Daniel Henrique Roos

Protective effect of Melissa officinalis aqueous extract against Mn-induced oxidative stress in chronically exposed mice.

Manganese (Mn) is an essential element for biological systems; however occupational exposure to high levels of this metal may lead to neurodegenerative disorders, resembling Parkinsons disease (PD). While its mechanisms of neurotoxicity have yet to be fully understood, oxidative stress plays a critical role. Thus, the main goal of this study was to investigate the efficacy of aqueous extract of Melissa officinalis in attenuating Mn-induced brain oxidative stress in mice. Sixteen male mice were randomly divided into two groups and treated for 3 months: the first group consumed tap water (control group) and the second group was treated with Mn (50 mg/kg/day for habituation during the first 15 days followed by 100 mg/kg/day for additional 75 days) in the drinking water. After 3 months both groups were sub divided (n=4 per group) and treated for additional 3 months with Mn and/or M. officinalis in the drinking water. The first group (control) was treated with water and served as control; the second group (M. officinalis) was treated with M. officinalis (100 mg/kg/day); the third group was treated with Mn (100 mg/kg/day); the fourth group (Mn+M. officinalis) was treated with both Mn and M. officinalis (100 mg/kg/day each). Mn-treated mice showed a significant increase in thiobarbituric acid reactive species (TBARS) levels (a marker of oxidative stress) in both the hippocampus and striatum. These changes were accompanied by a decrease in total thiol content in the hippocampus and a significant increase in antioxidant enzyme activity (superoxide dismutase and catalase) in the hippocampus, striatum, cortex and cerebellum. Co-treatment with M. officinalis aqueous extract in Mn-treated mice significantly inhibited the antioxidant enzyme activities and attenuated the oxidative damage (TBARS and decreased total thiol levels). These results establish that M. officinalis aqueous extract possesses potent antioxidative properties, validating its efficacy in attenuating Mn-induced oxidative stress in the mouse brain.

Modulation of methylmercury uptake by methionine: prevention of mitochondrial dysfunction in rat liver slices by a mimicry mechanism.

Methylmercury (MeHg) is an ubiquitous environmental pollutant which is transported into the mammalian cells when present as the methylmercury-cysteine conjugate (MeHg-Cys). With special emphasis on hepatic cells, due to their particular propensity to accumulate an appreciable amount of Hg after exposure to MeHg, this study was performed to evaluate the effects of methionine (Met) on Hg uptake, reactive species (RS) formation, oxygen consumption and mitochondrial function/cellular viability in both liver slices and mitochondria isolated from these slices, after exposure to MeHg or the MeHg-Cys complex. The liver slices were pre-treated with Met (250 μM) 15 min before being exposed to MeHg (25 μM) or MeHg-Cys (25 μM each) for 30 min at 37 °C. The treatment with MeHg caused a significant increase in the Hg concentration in both liver slices and mitochondria isolated from liver slices. Moreover, the Hg uptake was higher in the group exposed to the MeHg-Cys complex. In the DCF (dichlorofluorescein) assay, the exposure to MeHg and MeHg-Cys produced a significant increase in DFC reactive species (DFC-RS) formation only in the mitochondria isolated from liver slices. As observed with Hg uptake, DFC-RS levels were significantly higher in the mitochondria treated with the MeHg-Cys complex compared to MeHg alone. MeHg exposure also caused a marked decrease in the oxygen consumption of liver slices when compared to the control group, and this effect was more pronounced in the liver slices treated with the MeHg-Cys complex. Similarly, the loss of mitochondrial activity/cell viability was greater in liver slices exposed to the MeHg-Cys complex when compared to slices treated only with MeHg. In all studied parameters, Met pre-treatment was effective in preventing the MeHg- and/or MeHg-Cys-induced toxicity in both liver slices and mitochondria. Part of the protection afforded by Met against MeHg may be related to a direct interaction with MeHg or to the competition of Met with the complex formed between MeHg and endogenous cysteine. In summary, our results show that Met pre-treatment produces pronounced protection against the toxic effects induced by MeHg and/or the MeHg-Cys complex on mitochondrial function and cell viability. Consequently, this amino acid offers considerable promise as a potential agent for treating acute MeHg exposure.

Complex methylmercury-cysteine alters mercury accumulation in different tissues of mice

Methylmercury (MeHg) can cause deleterious effects in vertebrate tissues, particularly in the central nervous system. MeHg interacts with sulfhydryl groups from low and high molecular weight thiols in the blood, which can facilitate MeHg uptake into different tissues. The purpose of this study was to examine the effect of MeHg-Cysteine (MeHg-Cys) complex administration on Hg-uptake in cerebral areas (cortex and cerebellum), liver and kidney of adult mice. Animals were divided into four groups: control (1 mL/kg distilled water), MeHg (2 mg/kg), Cys (2 mg/kg) and MeHg-Cys complex (0.8 molar ratio). Mice received one intraperitoneal injection per day for 60 consecutive days. Treatment with MeHg significantly increased mercury concentrations in all tissues analysed when compared with the control group. The accumulation of mercury in brain and in liver was further increased in animals that received MeHg-Cys complex when compared with the MeHg alone group. However, renal Hg decreased in MeHg-Cys treated mice, when compared with the group treated only with MeHg. In summary, the transport of MeHg-Cys complex was tissue-specific, and we observed an increase in its uptake by liver and brain as well as a decrease in kidney.

Role of Calcium and Mitochondria in MeHg-Mediated Cytotoxicity

Methylmercury (MeHg) mediated cytotoxicity is associated with loss of intracellular calcium (Ca2+) homeostasis. The imbalance in Ca2+ physiology is believed to be associated with dysregulation of Ca2+ intracellular stores and/or increased permeability of the biomembranes to this ion. In this paper we summarize the contribution of glutamate dyshomeostasis in intracellular Ca2+ overload and highlight the mitochondrial dysfunctions induced by MeHg via Ca2+ overload. Mitochondrial disturbances elicited by Ca2+ may involve several molecular events (i.e., alterations in the activity of the mitochondrial electron transport chain complexes, mitochondrial proton gradient dissipation, mitochondrial permeability transition pore (MPTP) opening, thiol depletion, failure of energy metabolism, reactive oxygen species overproduction) that could culminate in cell death. Here we will focus on the role of oxidative stress in these phenomena. Additionally, possible antioxidant therapies that could be effective in the treatment of MeHg intoxication are briefly discussed.

Intense exercise potentiates oxidative stress in striatum of reserpine-treated animals.

Regular physical activity exerts beneficial effects for mental and physical health, but an intense exercise can cause oxidative stress (OS) in dopaminergic regions and intensify the harmful effects of reserpine. Reserpine-induced neurotoxicity can be accessed by behavioral and biochemical evaluations. The objective of this study was to examine the effect of a gradual intensifying exercise program on an animal model of oxidative stress. Male rats were submitted to swimming sessions (1 h/day, for eleven weeks), and they were loaded gradually during the adaptation period (two weeks) with a weight corresponding to 1-7% of their body weight tied to their back. After the last training, the animals were treated with two doses of vehicle or reserpine (1 mg/kg-sc), an agent that induces orofacial dyskinesia. After behavioral evaluations, the striatum was dissected for enzymatic and biochemical assays. Development of cardiac hypertrophy demonstrated the effectiveness of the physical training. The gradual intense exercise and reserpine increased lipid peroxidation and striatal catalase activity. The results confirm the importance of catalase activity in orofacial dyskinesia which can be related to lipid peroxidation in striatal dopaminergic brain tissue. These results indicate that intense exercise can have some deleterious effect on striatal dopaminergic system.

Mitochondrial electron transfer chain complexes inhibition by different organochalcogens.

Mitochondrial dysfunction plays a pivotal role in the cell toxicology and death decision. The aim of the present study was to investigate the effect of three organocompounds (ebselen [Ebs], diphenyl diselenide [(PhSe)(2)] and diphenyl ditelluride [(PhTe)(2)]) on mitochondrial complexes (I, II, I-III, II-III and IV) activity from rat liver and kidney to determine their potential role as molecular targets of organochalcogens. All studied organochalcogens caused a statistically significant inhibition of the mitochondrial complex I activity. Ebs and (PhTe)(2) caused a statistically significant inhibition of the mitochondrial complex II activity in both hepatic and renal membranes. Hepatic mitochondrial complex II activity was practically unchanged by (PhSe)(2), whereas it significantly inhibited renal complex II activity. Mitochondrial complex IV activity was practically unchanged by the organochalcogens. Furthermore, organochalcogens inhibited the mitochondrial respiration supported by complex I or complex II substrates. The inhibitory effect of Ebs, (PhSe)(2) and (PhTe)(2) on mitochondrial complex I was prevented by NADH, but it was not prevented by catalase (CAT) and/or superoxide dismutase (SOD). Additionally, the organochalcogens-induced inhibition of complex I and II was completely reversed by reduced glutathione (GSH). In conclusion, Ebs, (PhSe)(2) and (PhTe)(2) were more effective inhibitors of renal and hepatic mitochondrial complex I than complex II, whereas complexes III and IV were little modified by these compounds. Taking into account the presented results, we suggest that organochalcogen-induced mitochondrial complexes I and II inhibition can be mediated by their thiol oxidation activity, i.e., Ebs, (PhSe)(2) and (PhTe)(2) can oxidize critical thiol groups from mitochondrial complexes I and II. So, mitochondrial dysfunction can be considered an important factor in the toxicity of Ebs, (PhSe)(2) and (PhTe)(2).

In vitro antioxidant activity of stem bark of Trichilia catigua Adr. Juss

Antioxidant activity of the ethanolic extract and fractions from the stem bark of T. catigua was investigated. IC50 (for DPPH scavenging) by T. catigua varied from 9.17 ± 0.63 to 76.42 ± 5.87 mg mL-1 and total phenolic content varied from 345.63 ± 41.08 to 601.27 ± 42.59 mg GAE g-1 of dry extract. Fe2+-induced lipid peroxidation was significantly reduced by the ethanolic extract and fractions. Mitochondrial Ca2+-induced dichlorofluorescein oxidation was significantly reduced by the ethanolic extract in a concentration-dependent manner. Ethanolic extract reduced mitochondrial Dym only at high concentrations (40-100 mg mL-1), which indicates that its toxicity does not overlap with its antioxidant effects. Results suggest involvement of antioxidant activities of T. catigua in its pharmacological properties. U radu je opisano ispitivanje antioksidativnog u~inka etanolnog ekstrakta i pojedinih frakcija kore stabljike T. catigua. IC50 (za DPPH test) varirao je izme|u 9,17 ± 0,63 i 76,42 ± 5,87 mg mL-1, a ukupni sadr`aj fenola od 345,63 ± 41,08 i 601,27 ± 42,59 mg GAE po gramu suhog ekstrakta. Etanolni ekstrakt i frakcije zna~ajno su reducirale Fe2+-induciranu lipidnu peroksidaciju. Nadalje, reducirana je oksidacija diklorfluoresceina inducirana ionima kalcija u mitohondrijima, a redukcija je ovisila o dozi etanolnog ekstrakta. Etanolni ekstrakt smanjio je mitohondrijsku Dym samo pri visokim koncentracijama (40 ± 100 mg mL-1), {to ukazuje da se toksi~nost ne preklapa s antioksidativnim u~inkom. Rezultati pokazuju da u farmakolo{ko djelovanje T. catigua treba uklju~iti i antioksidativni u~inak.

Copper and selenium: Auxiliary measure to control infection by Haemonchus contortus in lambs

The aim of this study was to evaluate the effects of selenium and copper on oxidative stress and its performance in lambs experimentally infected with Haemonchus contortus. Twenty-eight five-months old lambs were experimentally infected by the oral route with 5000 third-stage infective larvae and allocated into four groups, i.e., untreated animals, animals treated intramuscularly with sodium selenite (0.2 mg kg(-1)), animals treated subcutaneously with copper (3.5 mg kg(-1)), and animals treated with sodium selenite (IM; 0.2 mg kg(-1)) and copper (SC; 3.5 mg kg(-1)). These animals received oat hay (Avena sativa) and commercial concentrate, totaling 15% of crude protein, 30% being derived from oat hay and 70% of the concentrate. Lipid peroxidation, antioxidant enzymes, eggs per gram of feces (EPG) and body weight were assessed on the day of infection and after 20, 40, 60 and 80 days post-infection. The number of H. contortus adults was assessed at the end of the experiment. The selenium associated or not with copper reduced the effects of oxidative stress caused by infection. The groups supplemented with copper had increased body weight, and the combination of these two minerals reduced the EPG and number of H. contortus adults in lambs. The use of selenium associated with copper may help the control of infection by H. contortus.

Ilex paraguariensis crude extract acts on protection and reversion from damage induced by t-butyl hydroperoxide in human erythrocytes: a comparative study with isolated caffeic and/or chlorogenic acids.

BACKGROUND Studies comparing the effects of phytochemicals under different regimens of exposure are necessary to give a better indication about their mechanism(s) of protection. Hence, in the present study, we investigated the preventive (pre-incubation), protective (co-incubation) and/or remediative (post-incubation) activity of chlorogenic acid and caffeic acids, in comparison with Ilex paraguariensis crude extract, against t-butyl hydroperoxide (t-BHP)-induced damage to human erythrocytes. RESULTS We found that both caffeic and chlorogenic acids were able to prevent and revert the hemolysis associated with t-BHP exposure. By contrast, isolated compounds (alone or in combination) presented no effect on basal and/or t-BHP-induced non-protein thiol (NPSH) oxidation or production of thiobarbituric acid reactive substances (TBBARS). In turn, I. paraguariensis extract was effective to prevent, protect and revert the hemolysis associated with t-BHP exposure. Moreover, I. paraguariensis significantly protects and reverts t-BHP-induced NPSH oxidation and TBARS production. CONCLUSIONS We have found that I. paraguariensis extract acts better with respect to the protection and reversion of t-BHP-associated changes, whereas isolated compounds are more active in preventing and reverting t-BHP pro-hemolytic action. Moreover, our data suggest that the pro-hemolytic activity of t-BHP may occur via mechanism(s) other(s) than lipid peroxidation and/or NPSH oxidation.

Organochalcogens Inhibit Mitochondrial Complexes I and II in Rat Brain: Possible Implications for Neurotoxicity

Organochalcogens, such as organoselenium and organotellurium compounds, can be neurotoxic to rodents. Since mitochondrial dysfunction plays a pivotal role in neurological disorders, the present study was designed to test the hypothesis that rat brain mitochondrial complexes (I, II, I–III, II–III and IV) could be molecular targets of organochalcogens. The results show that organochalcogens caused statistically significant inhibition of mitochondrial complex I activity, which was prevented by preincubation with NADH and fully blunted by reduced glutathione (GSH). Mitochondrial complex II activity remained unchanged in response to (PhSe)2 treatment. Ebs and (PhTe)2 caused a significant concentration-dependent inhibition of complex II that was also blunted by GSH. Mitochondrial complex IV activity was not modified by organochalcogens. Collectively, Ebs, (PhSe)2 and (PhTe)2 were more effective inhibitors of brain mitochondrial complex I than of complex II, whereas they did not affect complex IV. These observations are consistent with organochalcogens inducing mitochondrial complex I and II inhibition via their thiol-oxidase-like activity, with Ebs, (PhSe)2 and (PhTe)2 effectively oxidising critical thiol groups of these complexes.