Effects of Rosmarinus officinalis L. (Laminaceae) essential oil on adult and larvae of Drosophila melanogaster.

Rosmarinus officinalis (Lamiaceae family), also known as "alecrim," is a perennial herb, typical of the Mediterranean region and widely distributed in Brazilian territory. Despite having demonstrated several properties of human interest, insecticide/larvicidal effect of essential oil from R. officinalis on insects remains unclear. In this study, we tested the effects of R. officinalis essential oil on biomarkers of oxidative damage in Drosophila melanogaster. Exposure to R. officinalis essential oil increased adult mortality and decreased geotaxis behavior in adult fruit flies. In addition, essential oil increased of larval mortality and impaired the developmental success in D. melanogaster. R. officinalis essential oil showed a significant repellent effect, with duration time of about 6 h. To understand the mechanism underlying the toxicity of essential oil both pro-oxidant effects and biomarkers of oxidative damage were evaluated in exposed flies. Exposure to essential oil caused a significant redox imbalance with impairment of both enzymatic and non-enzymatic antioxidant system and increased the lipid peroxidation levels. These results suggest that R. officinalis essential oil can be used as a bioinsecticide and/or larvicide as well as an alternative insect repellent.

Mancozeb impairs mitochondrial and bioenergetic activity in Drosophila melanogaster.

Mancozeb (MZ) is a broad-spectrum fungicide used worldwide in several crops. Neurological disorders in humans and animals have been associated with exposure to this compound by mechanisms still not fully understood. Drosophila melanogaster represents a reliable model in toxicological studies, presenting genetic and biochemical similarities with mammals. In this study, D. melanogaster flies were exposed for 15 days to MZ through the food (5 and 10 mg/mL). After that period, the efficiency of mitochondrial respiration complexes and metabolic markers were analyzed and evaluated. Flies presented weight loss, lower glucose, trehalose, and glycogen levels, and augmented levels of triglycerides concerning control (non-treated group). Acetyl-CoA Synthetase (ACeCS-1) and Acyl-Coenzyme Synthetase (ACSL1) contents were unchanged by MZ treatment. Mitochondrial respiration of flies was targeted by MZ treatment, evidenced by a decrease in oxygen consumption and bioenergetics rate and inhibition in mitochondrial complexes I/II. These results suppose that an impairment in mitochondrial respiration jointly with reduced levels of energetic substrates might be a mechanism involved in MZ deleterious effects, possibly by the limitation of ATP's availability, necessary for essential cellular processes.

Guanosine protects against behavioural and mitochondrial bioenergetic alterations after mild traumatic brain injury.

Traumatic brain injury (TBI) constitutes a heterogeneous cerebral insult induced by traumatic biomechanical forces. Mitochondria play a critical role in brain bioenergetics, and TBI induces several consequences related with oxidative stress and excitotoxicity clearly demonstrated in different experimental model involving TBI. Mitochondrial bioenergetics alterations can present several targets for therapeutics which could help reduce secondary brain lesions such as neuropsychiatric problems, including memory loss and motor impairment. Guanosine (GUO), an endogenous neuroprotective nucleoside, affords the long-term benefits of controlling brain neurodegeneration, mainly due to its capacity to activate the antioxidant defense system and maintenance of the redox system. However, little is known about the exact protective mechanism exerted by GUO on mitochondrial bioenergetics disruption induced by TBI. Thus, the aim of this study was to investigate the effects of GUO in brain cortical and hippocampal mitochondrial bioenergetics in the mild TBI model. Additionally, we aimed to assess whether mitochondrial damage induced by TBI may be related to behavioral alterations in rats. Our findings showed that 24 h post-TBI, GUO treatment promotes an adaptive response of mitochondrial respiratory chain increasing oxygen flux which it was able to protect against the uncoupling of oxidative phosphorylation (OXPHOS) induced by TBI, restored the respiratory electron transfer system (ETS) established with an uncoupler. Guanosine treatment also increased respiratory control ratio (RCR), an indicator of the state of mitochondrial coupling, which is related to the mitochondrial functionality. In addition, mitochondrial bioenergetics failure was closely related with locomotor, exploratory and memory impairments. The present study suggests GUO treatment post mild TBI could increase GDP endogenous levels and consequently increasing ATP levels promotes an increase of RCR increasing OXPHOS and in substantial improve mitochondrial respiration in different brain regions, which, in turn, could promote an improvement in behavioral parameters associated to the mild TBI. These findings may contribute to the development of future therapies with a target on failure energetic metabolism induced by TBI.

Acetaminophen Oxidation and Inflammatory Markers - A Review of Hepatic Molecular Mechanisms and Preclinical Studies.

Acetaminophen is a widely used analgesic for pain management, especially useful in chronic diseases, such as rheumatoid arthritis. However, easy access to this medicine has increased the occurrence of episodes of poisoning. Patients often develop severe liver damage, which may quickly lead to death. Consequently, numerous studies have been conducted to identify new biomarkers that allow the prediction of the degree of acetaminophen intoxication and thus intervene in a timely manner to save patients' lives. This review highlights the main mechanisms of the induction and progression of liver damage arising from acetaminophen poisoning. In addition, we have discussed the possibility of using new clinical biomarkers for detecting acetaminophen poisoning.

Acute Exposure to Permethrin Modulates Behavioral Functions, Redox, and Bioenergetics Parameters and Induces DNA Damage and Cell Death in Larval Zebrafish.

Permethrin (PM) is a synthetic pyrethroid insecticide widely used as domestic repellent. Damage effects to nontarget organisms have been reported, particularly in the early stages of development. Studies indicate redox unbalance as secondary PM effect. Therefore, our goal was to investigate the acute PM effects on larval zebrafish. Larvae (6 days postfertilization) were exposed to PM (25-600 µg/L) during 24 hours, and 50% lethal concentration was estimated. For subsequent assays, the sublethal PM concentrations of 25 and 50 µg/L were used. PM increased anxiety-like behaviors according to the Novel Tank and Light-Dark tests. At the molecular level, PM induced increased ROS, which may be related to the increased lipid peroxidation, DNA damage, and apoptosis detected in PM-exposed organisms. In parallel, upregulation of the antioxidant system was detected after PM exposure, with increased superoxide dismutase, glutathione S-transferase and glutathione reductase activities, and thiol levels. The increased of Nrf2 target genes and the activation of an electrophile response element-driven reporter Tg(EPRE:LUC-EGFP) suggest that the Nrf2 pathway can mediate a fast response to PM, leading to antioxidant amplification. By using high-resolution respirometry, we found that exposure to PM decreased the oxygen consumption in all respiratory stages, disrupting the oxidative phosphorylation and inhibiting the electron transfer system, leading to decrease in bioenergetics capacity. In addition, PM led to increases of residual oxygen consumption and changes in substrate control ratio. Glucose metabolism seems to be affected by PM, with increased lactate dehydrogenase and decreased citrate synthase activities. Taken together, our results demonstrated the adverse effects of acute sublethal PM concentrations during larval development in zebrafish, causing apparent mitochondrial dysfunction, indicating a potential mechanism to redox unbalance and oxidative stress, which may be linked to the detected cell death and alterations in normal behavior patterns caused by acute PM exposure.

Guanosine protects against Ca2+-induced mitochondrial dysfunction in rats.

Mitochondria play an important role in cell life and in the regulation of cell death. In addition, mitochondrial dysfunction contributes to a wide range of neuropathologies. The nucleoside Guanosine (GUO) is an endogenous molecule, presenting antioxidant properties, possibly due to its direct scavenging ability and/or from its capacity to activate the antioxidant defense system. GUO demonstrate a neuroprotective effect due to the modulation of the glutamatergic system and maintenance of the redox system. Thus, considering the few studies focused on the direct effects of GUO on mitochondrial bioenergetics, we designed a study to evaluate the in vitro effects of GUO on rat mitochondrial function, as well as against Ca2+-induced impairment. Our results indicate that GUO prevented mitochondrial dysfunction induced by Ca2+ misbalance, once GUO was able to reduce mitochondrial swelling in the presence of Ca2+, as well as ROS production and hydrogen peroxide levels, and to increase manganese superoxide dismutase activity, oxidative phosphorylation and tricarboxylic acid cycle activities. Our study indicates for the first time that GUO could direct prevent the mitochondrial damage induced by Ca2+ and that these effects were not related to its scavenging properties. Our data indicates that GUO could be included as a new pharmacological strategy for diseases linked to mitochondrial dysfunction.

Anacardium microcarpum Promotes Neuroprotection Dependently of AKT and ERK Phosphorylation but Does Not Prevent Mitochondrial Damage by 6-OHDA.

Parkinson's disease is a degenerative and progressive illness characterized by the degeneration of dopaminergic neurons. 6-hydroxydopamine (6-OHDA) is a widespread model for induction of molecular and behavioral alterations similar to Parkinson and has contributed for testing of compounds with neuroprotective potential. The Brazilian plant Anacardium microcarpum is used in folk medicine for treatment of several illnesses; however, the knowledge about toxicology and biological effects for this plant is very rare. The neuroprotective effect from hydroalcoholic extract and methanolic and acetate fraction of A. microcarpum on 6-OHDA-induced damage on chicken brain slices was investigated in this study. 6-OHDA decreased cellular viability measured by MTT reduction assay, induced lipid peroxidation by HPLC, stimulated Glutathione-S-Transferase and Thioredoxin Reductase activity, and decreased Glutathione Peroxidase activity and the total content of thiols containing compounds. The methanolic fraction of A. microcarpum presented the better neuroprotective effects in 6-OHDA-induced damage in relation with hydroalcoholic and acetate fraction. The presence of AKT and ERK1/2 pharmacological inhibitors blocked the protective effect of methanolic fraction suggesting the involvement of survival pathways in the neuroprotection by the plant. The plant did not prevent 6-OHDA autoxidation or 6-OHDA-induced mitochondrial dysfunction. Thus, the neuroprotective effect of the methanolic fraction of A. microcarpum appears to be attributed in part to chelating properties of extract toward reactive species and is dependent on ERK1/2 and AKT phosphorylation. This study contributes to the understanding of biochemical mechanisms implied in neuroprotective effects of the vegetal species A. microcarpum.

Exposure of Drosophila melanogaster to Mancozeb Induces Oxidative Damage and Modulates Nrf2 and HSP70/83.

Mancozeb (MZ), a manganese- and zinc-containing ethylene-bis-dithiocarbamate, is a broad-spectrum fungicide. Harmful effects of this fungicide have been reported in nontarget organisms via a not fully understood mechanism. Drosophila melanogaster has provided remarkable contributions for toxicological studies. This work was aimed at evaluating the biochemical targets and implication of oxidative stress in MZ-mediated toxicity in drosophilas. Exposure of flies for fifteen days to MZ at 5 and 10 mg/mL through the diet impaired locomotor performance and induced fly mortality. In parallel, it caused lipid peroxidation and reactive oxygen species (ROS) formation and Mn overload. MZ inhibited superoxide dismutase and inducted catalase and glutathione S-transferase activities. Nitric oxide and reduced glutathione levels were significantly decreased by MZ. Heat shock proteins (HSP70 and HSP83) and Nrf2 mRNA levels were significantly augmented in MZ-exposed flies. Our study reinforced the use of Drosophila melanogaster as a reliable model for the study of biochemical targets of pesticides, and based on our data, MZ induced oxidative damage and Mn accumulation in a concentration-dependent manner. An adaptative cellular state was inducted by the lower concentration of pesticide, possibly contributing to the slighter damage observed.

N-acetylcysteine inhibits Mancozeb-induced impairments to the normal development of zebrafish embryos.

Mancozeb (MZ), a manganese/zinc-containing ethylene-bis-dithiocarbamate (EBCD) fungicide has been claimed to present low acute toxicity and short environmental persistence, however, its effects on embryogenesis in non-target organisms is unclear. Here, we used zebrafish embryos (5 hpf) to assess the potential embryotoxic effects induced by MZ (up to 72 hpf) as well as the role of reactive oxygen species (ROS) in this process by pre-treatment with a classical antioxidant (N-acetylcysteine, NAC). Markers of reactive oxygen species production (ROS), glutathione (GSH) levels and glutathione S-transferase (GST) activity were measured along with genotoxicity (comet assay), cell death (Acridine Orange) and behavioral parameters (spontaneous movement, touch stimulation and swimming response), in order to determine potential mechanisms of embryotoxicity. According to results, MZ was able to induce morphological abnormalities such as body axis distortion, DNA damage, cell death, increased ROS generation and changes in behavioral endpoints during zebrafish development. All these toxic effects were inhibited by the pre-treatment with NAC indicating a key role of redox unbalance during MZ-induced embryotoxicity. At least in our knowledge, this is the first report on the deleterious effect of MZ to the normal embryogenesis of zebrafish. In addition, the importance of ROS generation during this pathophysiological condition was highlighted.

Mancozeb exposure results in manganese accumulation and Nrf2-related antioxidant responses in the brain of common carp Cyprinus carpio.

Manganese (Mn)-containing dithiocarbamates such as Mancozeb (MZ) have been shown to induce oxidative stress-related toxicity in rodents and humans. However, little is known about the neurotoxic effects induced by MZ in fish. In this study, carp (Cyprinus carpio) were exposed to non-lethal waterborne concentrations of MZ, and oxidative stress parameters as well as metal accumulation in fish brains were evaluated. The experimental groups were as follows: control, MZ 5 mg/L, and MZ 10 mg/L. Fish were exposed for 7 days, and then brain was removed and prepared for subsequent analysis of antioxidant enzymes, reactive oxygen species (ROS), and expression of Nrf2 and phosphoNrf2. In parallel, manganese (Mn) levels were evaluated in blood and brain tissues. Mn levels were significantly increased in blood and brain of MZ-exposed carps. In addition, a concentration-dependent increase (p < 0.05) in ROS levels was observed in parallel to increments (p < 0.05) in the activity of major antioxidant enzymes, such as GPx, GR, and GST. On the other hand, significant decreases (p < 0.05) in CAT and SOD activities were observed. The expression of total and phosphorylated forms of Nrf2 was significantly (p < 0.05) upregulated in the brain of carps exposed to Mz when compared to the control, indicating an activation of the Nrf2 antioxidant pathway. Our study showed for the first time the activation of the Nrf2/ARE pathway and bioaccumulation of Mn induced by MZ exposure in fish species, highlighting important mechanisms of action and its toxicological impacts to aquatic organisms.

Free radical scavenging in vitro and biological activity of diphenyl diselenide-loaded nanocapsules: DPDS-NCS antioxidant and toxicological effects.

Selenium compounds, such as diphenyl diselenide (DPDS), have been shown to exhibit biological activity, including antioxidant effects. However, the use of DPDS in pharmacology is limited due to in vivo pro-oxidative effects. In addition, studies have shown that DPDS-loaded nanocapsules (DPDS-NCS) have greater bioavailability than free DPDS in mice. Accordingly, the aim of this study was to investigate the antioxidant properties of DPDS-NCS in vitro and biological activity in mice. Our in vitro results suggested that DPDS-NCS significantly reduced the production of reactive oxygen species and Fe(II)-induced lipid peroxidation (LPO) in brain. The administration of DPDS-NCS did not result in death or change the levels of endogenous reduced or oxidized glutathione after 72 hours of exposure. Moreover, ex vivo assays demonstrated that DPDS-NCS significantly decreased the LPO and reactive oxygen species levels in the brain. In addition, the highest dose of DPDS-NCS significantly reduced Fe(II)- and sodium nitroprusside-induced LPO in the brain and Fe(II)-induced LPO in the liver. Also, δ-aminolevulinate acid dehydratase within the brain was inhibited only in the highest dose of DPDS-NCS. In conclusion, our data demonstrated that DPDS-NCS exhibited low toxicity in mice and have significant antioxidant characteristics, indicating that nanoencapsulation is a safer method of DPDS administration.

Neuroprotective effect of diphenyl diselenide in a experimental stroke model: maintenance of redox system in mitochondria of brain regions.

Acute stroke is a major risk for morbidity and mortality in aging population. Mitochondrion has been the focus of a wide stroke-related research. This study investigated if treatment or pre-treatment with diphenyl diselenide (PhSe)2 can prevent mitochondrial damage in cerebral structures of rats induced by an ischemia and reperfusion (I/R) model. Adult male Wistar rats were assigned into five experimental groups: sham operation, ischemia/reperfusion, pre-treated + I/R, treated + I/R, and Sham + (PhSe)2. Neurological score showed the damage caused by I/R, which was partially prevented by (PhSe)2. Moreover, mitochondria of hippocampus and cortex were impaired by I/R through an increase of reactive oxygen species production, mitochondrial membrane potential (ΔΨm) and electrons flow alteration, activity of complex I deregulation as well as mitochondrial swelling. However, the ischemic damage did not induce an increase in pro-apoptotic proteins expression, but demonstrated an enhanced expression of Hsp70. The mitochondrial redox state was also altered (GSH/GSSG ratio, MnSOD, and GPx activities). Our results revealed that all treatments with (PhSe)2 significantly reduced the mitochondrial damage induced by I/R. These findings suggest that neuroprotective properties of (PhSe)2 may be attributed to the maintenance of mitochondrial redox balance.

Protective action of ethanolic extract of Rosmarinus officinalis L. in gastric ulcer prevention induced by ethanol in rats.

The pathology of a gastric ulcer is complex and multifactorial. Gastric ulcers affect many people around the world and its development is a result of the imbalance between aggressive and protective factors in the gastric mucosa. In this study, we evaluated the ethanolic extract of Rosmarinus officinalis L. (eeRo); this plant, more commonly known as rosemary, has attracted the interest of the scientific community due to its numerous pharmacological properties and their potential therapeutic applications. Here, we tested the preventive effects of eeRo against gastric ulcer induced by 70% ethanol in male Wistar rats. In addition, we aimed to clarify the mechanism involved in the preventive action of the eeRo in gastric ulcers. Based on the analysis of markers of oxidative damage and enzymatic antioxidant defense systems, the measurement of nitrite and nitrate levels and the assessment of the inflammatory response, the eeRo exhibited significant antioxidant, vasodilator and antiinflammatory properties.

Acute brain damage induced by acetaminophen in mice: effect of diphenyl diselenide on oxidative stress and mitochondrial dysfunction.

Organoselenium compounds exhibit antioxidant activity, as well as a variety of biological activities, with potential pharmacological and therapeutic applications. The aim of this study was to investigate the effect of diphenyl diselenide (PhSe)(2) in reversing oxidative brain damage and mitochondrial dysfunction caused by administration of acetaminophen (APAP) in mice. Mice received a toxic dose of APAP, followed by a dose of (PhSe)(2) 1 h later. Four hours after the administration of APAP, plasma was withdrawn from the mice and used for biochemical assays of aspartate aminotransferase (AST) and alanine aminotransferase (ALT) as markers of hepatotoxicity. Brain homogenate was examined to determine oxidative stress. Isolated brain mitochondria were examined to quantify mitochondrial transmembrane's electrical potential and mitochondrial swelling and to estimate reactive oxygen species (ROS) production. APAP administration caused an increase in plasma ALT and AST activities. APAP administration also caused a significant increase in the levels of thiobarbituric acid reactive substances (TBARS) and dichlorofluorescein oxidation in brain homogenate. Similarly, mitochondrial swelling and ROS production increased after APAP administration. APAP treatment also caused a decrease in Na(+), K(+)- ATPase activity and in mitochondrial membrane potential. These alterations observed in the brain of APAP-treated mice were restored by (PhSe)(2). Glutathione levels were decreased by APAP, but (PhSe)(2) did not reverse this change. Treatment with (PhSe)(2) after APAP administration can reverse the neurotoxicity caused by a single toxic dose of APAP. The neuroprotective effect of (PhSe)(2) is likely associated with its antioxidant properties.

Butane-2,3-dionethiosemicarbazone: an oxime with antioxidant properties.

Oximes are compounds generally used to reverse the acetylcholinesterase (AChE) inhibition caused by organophosphates (OPs). The aim of this study was to examine the capacity of the butane-2,3-dionethiosemicarbazone oxime to scavenge different forms of reactive species (RS) in vitro, as well as counteract their formation. The potential antioxidant and toxic activity of the oxime was assayed both in vitro and ex vivo. The obtained results indicate a significant hydrogen peroxide (H2O2), nitric oxide (NO) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activity at 0.275, 0.5 and 5microM of oxime, respectively (p< or =0.05). The oxime exhibited a powerful inhibitory effect on dihydroxybenzoate formation (25microM) (p< or =0.05) and also decreased deoxyribose degradation induced by Fe2+ and via Fenton reaction (0.44 and 0.66mM, respectively) (p< or =0.05). The oxime showed a significant inhibitory effect on sigma-phenantroline reaction with Fe2+ (0.4mM) suggesting a possible interaction between the oxime and iron. A significant decrease in the basal and pro-oxidant-induced lipid peroxidation in brain, liver, and kidney of mice was observed both in vitro and ex vivo (p< or =0.05). In addition, in our ex vivo experiments the oxime did not depict any significant changes in thiol levels of liver, kidney and brain as well as did not modify the delta-aminolevulinate dehydratase (delta-ALA-D) activity in these tissues. Taken together our results indicate an in vitro and ex vivo antioxidant activity of the oxime possibly due to its scavenging activity toward different RS and a significant iron interaction.