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Background/Aim. Exposure of living organisms to γ- radiation results in the overproduction of free radicals. The aim of the study was to test if the subacute administration of micronized zeolite (MZC) accomplishes radioprotective role based on the evaluation of the status of oxidative stress (OS) in the brain and 8-hydroxyguanosine (8-OH-dG) in the plasma of the rats exposed to the single γ-ray irradiation of 2 and/or 10 Gray (Gy). Methods. Wistar rats were on a four-week normal or 5% MZC supplemented diet and af- terward exposed to the single γ-ray irradiation of 2 and 10 Gy. Groups of rats were: a) on a normal diet (the control group, and 2Gy and 10Gy groups); b) on 5% MZC supple- mented diet (the control group – MZC, MZC + 2Gy, and MZC + 10Gy groups). We measured malondialdehyde (MDA), glutathione (GSH) total, and activity of total super- oxide dismutase (tSOD) and manganese superoxide dis- mutase (MnSOD) in vulnerable brain regions (cerebellum, hippocampus, and cerebral cortex) and 8-OH-dG in plasma. Results. Lower MDA was found in the MZC+2Gy and MZC+10Gy groups compared to the 2Gy and 10Gy groups. Activity od total SOD was higher in the MZC+10Gy group than in the 10Gy one. GSH was the highest in the 10Gy group. Compared to the control group, 8-OH-dG was extremely higher in groups radiated with 10 Gy regardless of a diet, but slightly lower in the MZC+2Gy and 2Gy groups. Conclusion. Subacute MZC pretreatment accomplished partial radioprotective effect in irradiated rats compared to non-irradiated rats, based on suppressed SOD activity at 2 Gy, and reduced brain MDA when exposed to 2 Gy and 10 Gy. Uvod/Cilj. Izlaganje živih organizama gama zračenju re- zultira hiperprodukcijom slobodnih radikala. Cilj istraživanja je bio da se ispita da li subakutna ishrana dopunjena sa 5% mikronizovanog zeolita (MZC) ispoljava radiozaštitnu ulogu na osnovu statusa oksidativnog stresa (OS) u mozgu i 8- hidroksiguanozina (8-OH-dG) u plazmi pacova izloženih pojedinačnim dozama jonizujućeg zračenja od 2 i 10 Gray (Gy). Metode. Wistar pacovi su bili na četvoronedeljnoj normalnoj ishrani ili ishrani obogaćenoj sa 5% MZC, posle čega su bili izloženi pojedinačnom jonizujućem zračenju od 2 Gy, odnosno 10 Gy. Grupe pacova bile su: a) gru pa pacova na normalnoj ishrani (kontrolna grupa i grupe 2Gy i 10Gy); b) grupa pacova na ishrani obogaćenoj sa 5% MZC (kontrolna grupa – MZC i grupe MZC+2Gy i MZC+10Gy). Meren je malondialdehid (MDA), glutation (GSH) i aktivnost ukupne (tSOD) i mangan superoksid dizmutaze (MnSOD) u osetljivim strukturama mozga (cerebelum, hipokampus i cerebralni korteks), a 8-OH-dG u plazmi. Rezultati. Biomarker MDA je bio niži u MZC+2Gy i MZC+10Gy grupama, u odnosu na grupe 2Gy i 10Gy. Aktivnost ukupne SOD je bila viša u grupi MZC+10Gy, u odnosu na grupu 10Gy. Najviši nivo GSH bio je u grupi 10Gy. U pređenju sa kontrolnom grupom, 8- OH-dG je bio izuzetno viši u grupama ozračenim sa 10 Gy, bez obzira na dijetetski režim i niži u grupama MZC+2Gy i 2Gy. Zaključak. Pacovi koji su bili na režimu ishrane obogaćene sa 5% MZC bili su delimično zaštićeni od zračenja, shodno redukovanoj moždanoj aktivnosti SOD pri 2 Gy i sniženom nivou MDA pri izlaganju zračenju od 2 i 10 Gy.

The use of the antidepressant drug chlorpromazine (CPZ) is linked to the occurrence of oxidative stress in some brain structures. Thus, overcoming the side effects of CPZ is of great importance. Because agmatine (AGM) can act as a free radical scavenger, it is an interesting compound as an adjunct to CPZ therapy. The aim of our study was to investigate the enzymatic parameters of oxidative stress in the hippocampus and striatum of rats after CPZ treatment, and the potential protective effects of AGM. Rats were injected as follows with (i) 1 mL/kg b.w. saline; (ii) a single intraperitoneal (i.p.) dose of CPZ (38.7 mg/kg); (iii) CPZ (38.7 mg/kg) and AGM (75 mg/kg); (iv) AGM (75 mg/kg). CPZ induced an increase in superoxide anion radical (O2 •-) concentration, while the activities of the antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glutathione reductase (GR), were lowered in both the hippocampus and striatum. Cotreatment with CPZ and AGM protected the examined brain structures by reversing the antioxidant enzyme activities to the control values. Following CPZ treatment, the effects were more pronounced for SOD and GPx in the hippocampus, and for SOD, CAT and GPx in the striatum. The full effect of restored superoxide production was achieved in the striatum, which points to the role of CAT. The obtained results suggest that CPZ in combination with AGM may be considered as a new treatment strategy.

Multiple sclerosis (MS) is an inflammatory, demyelinating disease with an unknown origin. Previous studies showed the involvement of the hypothalamic–pituitary–adrenal (HPA) axis to susceptibility to autoimmune diseases, including MS, and its best-characterized animal model, experimental autoimmune encephalomyelitis (EAE). During MS/EAE, innate immune cells are activated and release cytokines and other inflammatory mediators, leading to a vicious cycle of inflammation. In response to inflammation, the activated HPA axis modulates immune responses via glucocorticoid activity. Because the mechanisms involving oxidative stress to the HPA axis are relatively unrevealed, in this study, we investigate the inflammatory and oxidative stress status of HPA axis during EAE. Our results reveal an upregulation of Pomc gene expression, followed by POMC and ACTH protein increase at the peak of the EAE in the pituitary. Also, prostaglandins are well-known contributors of HPA axis activation, which increases during EAE at the periphery. The upregulated Tnf expression in the pituitary during the peak of EAE occurred. This leads to the activation of oxidative pathways, followed by upregulation of inducible NO synthase expression. The reactive oxidant/nitrosative species (ROS/RNS), such as superoxide anion and NO, increase their levels at the onset and peak of the disease in the pituitary and adrenal glands, returning to control levels at the end of EAE. The corticotrophs in the pituitary increased in number and volume at the peak of EAE that coincides with high lipid peroxidation levels. The expression of MC2R in the adrenal glands increases at the peak of EAE, where strong induction of superoxide anion and malondialdehyde (MDA), reduced total glutathione (GSH) content, and catalase activity occurred at the peak and end of EAE compared with controls. The results obtained from this study may help in understanding the mechanisms and possible pharmacological modulation in MS and demonstrate an effect of oxidative stress exposure in the HPA activation during the course of EAE.

Paper description: The effect of disulfiram (DSF) on reproductive toxicity induced by cadmium (Cd) was studied. In the testes of Wistar rats, morphological, oxidative and testosterone changes were examined upon exposure to Cd and/or DSF. Cd triggered oxidative stress, affected the morphology of the testes and decreased testosterone production. DSF decreased the oxidative stress. DSF did not change the morphology of the testes and it did not affect plasma testosterone levels. Once impaired by Cd, Leydig cells could not be reactivated by DSF. This article has been corrected. Link to the correction 10.2298/ABS200226010E Abstract: We investigated the effect of disulfiram (DSF) on reproductive toxicity induced by subchronic exposure to cadmium (Cd). We examined the redox status and systemic testosterone changes in the testes and plasma of Cd-treated male Wistar rats. Rats were treated with 1 mg CdCl 2 /kg body weight (bw)/day (intraperitoneal administration) for 42 days; in the second experimental group, rats were given 178.5 mg DSF/kg bw/day by oral gavage for 21 days; in the third group, after administration of Cd for 21 days, DSF treatment was introduced on day 22 and lasted until day 42, with continuous Cd intake. Each experimental group had a matching control: untreated rats, rats that received for 21 days olive oil, the solvent for DSF; rats that started with olive oil intake from days 22-42. Exposure of rats to DSF modulated the oxidative status in the testes; thus, coexposure increased the Cd-induced reduction in total superoxide dismutase (tSOD), catalase (CAT), glutathione reductase (GR) and total glutathione-S-transferase (tGST) activities, and lowered the Cd-increased superoxide anion radical (O 2 ●– ) and malondialdehyde (MDA) concentrations. DSF did not affect testosterone production diminished by Cd, as Leydig cells, once impaired by Cd, could not be reactivated by DSF. https://doi.org/10.2298/ABS190814057P Received: August 14, 2019 ; Revised: September 4, 2019; Accepted: September 4, 2019; Published online: September 9, 2019 How to cite this article: Pavlovic M, Đuric A, Stevanovic I, Begic A, Vujanovic D, Ninkovic M, Đukic M. Disulfiram partially improves oxidative but not androgen status in rats exposed to cadmium. Arch Biol Sci. 2019;71(4):747-53.

Dexamethasone (DEX) is frequently used to treat women at risk of preterm delivery, but although indispensable for completion of organ maturation in fetus, antenatal DEX treatment may exert adverse sex-dimorphic neurodevelopmental effects. Literature findings implicated oxidative stress in adverse effects of DEX treatment.. Purinergic signalling is involved in neurodevelopment, and controled by ectonucleotidases, among which in the brain are most abundant ectonucleoside triphosphate diphosphohydrolase 1 (NTPDase1/CD39) and ecto-5ʹ-nucleotidase (e5ʹNT/CD73), which jointly dephosphorylate ATP to adenosine. They are also involved in cell adhesion and migration, critical for brain development. Up-regulation of CD39 and CD73 after DEX treatment was reported in adult rat hippocampus. We investigated the effects of maternal DEX treatment on CD39 and CD73 expression and enzymatic activity in the rat fetal brain of both sexes, in the context of oxidative status of the brain tissue. Fetuses were obtained at embryonic day (ED) 21, from Wistar rat dams treated with 0.5 mg DEX/kg/day, at ED 16, 17, and 18, and brains were processed and used for further analysis. Sex-specific increase in CD39 and CD73 expression and in the corresponding enzyme activities was induced in the brain of antenatally DEX-treated fetuses, more prominently in males. The oxidative stress induction after antenatal DEX treatment, was confirmed in both sexes, altghough showing a slight bias in males. Due to involvement of purinergic system in crucial neurodevelopmental processes, future investigations are needed to determine the role of observed changes in the adverse effects of antenatal DEX treatment.

Cortical theta burst stimulation (TBS) structured as intermittent (iTBS) and continuous (cTBS) could prevent the progression of the experimental autoimmune encephalomyelitis (EAE). The interplay of brain antioxidant defense systems against free radicals (FRs) overproduction induced by EAE, as well as during iTBS or cTBS, have not been entirely investigated. This study aimed to examine whether oxidative-nitrogen stress (ONS) is one of the underlying pathophysiological mechanisms of EAE, which may be changed in terms of health improvement by iTBS or cTBS. Dark Agouti strain female rats were tested for the effects of EAE and TBS. The rats were randomly divided into the control group, rats specifically immunized for EAE and nonspecifically immuno-stimulated with Complete Freund&rsquo, s adjuvant. TBS or sham TBS was applied to EAE rats from 14th&ndash, 24th post-immunization day. Superoxide dismutase activity, levels of superoxide anion (O2&bull, &ndash, ), lipid peroxidation, glutathione (GSH), nicotinamide adenine dinucleotide phosphate (NADPH), and thioredoxin reductase (TrxR) activity were analyzed in rat spinal cords homogenates. The severity of EAE clinical coincided with the climax of ONS. The most critical result refers to TrxR, which immensely responded against the applied stressors of the central nervous system (CNS), including immunization and TBS. We found that the compensatory neuroprotective role of TrxR upregulation is a positive feedback mechanism that reduces the harmfulness of ONS. iTBS and cTBS both modulate the biochemical environment against ONS at a distance from the area of stimulation, alleviating symptoms of EAE. The results of our study increase the understanding of FRs&rsquo, interplay and the role of Trx/TrxR in ONS-associated neuroinflammatory diseases, such as EAE. Also, our results might help the development of new ideas for designing more effective medical treatment, combining neuropsychological with noninvasive neurostimulation&ndash, neuromodulation techniques to patients living with MS.

Biocompatibility of dental materials (DM) can be evaluated by gingival crevicular fluid (GCF) oxidative stress (OS) status. The goal of the study was to ascertain influence of dental caries degree, teeth position, and type and amount of applied DM on GCF OS profile. For this purpose, we tested six DMs that were sealed in one session: amalgam (Amg), composites: Tetric EvoCeram and Beautifil (BF), phosphate cement—zinc phosphate and polycarboxylate cements—zinc polycarboxylate cements, and glass ionomer cement (GIC). The study included 88 dental outpatients. Follow-up was scheduled at 7th and 30th day. Oxidative stress parameters (malondialdehyde (MDA) and glutathione (GSH) levels and total superoxide dismutase (tSOD) activity) were measured before (0th day) and after the treatment (7th and 30th day) in GCF. Control teeth were mirror-positioned healthy teeth. The DM accomplished the following effects (listed in descending order): increase of GSH in GCF was realized by ZPoC > BF > GIC > Amg; tSOD activity increase by ZPoC > BF > Amg; and MDA decrease by ZPoC > ZPhC > Amg > TEC. Dental caries provokes insignificant rise of OS in GCF. ZPoC and ZPhC showed the highest antioxidant effect, contrary to GIC. Restorations with antioxidant properties may reduce gum diseases initiated by caries lesion, what is of great clinical relevance in dentistry.

The aim of our work was to optimize and apply simple high-performance liquid chromatography method with ultraviolet detection (HPLC-UV) for simultaneous determination of reduced (GSH) and oxidized (GSSG) glutathione in biological matrix (specifically, the rat liver tissue was used herein), since the ratio between oxidized and reduced glutathione forms (GSSG-GSH) has been recognized as an important biological marker of oxidatively depleted GSH in oxidative stress (OS)-associated diseases and poisonings. An isocratic chromatographic separation of GSH and GSSG (2.8 min and 6.3 min, respectively) was performed with the mobile phase consisted of sodium perchlorate solution (pH adjusted to 2.8) at flow rate of 1 mL min(-1), detection set at 215 nm, and column temperature of 40 degrees C. The method offers short run time, linearity in the range of 0.01-200 mu M concentration for both compounds (R-2 = 1), low limits of detection and quantification (GSH: 0.18 mu M and 0.56 mu M, GSSG: 0.52 mu M and 1.58 mu M, respectively), precision, accuracy (bias lt 2%), and high reproducibility. Through suitable sample handling, an overestimation of GSSG was prevented. High recovery (>99%) was achieved. The method was successfully applied for the analysis of GSH and GSSG in liver homogenates of Wistar rats intraperitoneally exposed to cadmium (Cd) (1 mg kg(-1) CdCl2/21 days). Regardless of other Cd-mediated hepatotoxicity mechanisms, herein, we have exclusively interpreted/emphasized oxidative GSH depletion. The presented method is acceptable for a routine analysis of GSH and GSSG in biological matrix, while the calculated ratio GSSG-GSH is considered as a valuable OS marker.

Background/Aim. Although chlorpromazine (CPZ) is an antipsychotic drug widely used in clinical practice for a long time, its mechanism of action has not been entirely defined. An extremely difficult managing of patients acutely poisoned with CPZ is additional reason for detailed studying its toxicity mechanisms. In this clinical study, we investigated whether the oxidative stress (OS) mediates CPZ toxic effects in the exposed patients. Methods. The patients were organized into 3 groups: the T-group - hospitalized patients receiving therapeutic doses of 75-150 mg CPZ/day; the overdosed group, divided into two subgroups: the group M and the group S - mildly (CPZ serum concentration: 0.21 ± 0.05 mg/L) and severely (CPZ serum concentration: 2.66 ± 0.25 mg/L) poisoned patients, respectively, and the group C (control group of healthy volunteers). Oxidative stress parameters [total antioxidative status (TAS) and malondialdehyde (MDA) in plasma)] and superoxide dismutase (SOD) activity in erythrocytes were measured spectrophotometrically, and CPZ concentrations in serum were monitored chromatographically. One set of measurements was performed in the group C and T, whereas two sets of measurements (after 24 hours and 48 hours) were done in the poisoned patients, groups M and S. Results. A decrease of TAS and increase of SOD activity were obtained in both subgroups of the poisoned patients, compared to the controls and the group receiving therapeutic doses of CPZ. A significant increase of MDA was achieved in severely poisoned patients, compared to all other groups. Conclusion. Changed oxidative stress parameters in patients poisoned with chlorpromazine indicate involvement of oxidative stress in the toxicity mechanism(s) of chlorpromazine. [Military Medical Academy, Project No. МФВМА/6/15-17]

Overproduction of free radicals accompanied with their insufficient removal/neutralization by antioxidative defense system impairs redox hemostasis in living organisms. Oxidative stress has been shown to be involved in all the stages of carcinogenesis and malignant melanocyte transformation. The aim of this study was to examine association between oxidative stress development and different stages of melanoma.The measured oxidative stress parameters included: superoxide anion radical, total and manganese superoxide dismutase, catalase and malondialdehyde. Oxidative stress parameters were measured spectrophotometrically in serum samples from melanoma patients (n=72) and healthy control subjects (n=30). Patients were classified according to AJCC clinical stage.Average superoxide anion and malondialdehyde concentrations were significantly higher in melanoma patients than in control group, with the highest value of superoxide anion in stage III, while malondialdehyde highest value was in stage IV. The activity of total and manganese superoxide dismutase was insignificantly higher in melanoma patients than in control group, while catalase activity was significantly higher. The highest activity of total activity of manganese superoxide dismutase was in stage IV. Catalase activity was increasing with the disease progression achieving the maximum in stage III.Results of our study suggest that melanoma is oxidative stress associated disease, as well as deteriorated cell functioning at mitochondrial level.Prekomerna produkcija slobodnih radikala i njihova nedovoljna eliminacija/neutralizacija putem sistema antioksidativne odbrane, narušava redoks homeostazu u organizmu. Oksidativni stres je uključen u sve faze razvoja karcinoma i maligne transformacije melanocita. Cilj ove studije bio je da se ispita razvoj oksidativnog stresa u razlićitim stadijumima melanoma.Parametri za procenu oksidativnog stresa su: superoksil anjon radikal, ukupna i mangan superoksidna dizmutaza, katalaza i malondialdehid. Parametri oksidativnog stresa su mereni metodom spektrofotometrije u serumu bo lesnika sa melanomom (n=72) i zdravih kontrolnih oso ba (n=30). Bolesnici su klasifikovani prema AJCC kriterijumu.Prosečne koncentracije superoksil anjon radikala i malonaldehida bile su značajno veće kod bolesnika sa melanomom u odnosu na kontrolnu grupu, najveća vrednost superoksil anjon radikala bila je u III stadijumu, a malondialdehida u IV stadijumu. Aktivnost ukupne i mangan superoksidne dizmutaze bila je neznačajno povećana kod obolelih od melanoma u odnosu na kontrolnu grupu, dok je aktivnost katalaze bila statistički značajno veća. Najveća aktivnost ukupne superoksidne dizmutaze bila je u III stadijumu, a mangan superoksidne dizmutaze u IV stadijumu.Zaključak Aktivnost katalaze je rasla sa napredovanjem bolesti i dostigla maksimum u III stadijumu.Rezultati našem studije ukazuju na povezanost melanoma i oksidativnog stresa, kao i na pogoršanu funkciju ćelija na nivou mitohondrija.

This study was aimed to study the potentially beneficial effects of agmatine on oxidative/nitrosative stress development in the brain of Wistar rats during subacute chlorpromazine treatment. The animals were divided into control (0.9% saline), chlorpromazine (38.7 mg/kg b.w.), chlorpromazine+agmatine (agmatine 75 mg/kg b.w. immediately after chlorpromazine, 38.7 mg/kg b.w. i.p.) and agmatine (75 mg/kg b.w.) groups. All the tested substances were administered intraperitoneally for 15 consecutive days and the rats were sacrificed by decapitation on day 15. Subacute administration of chlorpromazine resulted in increased lipid peroxidation, nitric oxide concentration and superoxide anion production, while completely damaging the antioxidant defence system in the cerebral cortex, striatum, and hippocampus. However, the combined treatment with chlorpromazine and agmatine significantly attenuated the oxidative/nitrosative stress indices and restored the antioxidant capacity to the control values in all of the examined brain regions. Western blot analysis supported biochemical findings in all groups, but the most notable changes were found in the hippocampus. Our results suggest potentially beneficial effects of agmatine, which may be useful in the modified antioxidant approach in chlorpromazine-therapy.

The present study was directed to potentially beneficial effects of agmatine (AGM) on oxidative/nitrosative stress development in selective vulnerable brain regions during chlorpromazine (HPZ) treatment in rats. All tested compounds were administered intraperitoneally (i.p.) in one single dose. The animals were divided into control (K, 0.9 % saline solution), HPZ (HPZ, 38.7 mg/kg b.w.), HPZ+AGM (AGM, 75 mg/kg b.w. immediately after HPZ, 38.7 mg/kg b.w. i.p.) and AGM (AGM, 75 mg/kg b.w.) groups. Rats were sacrificed by decapitation 24 hours after the treatment. Analysis of data showed that HPZ+AGM injection significantly decreased drug concentration compared with HPZ-animals (p

Summary The aim of this study was to investigate the exogenous agmatine influence on nitrosative and oxidative stress parameters in acute phase of multiple sclerosis (MS) experimental model, experimental autoimmune encephalomyelitis (EAE). EAE was induced by subcutaneous injection of myelin basic protein (50 μg per animal). Sprague-Dawley rats were divided into five groups: I group - (CG), treated by PBS (i.p.), II group - (EAE), III group - (CFA), treated with Complete Freund’s adjuvant (0.2 ml subcutaneously), IV group - (EAE+AGM), treated by agmatine (75 mg/kg bw i.p.) upon EAE induction and V group - (AGM), received only agmatine in the same dose. The animals were treated every day during experiment - from day 0 to 15, and clinically scored every day. They were sacrificed on day 16 from MBP application. NO2+NO3, S-nitrosothiols (RSNO), malondyaldehide (MDA) and reduced glutathione (GSH) concentrations and superoxide dismutase (SOD) activity were determined in rat whole encephalitic mass (WEM) and cerebellum homogenates. Agmatine exerted strong protective effects on EAE clinical symptoms (p2+NO3, RSNO and MDA concentrations were increased compared to CG values. Agmatine treatment diminished NO2+NO3, RSNO and MDA levels in EAE animals (p The CNS changes in EAE are successfully supressed by agmatine application, which could be the the new aspect of the neuroprotective effects of agmatine.

Examination of cadmium (Cd) toxicity and disulfiram (DSF) effect on liver was focused on oxidative stress (OS), bioelements status, morphological and functional changes. Male Wistar rats were intraperitoneally treated with 1mg CdCl2/kg BW/day; orally with 178.5 mg DSF/kg BW/day for 1, 3, 10 and 21 days; and co-exposed from 22nd to 42nd day. The co-exposure nearly restored previously suppressed total superoxide dismutase (SOD), catalase (CAT) and increased glutathione peroxidase (GPx) activities; increased previously reduced glutathione reductase (GR) and total glutathione-S-transferase (GST) activities; reduced previously increased superoxide anion radical (O-2(center dot-)) and malondialdehyde (MDA) levels; increased zinc (Zn) and iron (Fe), and decreased copper (Cu) (yet above control value), while magnesium (Mg) was not affected; and decreased serum alanine aminotransferases (ALT) levels. Histopathological examination showed signs of inflammation process as previously demonstrated by exposure to Cd. Overall, we ascertained partial liver redox status improvement, compared with the formerly Cd-induced impact.

The aim of the study was to investigate if alcohol and disulfiram (DSF) individually and in combination affect bioelements' and red-ox homeostasis in testes of the exposed rats. The animals were divided into groups according to the duration of treatments (21 and/or 42 days): C-21/C-42 groups (controls); OL21 and OL22-42 groups (0.5 mL olive oil intake); A(1-21) groups (3 mL 20% ethanol intake); DSF1-21 groups (178.5 mg DSF/kg/day intake); and A(21)+DSF22-42 groups (the DSF ingestion followed previous 21 days' treatment with alcohol). The measured parameters in testes included metals: zinc (Zn), copper (Cu), iron (Fe), magnesium (Mg) and selenium (Se); as well as oxidative stress (OS) parameters: superoxide anion radical (O-2(center dot-)), glutathione reduced (GSH) and oxidized (GSSG), malondialdehyde (MDA), hydrogen peroxide (H2O2) decomposition and activities of total superoxide dismutase (tSOD), glutathione-Stransferase (GST) and glutathione reductase (GR). Metal status was changed in all experimental groups (Fe rose, Zn fell, while Cu increased in A(21)+DSF24-32 groups). Development of OS was demonstrated in A(1-21) groups, but not in DSF1-21 groups. In A(21)+DSF22-42 groups, OS was partially reduced compared to A groups (A(1-21)>MDA>C; A(1-21) lt GSH lt C). High metal-binding affinity of DSF/DDTC changes red-ox homeostasis in rat testes.

The present study focused on potentially beneficial effects of agmatine on oxidative stress development in the liver during chlorpromazine treatment in rats. We wanted to examine the role of reactive oxygen species and efficiency of antioxidant protection through the determination of malondylaldehyde and total glutathione concentrations in rat liver homogenate, as well as plasma concentrations of malonylaldehyde and sulfhydryl groups after the treatment. Also, liver tissue sections were examined to follow histological changes. Chlorpromazine was applied intraperitoneally at a single dose of 38.7 mg/kg b.w. The second group was treated with both chlorpromazine (at a single dose of 38.7 mg/kg b.w.) and agmatine (at a single dose of 75 mg/kg b.w.). Agmatine was applied immediately after the chlorpromazine. The control group was treated with 0.9% saline solution in the same manner. Rats were sacrificed by decapitation 24 h after the treatment and biochemical and immunohistochemical examinations were performed. Analysis of data showed that treatment with agmatine significantly attenuated the oxidative stress indicators as evidenced by lowering malonylaldehyde concentrations in the liver and in plasma while not affecting liver concentrations of total glutathione and plasma concentration of sulfhydryl groups. Additionally, histological evaluation revealed the improvement of liver damage in this respect. The presented data indicated that intraperitoneally administered agmatine protects against chlorpromazine-induced liver disease in rats. Malondialdehyde, glutathione, liver, oxidative stress, plasma

Aluminium may have an important role in the aetiology/pathogenesis/precipitation of Alzheimer's disease. Because green tea (Camellia sinensis L.) reportedly has health-promoting effects in the central nervous system, we evaluated the effects of green tea leaf extract (GTLE) on aluminium chloride (AlCl3) neurotoxicity in rats. All solutions were injected into the cornu ammonis region 1 hippocampal region. We measured the performance of active avoidance (AA) tasks, various enzyme activities and total glutathione content (TGC) in the forebrain cortex (FbC), striatum, basal forebrain (BFb), hippocampus, brain stem and cerebellum. AlCl3 markedly reduced AA performance and activities of cytochrome c oxidase (COX) and acetylcholinesterase (AChE) in all regions. It decreased TGC in the FbC, striatum, BFb, hippocampus, brain stem and cerebellum, and increased superoxide dismutase activity in the FbC, cerebellum and BFb. GTLE pretreatment completely reversed the damaging effects of AlCl3 on AA and superoxide dismutase activity, markedly corrected COX and AChE activities, and moderately improved TGC. GTLE alone increased COX and AChE activities in almost all regions. GTLE reduces AlCl3 neurotoxicity probably via antioxidative effects and improves mitochondrial and cholinergic synaptic functions through the actions of (−)-epigallocatechin gallate and (−)-epicatechin, compounds most abundantly found in GTLE. Our results suggest that green tea might be beneficial in Alzheimer's disease. Copyright © 2013 John Wiley & Sons, Ltd.

This study was conducted to investigate whether agmatine (AGM) provides protection against oxidative stress induced by treatment with chlorpromazine (CPZ) in Wistar rats. In addition, the role of reactive oxygen species and efficiency of antioxidant protection in the brain homogenates of forebrain cortexes prepared 48 h after treatment were investigated. Chlorpromazine was applied intraperitoneally (i.p.) in single dose of 38.7 mg/kg body weight (BW) The second group was treated with both CPZ and AGM (75 mg/kg BW). The control group was treated with 0.9\% saline solution in the same manner. All tested compounds were administered i.p. in a single dose. Rats were sacrificed by decapitation 48 h after treatment Treatment with AGM significantly attenuated the oxidative stress parameters and restored antioxidant capacity in the forebrain cortex. The data indicated that i.p. administered AGM exerted antioxidant action in CPZ-treated animals. Moreover, reactive astrocytes and microglia may contribute to secondary nerve-cell damage and participate in the balance of destructive vs. protective actions involved in the pathogenesis after poisoning. Ministry of Education, Science and Technological Development of the Republic of Serbia {[}III41014]; Military Medical Academy {[}MBMA/3/13-15, MBMA/6/15-17]

Summary Nitric oxide (NO) is an important signalling molecule in a variety of physiological processes. NO, a gas, is produced from L-arginine by different isoforms of the nitric oxide synthase and serves as mediator in important physiological functions, such as promoting vasodilation of blood vessels and mediating communication between nervous system cells. Contradictory to its physiologic actions, free radical activity of NO can cause cellular damage by the induction of nitrosative stress with significant implications on nervous system diseases. Although the mechanism of NOmediated neurodegeneration still remains unclear, numerous studies suggest its crucial role in modification of protein functions by nitrosylation and nitro-tyrosination. NO contributes to glutamate excitotoxicity, participates in organelle fragmentation, inhibits mitochondrial respiratory complexes and mobilizes zinc from the internal stores. Recently, NO has been emerged as a mediator of epigenetic gene expression and chromatin changes. Besides, NO is a key mediator in the regulation of inflammatory and immune response of the central nervous system. It is involved in down regulation of several aspects of CNS inflammation, but also has a dual role in that it is required for inflammation in some situations.

Experimental autoimmune encephalomyelitis (EAE) is a well-established animal model of human multiple sclerosis (MS). We have evaluated the role of oxidative and nitrosative stress, as the causal factors in the development of EAE, responsible for the damage of cardinal cellular components, such as lipids, proteins and nucleic acids, resulting in demyelination, axonal damage, and neuronal death. EAE was induced in female Sprague-Dawley rats, 3 months old (300 ± 20 g), by immunization with myelin basic protein in combination with Complete Freund's adjuvant (CFA). The animals were divided into seven groups: control, EAE, CFA, EAE + aminoguanidine (AG), AG, EAE + N-acetyl-l-cysteine (NAC) and NAC. The animals were sacrificed 15 days after EAE induction, and the levels of nitrosative and oxidative stress were determined in 10% homogenate of the whole encephalitic mass. In EAE rats, brain NO production and MDA level were significantly increased (P < 0.001) compared to the control values, whereas AG and NAC treatment decreased both parameters in EAE rats compared to EAE group (P < 0.001). Glutathione (GSH) was reduced (P < 0.001) in EAE rats in comparison with the control and CFA groups, but increased in EAE + AG and EAE + NAC group compared to the EAE group (P < 0.01). Superoxide dismutase (SOD) activity was significantly decreased (P < 0.001) in the EAE group compared to all other experimental groups. The clinical expression of EAE was significantly decreased (P < 0.05) in the EAE groups treated with AG and NAC compared to EAE rats, during disease development. The obtained results prove an important role of oxidative and nitrosative stress in the pathogenesis of EAE, whereas AG and NAC protective effects offer new possibilities for a modified combined approach in MS therapy.

The present study was aimed at determining the effectiveness of nitric oxide synthase (NOS) inhibitors: N-nitro-L-arginine methyl ester, 7-nitroindazole and aminoguanidine in modulating the toxicity of AlCl3 on superoxide production and the malondialdehyde concentration of Wistar rats. The animals were sacrificed 10 min and 3 days after the treatment and the forebrain cortex was removed. The results show that AlCl3 exposure promotes oxidative stress in different neural areas. The biochemical changes observed in the neuronal tissues show that aluminum acts as pro-oxidant, while NOS inhibitors exert an anti-oxidant action in AlCl3-treated animals. U eksperimentu je određivana efikasnost inhibitora azot-oksid-sintaze (NOS): metil-estra N-nitro-L-arginina, 7-nitroindazola i aminogvanidina u modulaciji toksičnosti AlCl3 na stvaranje superoksidnog anjona i koncentraciju malondialdehida kod Wistar pacova. Životinje su žrtvovane 10 min i 3 dana nakon tretmana i izolovana je kora velikog mozga. Rezultati pokazuju da izlaganje AlCl3 pokreće oksidativni stres u različitim moždanim regionima. Biohemijske promene opisane u neuronskom tkivu pokazuju da aluminijum deluje kao pro-oksidant, dok inhibitori NOS imaju antioksidativno dejstvo kod životinja tretiranih AlCl3. null

Chronic microglial activation and resulting sustained neuroinflammatory reaction are generally associated with neurodegeneration. Activated microglia acquires proinflammatory cellular profile that generates oxidative burst. Their persistent activation exacerbates inflammation, which damages healthy neurons via cytotoxic mediators, such as superoxide radical anion and nitric oxide. In our recent study, we have shown that benfotiamine (S-benzoylthiamine 0-monophosphate) possesses anti-inflammatory effects. Here, the effects of benfotiamine on the pro-oxidative component of activity of LPS-stimulated BV -2 cells were investigated. The activation of microglia was accompanied by upregulation of intracellular antioxidative defense, which was further promoted in the presence of benfotiamine. Namely, activated microglia exposed to non-cytotoxic doses of benfotiamine showed increased levels and activities of hydrogen peroxide- and superoxide removing enzymes catalase and glutathione system, and superoxide dismutase. In addition, benfotiamine showed the capacity to directly scavenge superoxide radical anion. As a consequence, benfotiamine suppressed the activation of microglia and provoked a decrease in NO and O-center dot(2)- production and lipid peroxidation. In conclusion, benfotiamine might silence pro-oxidative activity of microglia to alleviate/prevent oxidative damage of neighboring CNS cells. Ministry of Education, Science and Technological Development of the Republic of Serbia {[}III41014]

Deficiency in methionine or choline can induce oxidative stress in various organs such as liver, kidney, heart, and brain. This study was to examine the effects of alpha-lipoic acid (LA) on oxidative stress induced by methionine and choline deficiency (MCD) in several brain structures. Male mice C57BL/6 ( n = 28) were divided into four groups: (1) control – continuously fed with standard chow; (2) LA – fed with standard chow and receiving LA; (3) MCD2 – fed with MCD diet for two weeks, and (4) MCD2+LA – fed with MCD diet for two weeks and receiving LA (100 mg/kg/day intraperitonealy [i.p.]). Brain tissue (cortex, hypothalamus, striatum and hippocampus) was taken for determination of oxidative stress parameters. MCD diet induced a significant increase in malondialdehyde and NOx concentration in all brain regions, while LA restored their content to normal values. Similar to this, in MCD2 group, activity of total SOD, MnSOD, and Cu/ZnSOD was reduced by MCD diet, while LA treatment improved their activities in all brain structures. Besides, in MCD2 group a decrease in catalase activity in cortex and GSH content in hypothalamus was evident, while LA treatment induced an increase in catalase activity in cortex and striatum and GSH content in hypothalamus. LA treatment can significantly reduce lipid peroxidation and nitrosative stress, caused by MCD diet, in all brain regions by restoring antioxidant enzymes activities, predominantly total SOD, MnSOD, and Cu/ZnSOD, and to a lesser extent by modulating catalase activity and GSH content. LA supplementation may be used in order to prevent brain oxidative injury induced by methionine and choline deficiency.

The aim of the study was to investigate the hypothesis that agmatine (AGM) provides protection against oxidative stress in experimental autoimmune encephalomyelitis (EAE). Wild-type (WT) and knockout (KO) CBA/H iNOS-/- 3 months old (15 ± 5 g) mice, were used for EAE induction by myelin basic protein (MBP), dissolved in Complete Freund’s Adjuvant (CFA). The animals were divided into control, EAE, CFA, EAE+AGM and AGM groups. After the development of full clinical remission, animals were decapitated and oxidative stress parameters were determined in whole encephalitic mass (WEM) and cerebellum homogenates. The EAE clinical expression manifested to greater extent in WT than KO mice, was significantly decreased during AGM treatment. We demonstrated significant elevations of superoxide dismutase activity in WT and KO EAE animals, in WEM and cerebellum tissues, which were decreased during AGM treatment in both groups. Superoxide anion content was increased in WEM of both study groups, with a decrease during AGM treatment. The observed changes were more pronounced in WT than in KO animals. Also, the increased expressions of transferrin receptor and glial fibrillary acidic protein observed in WT and KO EAE mice were significantly decreased during AGM treatment. The results suggest potentially beneficial AGM effects in EAE, which might be used for a modified antioxidative approach in MS therapy.

Diquat (DQ) neurotoxicity mechanisms are unknown, although, it's systemic toxicity is mediated by free radical reactions. The role of glutathione cycle was assessed by glutathione reductase (GR) applied in the pre-treatment of DQ poisoning. Wistar rats were used and tested compounds were administered intrastriatally (i.s.) in one single dose. Total glutathione (tGSH), glutathione disulfide (GSSG) and glutathione peroxidase (GPx) were measured in the vulnerable brain regions (VBRs) (striatum, hippocampus and cortex), at 30 minutes, 24 hours and 7 days post treatment. Results from the intact and the sham operated groups were not statistically different. Rapid spatial spreading of oxidative stress was confirmed in the examined VBRs.. Mortality (30-40%, within 24hrs) and signs of lethargy were observed in the DQ group. Activity of GPx activity was elevated and GSSG/GSH was higher in the examined VBRs during the experiment, compared to the controls. The i.s. pre-treatment with GR achieved neuroprotective role against DQ induced neurotoxicity, based on animal survival, absence of lethargy and decreased GPx activity and GSSG/GSH in the examined VBRs during the experiment, compared to the DQ group. Our results confirmed that oxidation of GSH was the reason for the reduced antioxidative defense against DQ neurotoxicity. Mehanizmi neurotoksičnosti dikvata (DK) su nepoznati, mada se zna da je sistemska toksičnost posredovana reakcijama slobodnih radikala. Uloga glutationskog ciklusa je isptivana primenom glutation reduktaze (GR) u predtretmanu trovanja DK. Wistar pacovi su korišćeni i testirana jedinjenja intrastrjiatalno (i.s.) primenjena u jednokratnoj dozi. Ukupni glutation (tGSH), glutation-disulfid (GSSG) i aktivnost glutation peroksidaze (GPx) su mereni u selektivno osetljivim regionima mozga (strijatum, hipokampus i korteks), 30. minuta, 24. sata i 7. dana posle tretmana. Rezultati netretiranih (intaktna grupa) i lažno operisanih pacova se ne razlikuju statistički. Vremensko i prostorno širenje oksidativnog stresa je potvrđeno kod ispitivanih moždanih struktura. Mortalitet (30-40%, u roku od 24 casa) i znaci letargije su uočeni u samo u DK grupi. Statistički povećana aktivnost GPx, kao i odnosa GSSG/GSH u ispitivanim moždanim strukturama tokom eksperimenta, potvrđuje oksidativno narušenu ravnotežu i oštećenja moždanog tkiva. Predtretman i.s. sa GR je ispoljio neurozaštitni efekat od neurotoksičnosti DK, bazirano na preživljavanju životinja, odsustvu letargije i smanjenoj aktivnost GPx i odnosa GSSG / GSH ispitivanih moždanih struktura tokom eksperimenta, u odnosu na DK grupu. Naši rezultati ukazuju da je oksidacija GSH kljucna za smanjenje antioksidativne odbrane od DK neurotoksicnosti.

Paraquat (PQ), a widely used herbicide is a well-known free radical producing agent. The mechanistic pathways of PQ neurotoxicity were examined by assessing oxidative/nitrosative stress markers. Focus was on the role of glutathione (GSH) cycle and to examine whether the pre-treatment with enzyme glutathione reductase (GR) could protect the vulnerable brain regions (VBRs) against harmful oxidative effect of PQ. The study was conducted on Wistar rats, randomly divided in five groups: intact-control group, (n = 8) and four experimental groups (n = 24). All tested compounds were administered intrastriatally (i.s.) in one single dose. The following parameters of oxidative status were measured in the striatum, hippocampus and cortex, at 30 min, 24 h and 7 days post treatment: superoxide anion radical (O-2(center dot-)), nitrate (NO3-). malondialdehyde (MDA), superoxide dismutase (SOD), total GSH (tGSH) and its oxidized, disulfide form (GSSG) and glutathione peroxidase (GPx). Results obtained from the intact and the sham operated groups were not statistically different, confirming that invasive i.s. route of administration would not influence the reliability of results. Also, similar pattern of changes were observed between ipsi- and contra- lateral side of examined VBRs, indicating rapid spatial spreading of oxidative stress. Mortality of the animals (10%), within 24 h, along with symptoms of Parkinsonism, after awakening from anesthesia for 2-3 h, were observed in the PQ group, only. Increased levels of O2(center dot-), NO3- and MDA, increased ratio of GSSG/GSH and considerably high activity of GPx were measured at 30 min after the treatment. Cytotoxic effect of PQ was documented by drastic drop of all measured parameters and extremely high peak of the ratio GSSG/GSH at 24th hrs after the PQ i.s. injection. In the GR + PQ group, markedly low activity of GPx and low content of NO3- (in striatum and cortex) were measured during whole experiment, while increase value was observed only for O-2(center dot-), at 7th days. We concluded that oxidative/nitrosative stress and excitotoxicity are the most important events since the early stage of PQ induced neurotoxicity. Based on the ratio GSSG/GSH, the oxidation of GSH to GSSG is probably dominant way of GHS depletion and main reason for reduced antioxidative defense against PQ harmful oxidative effect. The GR pre-treatment resulted in the absence of Parkinson's disease-like symptoms and mortality of the rats. Additionally, oxidative/nitrosative stress did not developed, as well as almost diminished metabolism of the VBRs at 24th hours (as has been documented in the PQgroup) did not occurred in the GR + PQ suggesting a neuroprotective role for the GR in PQ induced neurotoxicity.

In this study we examined if the response of the cortex against diquat (DQ), intrastriatally (i.s.) applied to Wistar rats, was mediated by oxidative/nitrosative stress (OS/NS). In particular, we were focused on the glutathione (GSH) antioxidative role, thus we applied i.s. glutathione reductase (GR) in the pre-treatment of DQ administration. Superoxide anion radical (O2 •-), nitrate (NO3 -), malondialdehyde (MDA) and superoxide dismutase (SOD), were measured in ipsi- and contra- lateral sides of the cortex, at 30 minutes, 24 hours and 7 days post treatment. The redox balance was not significantly changed in the cortex of sham operated and intact groups. Also, no differences were observed between the ipsi- and contra- lateral side of the cortex. Lethargy and mortality (30-40%) of the animals in the DQ group within 24 hrs, coincided with rapidly developed lipid peroxidation supported by OS/NS upon i.s. DQ administration. Strong redox potential of DQ probably resulted in a huge deprivation of molecular oxygen. The pretreatment with GR acted neuro-protectively, based on animal survival and absence of lethargy, although, lipid peroxidation was not developed in the GR+DQ group, OS was documented by a high concentration of O2 •- (within 24 hrs), descending and eventually inhibiting SOD activity (at 7 days). U ovoj studiji smo ispitali da li je oksidativni/nitrosativni stres (OS/NS), uključen u odgovor korteksa Wistar pacova nakon intrastrijatalne (i.s.) izloženosti dikvatu (DK). Posebno smo ispitivali značaj antioksidativne uloge glutationa (GSH), zbog čega smo primenili glutation reduktazu (GR) u predtretmanu davanja DK. Superoksid anjon radikal (O2•¯), nitrati (NO3¯), malondialdehid (MDA) i superoksid dismutaza (SOD), su mereni u obostranom korteksu (ipsi- i kontrastrana), nakon 30 minuta, 24 sati i 7 dana od tretmana. Redoks balans se nije značajno promenio u korteksu lažno operisanih i netretiranih pacova. Takođe, ne postoji statistički značajna razlika između ipsi- i kontra- strane korteksa. Letargija i mortalitet (30-40%) kod životinja u DK grupi su uočene tokom 24 časa od i.s. trovanja DK, što se poklopilo sa naglim razvojem OS/NS i lipidne peroksidacije. Visok redoks potencijal DK verovatno rezultira opsežnim utroškom molekularnog kiseonika. Zaključeno je da je ostvaren neuroprotektivni učinak predtretmana sa GR, na osnovu preživljavanja životinja i odsustva letargije. Lipidna peroksidacija nije bila razvijena u grupi predtretiranoj sa GR ali je ipak izmerena visoka koncentracija O2•¯ (tokom 24 sata) koja zatim opada i na kraju 7. dana u potpunosti inhibira aktivnost SOD.

The present experiment was carried out to determine the effectiveness of nitric oxide synthase inhibitors: 7-nitroindazole and aminoguanidine in modulating the toxicity of aluminium chloride on acetylcholine esterase activity, as well as behavioural and morphological changes of Wistar rats. For biochemical analysis the animals were killed 10 min, 3 h, 3 days and 30 days after the treatment and forebrain cortex, striatum, basal forebrain and hippocampus were removed. The biochemical changes observed in neuronal tissues show that nitric oxide synthase inhibitors exert as protective action in aluminium chloride-treated animals. In the present study, active avoidance learning was significantly impaired after aluminium chloride injection, while pretreatment with nitric oxide synthase inhibitors prevented the behavioural deficits caused between 26th and 30th day after intrahippocampal application of neurotoxin. Our data suggest that aluminium may cause learning and memory deficits, while the treatment with specific nitric oxide synthase inhibitors may prevent learning and memory deficits caused by aluminium chloride. We have also applied immunohistochemical techniques to identify neuronal- and inducible-nitric oxide synthase expression 30 days after aluminium chloride and nitric oxide synthase inhibitors injections. Our data suggest that 7-nitroindazole and aminoguanidine can be effective in the protection of toxicity induced by aluminium chloride.

The present experiments were done to determine the effectiveness of a non-specific nitric oxide synthase inhibitor, N-nitro-L-arginine methyl ester (L-NAME), on oxidative stress parameters induced by aluminium chloride (AlCl(3)) intrahippocampal injections in Wistar rats. Animals were sacrificed 3 h and 30 d after treatments, heads were immediately frozen in liquid nitrogen and forebrain cortices were removed. Crude mitochondrial fraction preparations of forebrain cortices were used for the biochemical analyses: nitrite levels, superoxide production, malondialdehyde concentrations, superoxide dismutase (SOD) activities and reduced glutathione contents. AlCl(3) injection resulted in increased nitrite concentrations, superoxide anion production, malondialdehyde concentrations and reduced glutathione contents in the forebrain cortex, suggesting that AlCl(3) exposure promoted oxidative stress in this brain structure. The biochemical changes observed in neuronal tissues showed that aluminium acted as a pro-oxidant. However, the nonspecific nitric oxide synthase (NOS) inhibitor, L-NAME, exerted anti-oxidant actions in AlCl(3)-treated animals. These results revealed that NO-mediated neurotoxicity due to intrahippocampal AlCl3 injection spread temporally and spatially to the forebrain cortex, and suggested a potentially neuroprotective effect for L-NAME.