RESUMO
The aim of this study was to evaluate the time course of oxidative stress markers and inflammatory mediators in human conjunctival epithelial cells (IOBA-NHC) exposed to diesel exhaust particles (DEP) for 1, 3, and 24â¯h. Reactive oxygen species (ROS) production, lipid and protein oxidation, Nrf2 pathway activation, enzymatic antioxidants, glutathione (GSH) levels and synthesis, as well as cytokine release and cell proliferation were analyzed. Cells exposed to DEP showed an increase in ROS at all time points. The induction of NADPH oxidase-4 appeared later than mitochondrial superoxide anion production, when the cell also underwent a proinflammatory response mediated by IL-6. DEP exposure triggered the activation of Nrf2 in IOBA-NHC, as a strategy for increasing cellular antioxidant capacity. Antioxidant enzyme activities were significantly increased at early stages except for glutathione reductase (GR) that showed a significant decrease after a 3-h-incubation. GSH levels were found increased after 1 and 3â¯h of incubation with DEP, despite the increase in its consumption by the antioxidant enzymes as it works as a cofactor. GSH recycling and the de novo synthesis were responsible for the maintenance of its content at these time points, respectively. After 24â¯h, the decrease in GR and glutamate cysteine ligase as wells as the enhanced activity of glutathione peroxidase and glutathione S-transferase produced a depletion in the GSH pool. Lipid-peroxidation was found increased in cells exposed to DEP after 1-h-incubation, whereas protein oxidation was found increased in cells exposed to DEP after a 3-h-incubation that persisted after a longer exposure. Furthermore, DEP lead IOBA-NHC cells to hyperplasia after 1 and 3â¯h of incubation, but a decrease in cell proliferation was found after longer exposure. ROS production seems to be an earlier event triggered by DEP on IOBA-NHC, comparing to the proinflammatory response mediated by IL-6. Despite the fact that under short periods of exposure to DEP lipids and then proteins are targets of oxidative damage, the viability of the cells is not affected at early stages, since cell hyperplasia was detected as compensatory mechanism. Although after 24â¯h Nrf2 pathway is still enhanced, the epithelial cell capacity to maintain redox balance is exceeded. The antioxidant enzymes activation and the depleted GSH pool are not capable of counteracting the increased ROS production, leading to oxidative damage.
Assuntos
Poluentes Atmosféricos/toxicidade , Túnica Conjuntiva/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Interleucina-6/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Emissões de Veículos/toxicidade , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Túnica Conjuntiva/metabolismo , Células Epiteliais/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Glutationa/metabolismo , Glutationa Redutase/metabolismo , Glutationa Transferase/metabolismo , Humanos , Peroxidação de Lipídeos , Potenciais da Membrana/fisiologia , Mitocôndrias/metabolismo , NADPH Oxidase 4/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Oxirredução/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Peroxidase/metabolismo , Superóxidos/metabolismoRESUMO
PURPOSE: The aim of this study was to evaluate oxidative stress markers in human conjunctival epithelial cells (IOBA-NHC) exposed to diesel exhaust particles (DEP). METHODS: Reactive oxygen (ROS) and nitrogen (RNS) species production; hydrogen peroxide (H2O2) levels; protein oxidation; antioxidant enzymes activities (superoxide dismutase [SOD], catalase [CAT], glutathione peroxidase [GPx], glutathione S-transferase [GST], and glutathione reductase [GR]); total reactive antioxidant potential (TRAP); reduced (GSH) and oxidized glutathione (GSSG) were evaluated. Transmission electron microscopy was performed to evaluate DEP uptake. RESULTS: Diesel exhaust particles were entrapped by membrane protrusions developed by IOBA-NHC. Cells exposed to DEP 50 and 100 µg/mL showed a significant increase in ROS, RNS, H2O2 levels, SOD, GPx, and GST compared with the control group. A significant decay in GR was observed in both groups, meanwhile CAT levels remained unchanged. The group exposed to DEP 100 µg/mL displayed a significant increase in protein oxidation. In both groups, TRAP was significantly reduced as well as the GSH/GSSG ratio. CONCLUSIONS: The decrease in nonenzymatic antioxidants and the compensatory increase of SOD, GPX, and GST activities are consequence of the increase in ROS and RNS production due to DEP exposure and its accumulation inside the cells. The decay in GR activity leads to the decrease in GSH/GSSG recycling. These results suggest that oxidative stress could play an important role in the development of DEP effects on human conjunctival epithelial cells.
Assuntos
Túnica Conjuntiva/metabolismo , Células Epiteliais/metabolismo , Estresse Oxidativo/fisiologia , Espécies Reativas de Oxigênio/metabolismo , Emissões de Veículos , Biomarcadores/metabolismo , Células Cultivadas , Túnica Conjuntiva/efeitos dos fármacos , Túnica Conjuntiva/ultraestrutura , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/ultraestrutura , Humanos , Microscopia Eletrônica de TransmissãoRESUMO
Sulforaphane is a naturally occurring isothiocyanate capable of stimulating cellular antioxidant defenses and inducing phase 2 detoxifying enzymes, which can protect cells against oxidative damage. Oxidative stress and apoptosis are intimately involved in the pathophysiology of cardiac diseases. Although sulforaphane is known for its anticancer benefits, its role in cardiac cells is just emerging. The aim of the present study was to investigate whether sulforaphane can modulate oxidative stress, apoptosis, and correlate with PGC-1α, a transcriptional cofactor involved in energy metabolism. H9c2 cardiac myoblasts were incubated with R-sulforaphane 5 µmol/L for 24 h. Cell viability, ANP gene expression, oxidative stress and apoptosis markers, and protein expression of PGC-1α were studied. In cells treated with sulforaphane, cellular viability increased (12 %) and ANP gene expression decreased (46 %) compared to control cells. Moreover, sulforaphane induced a significant increase in superoxide dismutase (103 %), catalase (101 %), and glutathione S-transferase (72 %) activity, reduced reactive oxygen species levels (15 %) and lipid peroxidation (65 %), as well as stimulated the expression of the cytoprotective enzyme heme oxygenase-1 (4-fold). Sulforaphane also promoted an increase in the expression of the anti-apoptotic protein Bcl-2 (60 %), decreasing the Bax/Bcl-2 ratio. Active Caspase 3\7 and p-JNK/JNK were also reduced by sulforaphane, suggesting a reduction in apoptotic signaling. This was associated with an increased protein expression of PGC-1α (42 %). These results suggest that sulforaphane offers cytoprotection to cardiac cells by activating PGC1-α, reducing oxidative stress, and decreasing apoptosis signaling.
Assuntos
Antioxidantes/farmacologia , Isotiocianatos/farmacologia , Mioblastos Cardíacos/efeitos dos fármacos , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo , Animais , Apoptose , Fator Natriurético Atrial/genética , Fator Natriurético Atrial/metabolismo , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Mioblastos Cardíacos/fisiologia , Estresse Oxidativo/efeitos dos fármacos , Coativador 1-alfa do Receptor gama Ativado por Proliferador de Peroxissomo , Ratos , Transdução de Sinais/efeitos dos fármacos , SulfóxidosRESUMO
Long-term intake of aspartame at the acceptable daily dose causes oxidative stress in rodent brain mainly due to the dysregulation of glutathione (GSH) homeostasis. N-Acetylcysteine provides the cysteine that is required for the production of GSH, being effective in treating disorders associated with oxidative stress. We investigated the effects of N-acetylcysteine treatment (150 mg kg(-1), i.p.) on oxidative stress biomarkers in rat brain after chronic aspartame administration by gavage (40 mg kg(-1)). N-Acetylcysteine led to a reduction in the thiobarbituric acid reactive substances, lipid hydroperoxides, and carbonyl protein levels, which were increased due to aspartame administration. N-Acetylcysteine also resulted in an elevation of superoxide dismutase, glutathione peroxidase, glutathione reductase activities, as well as non-protein thiols, and total reactive antioxidant potential levels, which were decreased after aspartame exposure. However, N-acetylcysteine was unable to reduce serum glucose levels, which were increased as a result of aspartame administration. Furthermore, catalase and glutathione S-transferase, whose activities were reduced due to aspartame treatment, remained decreased even after N-acetylcysteine exposure. In conclusion, N-acetylcysteine treatment may exert a protective effect against the oxidative damage in the brain, which was caused by the long-term consumption of the acceptable daily dose of aspartame by rats.
Assuntos
Acetilcisteína/farmacologia , Aspartame/administração & dosagem , Encéfalo/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Animais , Biomarcadores/metabolismo , Glicemia/análise , Peso Corporal , Encéfalo/metabolismo , Masculino , Ratos , Ratos WistarRESUMO
PURPOSE: The goal of the present study is to establish the antioxidant status in the brain of a high pressure-induced rat model. METHODS: Ocular hypertension was induced in rats (n = 12) cauterizing two episcleral veins under a surgical microscope. A sham procedure (n = 12) was performed in the control group. The markers evaluated in the brain 7 days after surgery were as follows: spontaneous chemiluminescence, protein carbonylation, nitrite concentration, total reactive antioxidant potential (TRAP), ascorbic acid, glutathione, vitamin E and activities of superoxide dismutase (SOD), glutathione peroxidase (GPx) and catalase. RESULTS: Chemiluminescence in glaucoma was 55% higher than in controls (393 ± 20 cpm/mg protein, p < 0.001). Protein carbonylation in glaucoma was 93% higher than in controls (1.15 ± 0.18 nmol/mg protein, p < 0.001). Nitrite concentration was 5.30 ± 0.25 µM for glaucoma (controls 4.41 ± 0.24 µM, p < 0.05). Total reactive antioxidant potential decreased by 42% in glaucoma (controls 153 ± 14 µM Trolox, p < 0.001). Ascorbic acid was 67 ± 26 µM for glaucoma (controls 275 ± 22 µM, p < 0.001). Vitamin E was 0.58 ± 0.05 µmol/g organ for glaucoma (controls 1.10 ± 0.06 µmol/g organ, p < 0.01). Glutathione was 1.98 ± 0.13 µmol/g organ for glaucoma (controls 8.19 ± 0.71 µmol/g organ, p < 0.001). Superoxide dismutase and GPx were increased in glaucoma by 42 and 59%, respectively (p < 0.05). CONCLUSIONS: Reactive oxygen and nitrogen species were increased in glaucoma, the increase in chemiluminescence, protein carbonylation and nitrite levels could be evidenced by this situation. The decrease in nonenzymatic antioxidants and a compensatory increase in SOD and GPx activity may have been a consequence of an increase in oxidative processes.
Assuntos
Antioxidantes/metabolismo , Encéfalo/metabolismo , Modelos Animais de Doenças , Pressão Intraocular , Hipertensão Ocular/metabolismo , Estresse Oxidativo , Animais , Ácido Ascórbico/metabolismo , Catalase/metabolismo , Feminino , Glutationa/metabolismo , Glutationa Peroxidase/metabolismo , Luminescência , Carbonilação Proteica , Ratos , Ratos Wistar , Espécies Reativas de Nitrogênio/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Superóxido Dismutase/metabolismo , Vitamina E/metabolismoRESUMO
The objective of this study was to evaluate the effect of chronic thorium (Th) exposure on bioaccumulation, metabolism (through biochemical parameters of the muscle) and oxidative parameters (lipidic peroxidation levels and antioxidant enzymes in the gills and in the hepatic and muscular tissues) of silver catfish (Rhamdia quelen). Silver catfish juveniles were exposed to different waterborne Th levels (in microg L(-1)): 0 (control), 25.3+/-3.2, 80.6+/-12.0, 242.4+/-35.6, and 747.2+/-59.1 for 30 d. The gills and skin were the organs that accumulated the highest Th levels. The increase in the waterborne Th concentration corresponded to a progressive increase in the Th levels in the gills and kidney. Chronic Th exposure causes alterations in the oxidative parameters of silver catfish gills, which are correlated with the Th accumulation in this organ. The levels of GST decreased in the gills of fish exposed to 747.2 microg L(-1) Th and SOD activity decreased in silver catfish exposed to 242.4 and 747.2 microg L(-1) Th. In addition, the increase in the LPO in the gills exposed to 242.4 and 747.2 microg L(-1) Th suggests that higher oxidative damage occurred in the gills. However, in the liver and muscle, these alterations occurred mainly in the lowest waterborne Th level. Metabolic intermediates in the muscle were altered by Th exposure, but no clear relationship was found.
Assuntos
Peixes-Gato/metabolismo , Tório/metabolismo , Tório/toxicidade , Poluentes Químicos da Água/metabolismo , Poluentes Químicos da Água/toxicidade , Amônia/metabolismo , Animais , Biomarcadores/metabolismo , Proteínas de Peixes/metabolismo , Brânquias/metabolismo , Glucose/metabolismo , Glicogênio/metabolismo , Ácido Láctico/metabolismo , Fígado/metabolismo , Estresse OxidativoRESUMO
Dehydroepiandrosterone (DHEA) is an endogenous steroid synthesized mainly in the adrenal cortex. It is known that DHEA is a precursor of sex steroids and that part of its effects depends on its conversion to estrogens and androgens. Sex steroids exert profound and controversial effects on cardiovascular function. Exogenous DHEA can exert a dual effect, antioxidant or prooxidant, depending on the dose and on the tissue specificity [1,2] (F. Celebi, I. Yilmaz, H. Aksoy, M. Gümüs, S. Taysi, D. Oren, Dehydroepiandrosterone prevents oxidative injury in obstructive jaundice in rats, J. Int. Med. Res. 32 (4) (2004) 400-405; S.K. Kim, R.F. Novak, The role of intracellular signaling in insulin-mediated regulation of drug metabolizing enzyme gene and protein expression, Pharmacol. Ther. 113 (1) (2007) 88-120). Akt signaling pathway is one of the anti-proliferative mechanisms of DHEA (Y. Jiang, T. Miyazaki, A. Honda, T. Hirayama, S. Yoshida, N. Tanaka, Y. Matsuzaki, Apoptosis and inhibition of the phosphatidylinositol 3-kinase/Akt signaling pathway in the anti-proliferative actions of dehydroepiandrosterone, J. Gastroenterol. 40 (5) (2005) 490-497). Heart homogenates were prepared to quantify lipid peroxidation (LPO), concentration of superoxide dismutase (SOD), catalase (CAT), glutathione-S-transferase (GST), 4-hydroxy-2-nonenal (HNE) and p-Akt/Akt ratio, and the activities of those antioxidant enzymes. When administrated to male Wistar rats in short-term (6 or 24h) intraperitoneally, DHEA produced significant differences in some parameters of oxidative stress in rat hearts among the distinct doses (1, 10, and 50mg/kg) used. The results here presented, regarding 6 and 24h oxidative stress status, have shown that DHEA injections promoted a prooxidant answer in healthy Wistar rat hearts.
Assuntos
Catalase/metabolismo , Sulfato de Desidroepiandrosterona/metabolismo , Miocárdio/metabolismo , Proteínas Proto-Oncogênicas c-akt/fisiologia , Superóxido Dismutase/metabolismo , Animais , Sulfato de Desidroepiandrosterona/farmacologia , Ativação Enzimática , Peroxidação de Lipídeos , Masculino , Estresse Oxidativo , Ratos , Ratos Wistar , Transdução de SinaisRESUMO
The objective of this study was to evaluate the effect of thorium (Th) bioaccumulation on the metabolism of silver catfish (Rhamdia quelen) through biochemical parameters of the muscle (glycogen, glucose, lactate, protein, and ammonia). In addition, lipidic peroxidation levels (TBARS), catalase (CAT) and glutathione-S-transferase (GST) in the gills and in hepatic and muscular tissues were also analyzed. Cytogenetic parameters were studied through the evaluation of nuclear abnormalities in red blood cells. Silver catfish juveniles were exposed to different waterborne Th levels (in microg L(-1)): 0 (control), 25.3+/-3.2, 69.2+/-2.73, 209.5+/-17.6, and 608.7+/-61.1 for 15 days. The organs that accumulated the highest Th levels were the gills and skin. The increase of waterborne Th concentration corresponded to a progressive increase of Th levels in the gills, liver, skin and kidneys, with the highest accumulation in the gills and skin. Metabolic intermediates in the muscle were altered by Th exposure, but no clear relationship was found. CAT and GST activities in the hepatic and muscular tissues of this species suggest that the enzymatic activities can be stimulated at the lowest Th levels and inhibited at the higher levels (mainly in 608.7 microg L(-1)). The results of the cytogenetic assay contribute to this hypothesis because the higher toxicity in blood samples was found in juveniles exposed to 69.2 and 209.5 microg L(-1) Th.
Assuntos
Peixes-Gato/metabolismo , Músculo Esquelético/metabolismo , Tório/farmacocinética , Amônia/metabolismo , Análise de Variância , Animais , Catalase/metabolismo , Relação Dose-Resposta a Droga , Glucose/metabolismo , Glutationa Transferase/metabolismo , Glicogênio/metabolismo , Ácido Láctico/metabolismo , Testes para Micronúcleos , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo , Tório/sangueRESUMO
PURPOSE: Oxidative stress and antioxidant status in eye tissues may be associated with glaucomatous damage. The aim of this study was to establish the antioxidant status of aqueous humor of patients with primary open-angle glaucoma. For this purpose the authors measured the total reactive antioxidant potential (TRAP) and the activities of the antioxidant enzymes superoxide dismutase (SOD), catalase, and glutathione peroxidase. DESIGN: Case control study. METHODS: Aqueous humor was obtained at the time of surgery from 24 patients with glaucoma and 24 cataract patients; TRAP was measured by chemiluminescence. Activities of the antioxidant enzymes were measured spectrophotometrically. Superoxide dismutase activity was determined by inhibition of the rate of adrenochrome formation at 480 nm. Catalase activity was evaluated by decrease of H(2)O(2) absorbance at 240 nm. Glutathione peroxidase (GPx) activity was determined following nicotinamide adenine dinucleotide phosphate oxidation at 340 nm. RESULTS: Total reactive antioxidant potential value of the cataract group was 124 +/- 5 micromol/l Trolox. This value was significantly decreased, by 64%, in glaucoma patients. An increase of 57% in SOD activity was observed in glaucoma patients when compared with cataract patients (41.7 +/- 2.7 U SOD/ml). Glutathione activity was threefold higher in glaucoma patients than in the cataract group (6.1 +/- 0.6 U/ml). No significant changes were found in catalase levels. CONCLUSIONS: Oxidative stress may lead to an induction of antioxidant enzymes and contribute to TRAP decrease. Superoxide dismutase, GPx activities, and TRAP may be useful oxidative stress markers in aqueous humor of glaucoma patients.
Assuntos
Humor Aquoso/enzimologia , Catalase/metabolismo , Glaucoma de Ângulo Aberto/enzimologia , Glutationa Peroxidase/metabolismo , Estresse Oxidativo , Superóxido Dismutase/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Antioxidantes/metabolismo , Biomarcadores/análise , Estudos de Casos e Controles , Catarata/enzimologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos ProspectivosRESUMO
OBJECTIVE: To evaluate the oxidative state in patients with familial amyloidotic polyneuropathy type 1 (FAP1). DESIGN: From 3 unrelated families, patients with FAP1 carrying a transthyretin Met-30 mutation were studied. The diagnosis was confirmed by genetic analysis. Eleven of 21 patients carried the mutation; all were symptomatic and were clinically assessed using a clinical score. All of the patients were evaluated for copper-zinc superoxide dismutase type 1 activity in red blood cells using spectrophotometry. Plasma total reactive antioxidant potential was studied using a chemiluminescent method. The results were compared with those obtained from an age-matched control group. SETTING: A public and academic multidisciplinary research clinic. RESULTS: Six of the 11 FAP1-positive patients disclosed superoxide dismutase type 1 activity values greater than 55 U/mg of protein (upper control limit), whereas 9 of 10 patients in whom total reactive antioxidant potential was measured had values below the lower limit of the control group. No relationship was found between the levels of superoxide dismutase type 1 activity and the severity of the clinical involvement. CONCLUSIONS: Oxidative stress may be part of the mechanisms leading to tissue damage in patients with FAP1. The lack of correlation between the laboratory findings and the severity of clinical involvement may signal that oxidative processes are at work throughout the natural history of the disease.