male Wistar rats, weaned at 3 weeks, were placed on diets normal, deficient or supplemented in vitamin E and selenium for 4 weeks. At the end of this period, rats were divided in two groups: control (sacrificed without any further treatment) and nephrotic (injected with puromycin aminonucleoside and sacrificed 7 and 22 days later). In control rats, the dietary deficiency or supplementation of antioxidants resulted in no significative differences in renal function, proteinuria or kidney ultrastructure. However, kidney lipoperoxidation, kidney glutathione peroxidase activity and circulating levels of vitamin E changed according to the amount of antioxidants in the diet. Seven days after the injection of puromycin aminonucleoside, rats fed normal, deficient or supplemented diets, developed nephrotic syndrome. However, proteinuria, hypoproteinemia, renal dysfunction and ultrastructural alterations were higher in rats fed a deficient diet. In contrast, proteinuria and kidney ultrastructural alterations were lower in rats fed a supplemented diet. Kidney lipoperoxidation and glutathione peroxidase activity increased on day 7 in rats fed a normal or a deficient diet, but not in rats fed a supplemented diet. This study shows that nephrotic syndrome induced by puromycin aminonucleoside in rats is modified by dietary antioxidants (vitamin E and selenium). Dietary supplementation ameliorates it and dietary deficiency exacerbates it. It is concluded that reactive oxygen species are involved in the pathophysiology of this experimental disease.