Oxidative DNA damage induced by equine estrogen metabolites: role of estrogen receptor alpha.
Department of Medicinal Chemistry and Pharmacognosy (M/C 781), College of Pharmacy, The University of Illinois at Chicago, 833 South Wood Street, Chicago, Illinois 60612, USA.Chemical Research in Toxicology (Impact Factor: 3.53). 05/2002; 15(4):512-9.
Excessive exposure to synthetic and endogenous estrogens has been associated with the development of cancer in several tissues. 4-Hydroxyequilenin (4-OHEN), a major metabolite of equine estrogens present in estrogen replacement formulations, has been shown to induce cytotoxic/carcinogenic effects. In the present study, we have found that 4-OHEN caused DNA damage in breast cancer cells, and cells that contain estrogen receptor alpha (S30) are more sensitive to 4-OHEN-mediated DNA damage as compared to estrogen receptor negative cells (MDA-MB-231). For example, concentration-dependent increases in 8-oxo-deoxyguanosine (8-oxo-dG), as measured by LC-MS-MS or by the Fpg comet assay, were only detected in the S30 cells, and the amount of this lesion could be enhanced by agents, which catalyze redox cycling (NADH) or deplete GSH (diethyl maleate). The role of the estrogen receptor in modulating DNA damage was further established in incubations with the ER antagonist tamoxifen, where decreases in 8-oxo-deoxyguanosine were observed. Another equine estrogen metabolite, 4,17 beta-hydroxyequilenin (4,17 beta-OHEN), was found to have the same cytotoxicity and a similar ability to induce reactive oxygen species (ROS), and caused the same oxidative DNA damage in S30 cells as compared to 4-OHEN. However, 4,17 beta-OHEN induced twice as much single strand DNA breaks in S30 cells compared to 4-OHEN. Also 4,17 beta-OHEN was more estrogenic than 4-OHEN as demonstrated by a higher binding affinity for ER alpha and an enhanced induction in activity of estrogen-dependent alkaline phosphatase in Ishikawa cells. These data suggest that the mechanism of DNA damage induced by equine estrogen metabolites could involve oxidative stress and that the estrogen receptor may play a role in this process.
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- "Imaging was performed using a fluorescence microscope (Zeiss Axiovert L410 Inc., Jena, Germany) equipped with a CCD camera (Olympus Inc., Tokyo, Japan). The Comets were analysed by a visual scoring method and computer image analysis using the Comet Assay Project Software (CAPS) (Liu et al., 2002). "
ABSTRACT: The primary aim of this study was to determine the potential modulatory role of the apitoxin (bee venom; BV) against propionic acid (PPA)-induced neurotoxicity. The biochemical responses to PPA exposure in rat pups were assayed, including changes in the antioxidant barrier systems and lipid peroxidation and protein oxidation biomarkers in the brain tissue. DNA damage was measured by single-cell gel electrophoresis and differences in Bcl-2 and Caspase-3 mRNA expression were assessed using real-time PCR. Changes in amygdala complex ultrastructure were visually assessed using electron microscopy. Sixty rat pups were assigned into six groups: a control group, a PPA-treated group, a BV-treated group, a protective co-treated group, a therapeutic co-treated group, and a protective/therapeutic co-treated group. The results indicate that PPA induced a pronounced increase (64.6%) in malondialdehyde (MDA), and in DNA damage (73.3%) with three-fold increase in protein carbonyl concentration. A significant reduction was observed in the enzyme activities of superoxide dismutase (SOD) (48.7%) and catalase (CAT) (74.8%) and reduced glutathione (GSH) level (52.6%). BV significantly neutralized the PPA-induced oxidative stress effects, especially in the BV protective/therapeutic co-treated group. In this group, GSH levels were restored to 64.5%, and MDA, protein carbonyl levels and tail moment % were diminished by 69.5, 21.1 and 18.8% relative to the control, respectively. Furthermore, while PPA induced significant apoptotic neural cell death, BV markedly inhibited apoptosis by promoting Bcl-2 expression and blocking Caspase-3 expression. BV markedly restored the normal ultrastructural morphology of the amygdala complex neurons. These results conclusively demonstrate that BV administration provides both protective and therapeutic effects in response to the PPA-induced deleterious effects, including oxidative stress, DNA damage, and neuronal death in the brains of rat pups.NeuroToxicology 06/2015; 49. DOI:10.1016/j.neuro.2015.05.011 · 3.38 Impact Factor
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- "The slides were examined with a 40X objective lens with epi-illuminated fluorescence microscopy (Olympus-Bx60, excitation filter: 515-560 nm; barrier filter: 590 nm) attached to a color CCD video camera and connected to an image analysis system (Comet II, Perspective Instruments, UK). The Comets were analyzed by a visual scoring method and computerized image analysis (Liu et al., 2002). To quantify DNA damage, tail length (TL), tail DNA (%) (TDNA) and tail moment (TM) were analyzed using Comet Assay Project Software (CAPS), generally 50-100 randomly selected cells are analyzed per sample. "
ABSTRACT: The genotoxic parameters are currently among the most valuable fish biomarkers for environmental risk assessment. So, the present study was directed to explore the toxic impacts of Aluminum chloride (AlCl 3) on DNA damage, apoptosis, antioxidant status and ultrastructural investigations of Nile catfish. The experiment was carried out on 48 fish that randomly divided into four equal groups with two replicates; the first group exposed to 1/20 LC 50 of AlCl 3 , the second group exposed to the combined doses of AlCl 3 (1/20 LC 50) and vitamin C (Vit. C) at dose of 5 ppm, the third group exposed to Vit C at the dose previously mentioned and the fourth group was kept as negative control. The experiment was terminated after six months where the fish were sacrificed and specimens from liver and gills of all groups were obtained and kept at -20°C till applying the required measurements and another specimen from the same organs were fixed in 10% neutral –buffered formalin and 3% glutaraldehyde solution for histopathological examination. The results indicated pronounced significant increase in malondialdehyde (MDA) concentration and significant decrease in both reduced glutathione (GSH) concentration and superoxide dismutase (SOD) and catalase (CAT) enzyme activity. AlCl 3 elicited an obvious increase in oxidative DNA damage and frequency of apoptotic cells, these manifestation were markedly ameliorated in the group exposed to the combined doses of AlCl 3 and Vit C. The ultrastructural histopathological findings proved the aforementioned results. It could be concluded that AlCl 3 elucidated a marked ruinous effects on the oxidative and genotoxic impacts as well as the histopathological alterations which were alleviated by Vitamin C.Life Science Journal 11/2012; 9(4). · 0.17 Impact Factor
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- "Each cell (or comet) was assigned a value according to the class it belonged to, so that an overall score was derived for each treatment ranging from 0 to 400 arbitrary units. The following formula (Liu et al., 2002) was used to calculate scores in which N is the number of cells in each category (e.g. N 4 is the number of cells in category 4). "
ABSTRACT: Peroxynitrite formed by the reaction of superoxide and nitric oxide is a highly reactive species with a role in various pathological processes such as cancer, chronic inflammation, and cardiovascular and neurological diseases. In the present study, the effect of the carotenoids, lycopene and beta-carotene, on peroxynitrite-mediated modifications in plasmid DNA as well as cellular DNA and proteins were investigated. In pUC18 plasmid DNA, these carotenoids strongly inhibited DNA strand breaks caused by peroxynitrite generated from 3-morpholinosydnonimine (SIN-1). SIN-1 was also used to determine effects on DNA damage and protein tyrosine nitration in Chinese hamster lung fibroblasts. SIN-1 dose-dependently increased nitration of proteins in cells above basal levels as determined by Western blotting. This nitration was inhibited in the presence of the uric acid as well as lycopene. Physiological concentrations (0.31-10 microM) of lycopene and beta-carotene also had protective effects on DNA damage, as measured by the comet assay. Lycopene significantly reduced DNA damage particularly, in the median range of concentrations (2.5 microM). The protective effects of lycopene and beta-carotene could be due to their scavenging of reactive oxygen (ROS) and/or nitrogen species (RNS) as they reduce the amount of intracellular ROS/RNS produced following treatment with SIN-1 by as much as 47.5% and 42.4%, respectively. The results obtained in this study suggest that carotenoids may alleviate some of the deleterious effects of peroxynitrite and possibly other reactive nitrogen species as well in vivo.Toxicology and Applied Pharmacology 10/2006; 215(3):330-40. DOI:10.1016/j.taap.2006.03.006 · 3.71 Impact Factor
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