RESUMO
Although copper (Cu) is an essential trace metal for cells, it can induce harmful effects as it participates in the Fenton reaction. Involuntary exposure to Cu overload is much more common than expected and has been linked with neurodegeneration, particularly with Alzheimer's disease (AD) evidenced by a positive correlation between free Cu in plasma and the severity of the disease. It has been suggested that Cu imbalance alters cholesterol (Chol) homeostasis and that high membrane Chol promotes the amyloidogenic processing of the amyloid precursor protein (APP) secreting the ß-amyloid (Aß) peptide. Despite the wide knowledge on the effects of Cu in mature brain metabolism, the consequence of its overload on immature neurons remains unknown. Therefore, we used an undifferentiated human neuroblastoma cell line (SH-SY5Y) to analyze the effect of sublethal concentrations of Cu on 1- de novo Chol synthesis and membrane distribution; 2-APP levels in cells and its distribution in membrane rafts; 3-the levels of Aß in the culture medium. Our results demonstrated that Cu increases reactive oxygen species (ROS) and favors Chol de novo synthesis in both ROS-dependent and independent manners. Also, at least part of these effects was due to the activation of 3-hydroxy-3-methyl glutaryl CoA reductase (HMGCR). In addition, Cu increases the Chol/PL ratio in the cellular membranes, specifically Chol content in membrane rafts. We found no changes in total APP cell levels; however, its presence in membrane rafts increases with the consequent increase of Aß in the culture medium. We conclude that Cu overload favors Chol de novo synthesis in both ROS-dependent and independent manners, being at least in part, responsible for the high Chol levels found in the cell membrane and membrane rafts. These may promote the redistribution of APP into the rafts, favoring the amyloidogenic processing of this protein and increasing the levels of Aß.
RESUMO
Mitochondrial dysfunction is a common hallmark in aging. In the female, reproductive senescence is characterized by loss of ovarian hormones, many of whose neuroprotective effects converge upon mitochondria. The functional integrity of mitochondria is dependent on membrane fatty acid and phospholipid composition, which are also affected during aging. The effect of long-term ovarian hormone deprivation upon mitochondrial function and its putative association with changes in mitochondrial membrane lipid profile in the hippocampus, an area primarily affected during aging and highly responsive to ovarian hormones, is unknown. To this aim, Wistar adult female rats were ovariectomized or sham-operated. Twelve weeks later, different parameters of mitochondrial function (O2 uptake, ATP production, membrane potential and respiratory complex activities) as well as membrane phospholipid content and composition were evaluated in hippocampal mitochondria. Chronic ovariectomy reduced mitochondrial O2 uptake and ATP production rates and induced membrane depolarization during active respiration without altering the activity of respiratory complexes. Mitochondrial membrane lipid profile showed no changes in cholesterol levels but higher levels of unsaturated fatty acids and a higher peroxidizability index in mitochondria from ovariectomized rats. Interestingly, ovariectomy also reduced cardiolipin content and altered cardiolipin fatty acid profile leading to a lower peroxidizability index. In conclusion, chronic ovarian hormone deprivation induces mitochondrial dysfunction and changes in the mitochondrial membrane lipid profile comparable to an aging phenotype. Our study provides insights into ovarian hormone loss-induced early lipidomic changes with bioenergetic deficits in the hippocampus that may contribute to the increased risk of Alzheimer's disease and other age-associated disorders observed in postmenopause.
Assuntos
Ácidos Graxos/fisiologia , Hormônios Esteroides Gonadais/deficiência , Hipocampo/metabolismo , Mitocôndrias/metabolismo , Fosfolipídeos/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Feminino , Potencial da Membrana Mitocondrial/fisiologia , Ovariectomia , Consumo de Oxigênio/fisiologia , Ratos , Ratos WistarRESUMO
OBJECTIVE: Brain stroke is the third most important cause of death in developed countries. We studied the effect of different dietary lipids on the outcome of a permanent ischemic stroke rat model. METHODS: Wistar rats were fed diets containing 7% commercial oils (S, soybean; O, olive; C, coconut; G, grape seed) for 35 d. Stroke was induced by permanent middle cerebral artery occlusion. Coronal slices from ischemic brains and sham-operated animals were supravitally stained. Penumbra and core volumes were calculated by image digitalization after 24, 48, and 72 h poststroke. Homogenates and mitochondrial fractions were prepared from different zones and analyzed by redox status, inflammatory markers, ceramide, and arachidonate content, phospholipase A2, NOS, and proteases. RESULTS: Soybean (S) and G diets were mainly prooxidative and proinflammatory by increasing the liberation of arachidonate and its transformation into prostaglandins. O was protective in terms of redox homeostatic balance, minor increases in lipid and protein damage, conservation of reduced glutathione, protective activation of NOS in penumbra, and net ratio of anti-to proinflammatory cytokines. Apoptosis (caspase-3, milli- and microcalpains) was less activated by O than by any other diet. CONCLUSION: Dietary lipids modulate NOS and PLA2 activities, ceramide production, and glutathione import into the mitochondrial matrix, finally determining the activation of the two main protease systems involved in programmed cell death. Olive oil appears to be a biological source for the isolation of protective agents that block the expansion of brain core at the expense of penumbral neurons.
Assuntos
Antioxidantes/uso terapêutico , Encéfalo/efeitos dos fármacos , Gorduras na Dieta , Inflamação/prevenção & controle , Estresse Oxidativo/efeitos dos fármacos , Óleos de Plantas/uso terapêutico , Acidente Vascular Cerebral , Animais , Antioxidantes/farmacologia , Apoptose , Biomarcadores/metabolismo , Encéfalo/citologia , Encéfalo/metabolismo , Isquemia Encefálica/dietoterapia , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patologia , Cocos , Dieta , Gorduras na Dieta/efeitos adversos , Gorduras na Dieta/metabolismo , Gorduras na Dieta/farmacologia , Gorduras na Dieta/uso terapêutico , Inflamação/etiologia , Inflamação/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Masculino , Neurônios , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Óxido Nítrico Sintase/metabolismo , Olea , Oxirredução , Óleos de Plantas/efeitos adversos , Óleos de Plantas/farmacologia , Ratos Wistar , Espécies Reativas de Oxigênio/efeitos adversos , Glycine max , Acidente Vascular Cerebral/dietoterapia , Acidente Vascular Cerebral/etiologia , Acidente Vascular Cerebral/metabolismo , Acidente Vascular Cerebral/patologia , VitisRESUMO
We have previously demonstrated that the sub-chronic administration of low doses of Toc or α-Toc, glyphosate and zineb to rats (i.p. 1/250 LD50, three times a week for 5 weeks) provoked severe oxidative stress (OS) in testicles. These effects were also reflected in plasma. Lipoic acid (LA) and α-tocopherol are considered as antioxidants due to their ability to neutralize reactive oxygenated species (ROS) and reset endogenous antioxidant levels. To investigate the possible protective effect on reproductive function, LA and Toc (i.p. 25, 50 and 100mg/kg) were administered simultaneously with the pesticide mixture (PM) for 5 weeks. Both drugs prevented OS and the damage to proteins and lipids caused by PM in a dose-dependent manner. The PM-induced increase levels of prostaglandins E2 and F2α was completely restored by LA but not by Toc. Similarly, only LA was able to restore the inhibition of testosterone production, the decrease of 3ß- and 17ß-hydroxysteroid dehydrogenases activities, and the elevation of gonatropins (FSH and LH) levels produced by PM. Furthermore, LA was more efficient than Toc in normalizing the histological alterations produced by PM administration, suggesting that pesticides act though other mechanisms that generate oxidative stress. In our experimental model LA displayed a higher protective role against pesticide-induced damage than that observed by Toc administration. Our results suggest that LA administration is a promising therapeutic strategy for coping with disorders suspected to be caused by OS generators - such as pesticides - in male reproductive system.
Assuntos
Antioxidantes/farmacologia , Poluentes Ambientais/toxicidade , Praguicidas/toxicidade , Testículo/efeitos dos fármacos , Ácido Tióctico/farmacologia , Tocoferóis/farmacologia , Animais , Hormônios Esteroides Gonadais/sangue , Masculino , Estresse Oxidativo/efeitos dos fármacos , Ratos , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismoRESUMO
We have previously demonstrated that the administration of low doses of dimethoate, glyphosate and zineb to rats (i.p. 1/250 LD50, three times a week for 5weeks) provokes severe oxidative stress (OS) in specific brain regions: substantia nigra, cortex and hippocampus. These effects were also observed in plasma. Lipoic acid (LA) is considered an "ideal antioxidant" due to its ability to scavenge reactive species, reset antioxidant levels and cross the blood-brain barrier. To investigate its protective effect we administered LA (i.p. 25, 50 and 100mg/kg) simultaneously with the pesticide mixture (PM) for 5weeks. After suppression of PM administration, we evaluated the restorative effect of LA for a further 5weeks. LA prevented OS and the production of nitrites+nitrates [NOx] caused by PM in a dose-dependent manner. The PM-induced decrease in reduced glutathione and α-tocopherol levels in all brain regions was completely restored by LA at both high doses. PM administration also caused an increase in prostaglandins E(2) and F(2α) in brain that was reduced by LA in a dose-dependent fashion. Taking into account the relationship between OS, inflammation and apoptosis, we measured caspase and calpain activity. Only milli- and micro-calpain isoforms were increased in the PM-treated group and LA reduced the activities to basal levels. We also demonstrated that interrupting PM administration is not enough to restore the levels of all the parameters measured and that LA is necessary to achieve basal status. In our experimental model LA displayed a protective role against pesticide-induced damage, suggesting that LA administration is a promising therapeutic strategy to cope with disorders suspected to be caused by OS generators, especially in brain.
Assuntos
Encéfalo/efeitos dos fármacos , Encefalite/prevenção & controle , Estresse Oxidativo/efeitos dos fármacos , Praguicidas/toxicidade , Ácido Tióctico/farmacologia , Animais , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Encéfalo/metabolismo , Encéfalo/patologia , Encefalite/induzido quimicamente , Masculino , Fosfolipases A2/metabolismo , Prostaglandinas/metabolismo , Ratos , Ratos WistarRESUMO
The present work studies the potential restorative effect of polyunsaturated fatty acids (PUFA, 5 µM/24 h) on the dimethoate (DMT)-induced inhibition of testosterone biosynthesis in Leydig cells isolated from rat testes. Various fatty acids (FA) from the n-6 (18:2, 20:3, 20:4, 22:4 and 22:5) and n-3 (18.3, 20:5, 22:5, 22:6) series were assayed in Leydig cells, alone (as delipidated BSA complexes) and in combination with DMT (1 ppm). The n-6 FA stimulated lipid peroxidation (LPO) and inhibited the activities of steroidogenic enzymes (3ß- and 17ß-hydroxysteroid dehydrogenases). The n-3 FA exerted an anti-oxidant effect, decreasing the production of thiobarbituric-acid reactive substances (TBARS) and inhibiting phospholipase A(2) activity. The biosynthesis of testosterone in DMT-treated cultures was completely normalized by ARA (20:4n-6) and partially restored by the addition of 20:3n-6, increasing ARA content inside the mitochondria. The other FA assayed failed to restore androgenesis. COX-2 protein and prostaglandin F2α and E2 production were stimulated by 20:3n-6, ARA, 18:3n-3 and 20:5 n-3. COX-2 protein decreased upon addition of 22:5n-3 and 22:6n-3. StAR protein was increased by ARA and partially increased by 20:3n-6, likely due to its metabolic conversion into ARA. Both FA increased the mitochondrial cholesterol pool available for testosterone biosynthesis. The rate of androgenesis is likely the result of various regulatory factors acting concomitantly on the physiology of Leydig cells.
Assuntos
Ácido Araquidônico/farmacologia , Inibidores da Colinesterase/farmacologia , Dimetoato/farmacologia , Células Intersticiais do Testículo/metabolismo , Testosterona/biossíntese , 17-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , 17-Hidroxiesteroide Desidrogenases/metabolismo , 3-Hidroxiesteroide Desidrogenases/antagonistas & inibidores , 3-Hidroxiesteroide Desidrogenases/metabolismo , Animais , Antioxidantes/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Colesterol , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Ácidos Graxos/farmacologia , Expressão Gênica , Células Intersticiais do Testículo/efeitos dos fármacos , Células Intersticiais do Testículo/enzimologia , Peroxidação de Lipídeos , Masculino , Mitocôndrias/metabolismo , Inibidores de Fosfolipase A2 , Fosfolipases A2/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Prostaglandinas/metabolismo , Ratos , Ratos Wistar , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismoRESUMO
The impact of involuntary exposure to pesticides was studied in a group of professional sprayers (S) (25±5 years old) exposed to various agrochemicals for about 10 years. The results were compared with a group of non exposed people (C). S group showed hematological, renal, pancreatic and hepatic biomarkers within the reference values established for the general population, including cholinesterase activity. In spite of that, all the biochemical tests were statistically different compared to C. On the other hand, oxidative stress biomarkers (OSB) such as plasma tocopherol and the total reducing ability of plasma were significantly decreased, while protein carbonyls, thiobarbituric acid-reactive substances, total glutathione and the sum of nitrites and nitrates were increased in the exposed group. Results demonstrated that screening laboratory tests could not be fully sensitive in detecting sub-clinical exposure to pesticides, and also suggest that OSB could be validated and included in health surveillance protocols.
Assuntos
Biomarcadores/sangue , Exposição Ocupacional , Ocupações , Estresse Oxidativo , Praguicidas/toxicidade , Adulto , Agricultura/métodos , Argentina , Butirilcolinesterase/metabolismo , Feminino , Glutationa/sangue , Humanos , Peroxidação de Lipídeos , Masculino , Distribuição Aleatória , Inquéritos e Questionários , Adulto JovemRESUMO
Copper based-pesticides are widely used in agricultural practice throughout the world. We studied the (i) concentration of Cu and proteins involved in Cu homeostasis, (ii) plasma redox status, and (iii) biomarkers of exposure in Cu-based pesticide applicators in order to compare them with clinical biochemical tests. Thirty-one professional applicators and 32 control subjects were recruited. Oxidative stress biomarkers, ceruloplasmin (CRP), metallothioneins (MTs), copper, hematological parameters, and biochemical markers for pancreatic, hepatic and renal function were measured in plasma. Copper was increased in the exposed group compared to the control group concomitantly with TBARS, protein carbonyls, and nitrate+nitrite levels. In the exposed group, α-tocopherol and the FRAP assay were lower and LDH, transaminases, GGT, ALP, urea, creatinine, CRP and MTs were higher than in the control group. The relative leukocyte subclasses were also different between the two groups. Clinical chemistry tests did not surpass the upper reference limit. Our results suggest that the incorporation of oxidative stress biomarkers to biochemical/clinical tests should be considered for validation and included in the human health surveillance protocols.
Assuntos
Cobre/toxicidade , Exposição Ocupacional/análise , Estresse Oxidativo , Praguicidas/toxicidade , Adulto , Agricultura , Biomarcadores/sangue , Proteína C-Reativa/metabolismo , Ceruloplasmina/metabolismo , Testes de Química Clínica , Cobre/sangue , Creatinina/sangue , Feminino , Humanos , Masculino , Metalotioneína/sangue , Exposição Ocupacional/efeitos adversos , Exposição Ocupacional/estatística & dados numéricos , Praguicidas/sangue , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Transaminases/sangue , alfa-Tocoferol/metabolismoRESUMO
The mechanism involved in the inhibition of testosterone (Te) biosynthesis after a sub-chronic exposure to low doses of dimethoate (D) was studied in rat interstitial cells (IC). Expression of COX-2 in IC isolated from D-treated rats increased by 44% over C data, while transcription of StAR decreased by approx. 50% and the expression of this protein was diminished by approximately 40%. PGE(2) and PGF(2alpha) were increased by 61 and 78%, respectively. Te concentration decreased by 49% in IC homogenates. Concomitantly, plasma concentration of LH and FSH both increased. Araquidonate (ARA) and C(22) fatty acyl chains in phospholipids from IC mitochondrial fraction decreased by approx. 30% after D treatment. Protein carbonyls, lipoperoxides and nitrite content increased while alpha-tocopherol and the antioxidant capacity of the soluble cellular fraction decreased significantly. Stimulation with h-CG 10 nM overnight failed to overcome the inhibition caused by D on both Te biosynthesis and 3beta- and 17beta-hydroxysteroid dehydrogenases. Decreased Te biosynthesis may be attributed to (1) inhibition of StAR protein activity due to the stimulation of COX-2 and the overproduction of PGF(2alpha), (2) decreased stimulatory effect of ARA on StAR with a subsequent reduction in the availability of CHO for the androgenic pathway, and/or (3) indirect inhibition of steroidogenic enzymes by a lower transcriptional rate caused by elevated PGF(2alpha). Rofecoxib administration prevents the deleterious effect(s) exerted by D.