Oxidative stress and apoptosis induced by titanium dioxide nanoparticles in cultured BEAS-2B cells

As the applications of industrial nanoparticles are being developed, the concerns on the environmental health are increasing. Cytotoxicities of titanium dioxide nanoparticles of different concentrations (5, 10, 20 and 40mug/ml) were evaluated in this study using a cultured human bronchial epithelial cell line, BEAS-2B. Exposure of the cultured cells to nanoparticles led to cell death, reactive oxygen species (ROS) increase, reduced glutathione (GSH) decrease, and the induction of oxidative stress-related genes such as heme oxygenase-1, thioredoxin reductase, glutathione-S-transferase, catalase, and a hypoxia inducible gene. The ROS increase by titanium dioxide nanoparticles triggered the activation of cytosolic caspase-3 and chromatin condensation, which means that titanium dioxide nanoparticles exert cytotoxicity by an apoptotic process. Furthermore, the expressions of inflammation-related genes such as interleukin-1 (IL-1), interleukin-6 (IL-6), interleukin-8 (IL-8), TNF-a, and C-X-C motif ligand 2 (CXCL2) were also elevated. The induction of IL-8 by titanium dioxide nanoparticles was inhibited by the pre-treatment with SB203580 and PD98059, which means that the IL-8 was induced through p38 mitogen-acitvated protein kinase (MAPK) pathway and/or extracellular signal (ERK) pathway. Uptake of the nanoparticles into the cultured cells was observed and titanium dioxide nanoparticles seemed to penetrate into the cytoplasm and locate in the peri-region of the nucleus as aggregated particles, which may induce direct interactions between the particles and cellular molecules, to cause adverse biological responses.     

药用纳米SiO2对人正常肺细胞的氧化损伤

目的：探讨药用纳米SiO2对人正常肺细胞MRC-5的生长抑制与氧化损伤作用。方法：纳米SiO2暴露于MRC-5细胞48h后,以MTT法测定其对细胞增殖的影响,HE染色观察细胞的形态学变化,并检测暴露后细胞内活性氧（ROS）和还原型谷胱甘肽（GSH）含量以及超氧化物歧化酶（SOD）活性的改变,分析纳米材料对MRC-5细胞的氧化损伤作用。结果：两种尺度（粒径21.6、48.6nm）纳米SiO2暴露浓度分别达到0.4mg/mL与1.0mg/mL以上时,细胞存活率随暴露剂量的增加而降低,IC50分别为0.8mg/mL和1.9mg/mL。细胞形态皱缩,核质凝聚。细胞内活性氧明显升高（P〈0.01）,GSH含量和SOD活性显著降低（P〈0.05）,且呈现明显的剂量效应关系。结论：较高浓度纳米SiO2直接暴露可抑制人正常肺细胞MRC-5的增殖,其机理与细胞的氧化损伤有关。     

The size‐dependent genotoxic potentials of titanium dioxide nanoparticles to endothelial cells

Despite intensive research activities, there are still many major knowledge gaps over the potential adverse effects of titanium dioxide nanoparticles (TiO₂‐NPs), one of the most widely produced and used nanoparticles, on human cardiovascular health and the underlying mechanisms. In the present study, alkaline comet assay and cytokinesis‐block micronucleus test were employed to determine the genotoxic potentials of four sizes (100, 50, 30, and 10 nm) of anatase TiO₂‐NPs to human umbilical vein endothelial cells (HUVECs) in culture. Also, the intracellular redox statuses were explored through the measurement of the levels of reactive oxygen species (ROS) and reduced glutathione (GSH) with kits, respectively. Meanwhile, the protein levels of nuclear factor erythroid 2‐related factor 2 (Nrf2) were also detected by western blot. The results showed that at the exposed levels (1, 5, and 25 μg/mL), all the four sizes of TiO₂‐NPs could elicit an increase of both DNA damage and MN frequency in HUVECs in culture, with a positive dose‐dependent and negative size‐dependent effect relationship (T100 < T50 < T30 < T10). Also, increased levels of intracellular ROS, but decreased levels of GSH, were found in all the TiO₂‐NP‐treated groups. Intriguingly, a very similar manner of dose‐dependent and size‐dependent effect relationship was observed between the ROS test and both comet assay and MN test, but contrary to that of GSH assay. Correspondingly, the levels of Nrf2 protein were also elevated in the TiO₂‐NP‐exposed HUVECs, with an inversely size‐dependent effect relationship. These findings indicated that induction of oxidative stress and subsequent genotoxicity might be an important biological mechanism by which TiO₂‐NP exposure would cause detrimental effects to human cardiovascular health.     

ROS-mediated genotoxicity induced by titanium dioxide nanoparticles in human epidermal cells

Titanium dioxide nanoparticles (TiO₂ NPs) are among the top five NPs used in consumer products, paints and pharmaceutical preparations. Since, exposure to such nanoparticles is mainly through the skin and inhalation, the present study was conducted in the human epidermal cells (A431). A mild cytotoxic response of TiO₂ NPs was observed as evident by the MTT and NR uptake assays after 48 h of exposure. However, a statistically significant (p < 0.05) induction in the DNA damage was observed by the Fpg-modified Comet assay in cells exposed to 0.8 μg/ml TiO₂ NPs (2.20 ± 0.26 vs. control 1.24 ± 0.04) and higher concentrations for 6 h. A significant (p < 0.05) induction in micronucleus formation was also observed at the above concentration (14.67 ± 1.20 vs. control 9.33 ± 1.00). TiO₂ NPs elicited a significant (p < 0.05) reduction in glutathione (15.76%) with a concomitant increase in lipid hydroperoxide (60.51%; p < 0.05) and reactive oxygen species (ROS) generation (49.2%; p < 0.05) after 6 h exposure. Our data demonstrate that TiO₂ NPs have a mild cytotoxic potential. However, they induce ROS and oxidative stress leading to oxidative DNA damage and micronucleus formation, a probable mechanism of genotoxicity. This is perhaps the first study on human skin cells demonstrating the cytotoxic and genotoxic potential of TiO₂ NPs.     

Evaluation of cytotoxic,genotoxic and inflammatory response in human alveolar and bronchial epithelial cells exposed to titanium dioxide nanoparticles

The toxicity of titanium dioxide nanoparticles (TiO₂‐NPs), used in several applications, seems to be influenced by their specific physicochemical characteristics. Cyto‐genotoxic and inflammatory effects induced by a mixture of 79% anatase/21% rutile TiO₂‐NPs were investigated in human alveolar (A549) and bronchial (BEAS‐2B) cells exposed to 1–40 µg ml^–1 30 min, 2 and 24 h to assess potential pulmonary toxicity. The specific physicochemical properties such as crystallinity, NP size and shape, agglomerate size, surface charge and specific surface area (SSA) were analysed. Cytotoxic effects were studied by evaluating cell viability using the WST1 assay and membrane damage using LDH analysis. Direct/oxidative DNA damage was assessed by the Fpg‐comet assay and the inflammatory potential was evaluated as interleukin (IL)‐6, IL‐8 and tumour necrosis factor (TNF)‐α release by enzyme‐linked immunosorbant assay (ELISA). In A549 cells no significant viability reduction and moderate membrane damage, only at the highest concentration, were detected, whereas BEAS‐2B cells showed a significant viability reduction and early membrane damage starting from 10 µg ml^–1. Direct/oxidative DNA damage at 40 µg ml^–1 and increased IL‐6 release at 5 µg ml^–1 were found only in A549 cells after 2 h. The secretion of pro‐inflammatory cytokine IL‐6, involved in the early acute inflammatory response, and oxidative DNA damage indicate the promotion of early and transient oxidative‐inflammatory effects of tested TiO₂‐NPs on human alveolar cells. The findings show a higher susceptibility of normal bronchial cells to cytotoxic effects and higher responsiveness of transformed alveolar cells to genotoxic, oxidative and early inflammatory effects induced by tested TiO₂‐NPs. This different cell behaviour after TiO₂‐NPs exposure suggests the use of both cell lines and multiple end‐points to elucidate NP toxicity on the respiratory system. Copyright © 2014 John Wiley & Sons, Ltd.     

The role of oxidative stress in Sudan IV‐induced DNA damage in human liver‐derived HepG2 cells

Objectives: We evaluated the role of oxidative stress in Sudan IV‐induced DNA damage, using human liver‐derived HepG2 cells. Methods: The DNA damaging effects of Sudan IV in HepG2 cells were evaluated by alkaline single cell gel electrophoresis assay and micronucleus test (MNT). To clarify the underlying mechanisms, we monitored the intracellular generation of reactive oxygen species (ROS) by 2, 7‐dichlorofluorescein diacetate assay and the level of oxidative DNA damage by immunoperoxidase staining for 8‐hydroxydeoxyguanosine (8‐OHdG). Furthermore, the intracellular glutathione (GSH) level was moderated by pretreatment with buthionine‐(S,R)‐sulfoximine (BSO), a specific GSH synthesis inhibitor. Results: A significant dose‐dependent increment in DNA migration was detected at all tested concentrations (25–100 μM) of Sudan IV. And in the MNT, a significant increase of the frequency of micronuclei was found at higher tested concentrations (50–100 μM). The data suggested that Sudan IV caused DNA strand breaks and chromosome breaks. In addition, significantly increased levels of ROS, 8‐OHdG formation were observed in HepG2 cells. It was also found that depletion of GSH in HepG2 cells with BSO dramatically increased the susceptibility of HepG2 cells to Sudan IV‐induced DNA damage. Conclusions: Based on these data we believe that Sudan IV exerts toxic effects in HepG2 cells, probably through oxidative DNA damage induced by intracellular ROS and depletion of GSH. © 2009 Wiley Periodicals, Inc. Environ Toxicol, 2011.     

Metabolic profiling reveals disorder of carbohydrate metabolism in mouse fibroblast cells induced by titanium dioxide nanoparticles

As titanium dioxide (TiO₂) nanoparticles are widely used commercially, their potential biosafety and metabolic mechanism needs to be fully explained. In this study, the cytotoxicity of homogeneous and weakly aggregated (< 100 nm) TiO₂ nanoparticles was investigated by analyzing the changes in metabolite profiles both in mouse fibroblast (L929) cells and their corresponding culture media using gas chromatograph with a time‐of‐flight mass spectrometry (GC/TOFMS)‐based metabolomic strategy. With multivariate statistics analysis, satisfactory separations were observed in principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS‐DA) models. Based on the variable importance in the OPLS‐DA models, a series of differential metabolites were identified by comparison between TiO₂ nanoparticle‐treated L929 cells or their corresponding culture media and the control groups. It was found that the major biochemical metabolism (carbohydrate metabolism) was suppressed in TiO₂ nanoparticle‐treated L929 cells and their corresponding culture media. These results might account for the serious damage to energy metabolism in mitochondria and the increased cellular oxidation stress in TiO₂ nanoparticle‐induced L929 cells. These results also suggest that the metabolomic strategy had a great potential in evaluating the cytotoxicity of TiO₂ nanoparticles and thus was very helpful in understanding its underlying molecular mechanisms. Copyright © 2012 John Wiley & Sons, Ltd.     

The role of oxidative stress in citreoviridin‐induced DNA damage in human liver‐derived HepG2 cells

We hypothesize that citreoviridin (CIT) induces DNA damage in human liver‐derived HepG2 cells through an oxidative stress mechanism and that N‐acetyl‐l ‐cysteine (NAC) protects against CIT‐induced DNA damage in HepG2 cells. CIT‐induced DNA damage in HepG2 cells was evaluated by alkaline single‐cell gel electrophoresis assay. To elucidate the genotoxicity mechanisms, the level of oxidative DNA damage was tested by immunoperoxidase staining for 8‐hydroxydeoxyguanosine (8‐OHdG); the intracellular generation of reactive oxygen species (ROS) and reduced glutathione (GSH) were examined; mitochondrial membrane potential and lysosomal membranes' permeability were detected; furthermore, protective effects of NAC on CIT‐induced ROS formation and CIT‐induced DNA damage were evaluated in HepG2 cells. A significant dose‐dependent increment in DNA migration was observed at tested concentrations (2.50–10.00 µM) of CIT. The levels of ROS, 8‐OHdG formation were increased by CIT, and significant depletion of GSH in HepG2 cells was induced by CIT. Destabilization of lysosome and mitochondria was also observed in cells treated with CIT. In addition, NAC significantly decreased CIT‐induced ROS formation and CIT‐induced DNA damage in HepG2 cells. The data indicate that CIT induces DNA damage in HepG2 cells, most likely through oxidative stress mechanisms; that NAC protects against DNA damage induced by CIT in HepG2 cells; and that depolarization of mitochondria and lysosomal protease leakage may play a role in CIT‐induced DNA damage in HepG2 cells. © 2014 The Authors. Published by Wiley Periodicals Inc. Environ Toxicol 30: 530–537, 2015.     

DNA damage and alterations in expression of DNA damage responsive genes induced by TiO2 nanoparticles in human hepatoma HepG2 cells

Abstract

We investigated the genotoxic responses to two types of TiO₂ nanoparticles (<25 nm anatase: TiO₂-An, and <100 nm rutile: TiO₂-Ru) in human hepatoma HepG2 cells. Under the applied exposure conditions the particles were agglomerated or aggregated with the size of agglomerates and aggregates in the micrometer range, and were not cytotoxic. TiO₂-An, but not TiO₂-Ru, caused a persistent increase in DNA strand breaks (comet assay) and oxidized purines (Fpg-comet). TiO₂-An was a stronger inducer of intracellular reactive oxygen species (ROS) than TiO₂-Ru. Both types of TiO₂ nanoparticles transiently upregulated mRNA expression of p53 and its downstream regulated DNA damage responsive genes (mdm2, gadd45α, p21), providing additional evidence that TiO₂ nanoparticles are genotoxic. The observed differences in responses of HepG2 cells to exposure to anatase and rutile TiO₂ nanoparticles support the evidence that the toxic potential of TiO₂ nanoparticles varies not only with particle size but also with crystalline structure.     

OGG1 Ser326Cys polymorphism interacts with cigarette smoking to increase oxidative DNA damage in human sperm and the risk of male infertility

8-Oxoguanine DNA glycosylase 1 (OGG1) plays an important role in repairing oxidative DNA damage induced by chemical agents, such as tobacco. This study examined the effects of OGG1 Ser326Cys polymorphism and cigarette smoking, alone or combined, on sperm oxidative DNA damage and the risk of male infertility. A total of 620 idiopathic infertile subjects and 480 fertile controls were recruited in this study. Sperm 8-hydroxydeoxyguanine (8-OHdG) was measured by immunofluorescent assay using flow cytometry and genotypes were determined by OpenArray platform with a chip-based Taq-Man genotyping technology. Our results demonstrated that both cigarette smoking and OGG1 polymorphism can affect the sperm 8-OHdG levels. Individuals with variant Cys/Cys homozygote showed higher levels of sperm 8-OHdG than wide-type homozygote carriers (Ser/Ser). Stratified analysis found that the association between OGG1 polymorphism and sperm 8-OHdG levels was only observed among smokers with pack-years ≥5 but not among those subjects with pack-years < 5 (pack-years = packs smoked per day × years as a smoker). Further analysis based on the case–control study revealed that variant allele (Cys) of OGG1 was significantly associated with male infertility risk in a dominant model (OR = 1.35, 95% CI: 1.01–1.82; trend P < 0.001). Furthermore, we found a significant gene–environment interaction between OGG1 Ser326Cys polymorphism and cigarette smoking in relation to male infertility risk (P_interation = 0.0003). These findings provided the first evidence about potential interactive effects of OGG1 polymorphism and cigarette smoking on male infertility risk.     

Combined effect of titanium dioxide nanoparticles and glucose on the cardiovascular system in young rats after oral administration

Titanium dioxide nanoparticles (TiO₂ NPs) have already been used as food additive in various products and are usually consumed with a considerable amount of sugar. Oral consumption of TiO₂ NPs poses concerning health risks; however, research on the combined effect of ingested TiO₂ NPs and glucose is limited. We examined young Sprague‐Dawley rats administrated TiO₂ NPs orally at doses of 0, 2, 10 and 50 mg/kg body weight per day with and without 1.8 g/kg body weight glucose for 30 and 90 days. Heart rate, systolic and diastolic blood pressure, blood biochemical parameters and histopathology of cardiac tissues was assessed to quantify cardiovascular damage. The results showed that oral exposure to TiO₂ NPs and high doses of glucose both could induce cardiovascular injuries. The toxic effects were dose‐, time‐ and gender‐dependent. The interaction effects between oral‐exposed TiO₂ NPs and glucose existed and revealed to be antagonism in most of the biological parameters. However, toxic effects of the high‐dose glucose seemed to be more severe than TiO₂ NPs and the interaction of TiO₂ NPs with glucose. These results suggest that it may be more important to control the sugar intake than TiO₂ NPs for protecting the health of TiO₂ NP consumers.     

Cytotoxicity and oxidative DNA damage by nanoparticles in human intestinal Caco-2 cells

Abstract

The use of engineered nanoparticles in the food sector is anticipated to increase dramatically, whereas their potential hazards for the gastrointestinal tract are still largely unknown. We investigated the cytotoxic and DNA-damaging effects of several types of nanoparticles and fine particles relevant as food additives (TiO₂ and SiO₂) or for food packaging (ZnO and MgO) as well as carbon black on human intestinal Caco-2 cells. All particles, except for MgO, were cytotoxic (LDH and WST-1 assay). ZnO, and to lesser extent SiO₂, induced significant DNA damage (Fpg-comet), while SiO₂ and carbon black were the most potent in causing glutathione depletion. DNA damage by TiO₂ was found to depend on sample processing conditions. Interestingly, application of different TiO₂ and ZnO particles revealed no relation between particle surface area and DNA damage. Our results indicate a potential hazard of several food-related nanoparticles which necessitate investigations on the actual exposure in humans.     

Beneficial effects of quercetin on oxidative stress in liver and kidney induced by titanium dioxide (TiO2) nanoparticles in rats

Abstract

TiO₂ nanoparticles used as vectors for the delivery of drugs have shown greater effectiveness. However, TiO₂ nanoparticles can cause oxidative stress in liver and kidney, so we analyzed if a previous or simultaneous quercetin treatment could counteract this in rats. Five groups of male Wistar rats (200–250?g) were included: (1) healthy controls, (2) TiO₂ group, (3) quercetin group, (4) preventive group: quercetin for 5 days prior to exposure of TiO₂, and (5) therapeutic group: TiO₂ (5?mg/kg, i.v.) plus quercetin single dose for 5 days (5?mg/kg/day, i.p.). Hepatic and renal function tests were made. Five animals from each group were sacrificed (0, 14 and 28 days), and liver and kidney tissue were obtained. Malondialdehyde (MDA), reduced/oxidized glutathione, and activity of glutathione peroxidase/reductase were measured, as well as the level of gene expression by q-PCR. There were no significant changes in serum ALT and AST activities. More damage was observed at 14 versus 28 days, because TiO₂ was excreted in urine. Quercetin indeed showed a renal protective effect by increasing glutathione reductase and peroxidase levels and reducing MDA levels. On the other hand, TiO₂ liver damage was less pronounced with quercetin as therapeutic treatment. TiO₂ induces significantly the glutathione reductase expression and it can be down-regulated by quercetin. Biochemical tests in serum and urine showed a better effect of quercetin administered in the therapeutic group. Care should be taken with the dose and time of administration of quercetin, because this antioxidant could also have a pro-oxidant effect.     

Protective effect of amifostine on busulfan induced DNA damage in human hepatoma cells

Busulfan is one of the most effective chemotherapeutic agents used for the treatment of chronic myeloid leukemia. However, as a bifunctional alkylating agent, during clinical use several side effects may occur. In addition, several in vivo and in vitro studies of busulfan have shown a range of genotoxic effects including DNA strand break and inhibition of DNA synthesis. Amifostine, an organic thiophosphate compound, has been shown to exert an important cyto-protective effect in many tissues. The aim of this study was to explore whether amifostine protects against busulfan-induced genotoxicity in HepG2 cell line. Our results showed that amifostine reduced the genotoxic effects of busulfan significantly in both type of experiment conditions, as measured via comet assay. Furthermore, amifostine decreased the intracellular ROS generation induced by busulfan and also increased the intracellular GSH levels in HepG2 cells. Altogether, our results suggest a protective action of amifostine against busulfan cytotoxicity and genotoxicity via various pathways. The most protective effect was observed with amifostine when it was administrated 24?h before busulfan treatment.     

DNA damage and repair following In vitro exposure to two different forms of titanium dioxide nanoparticles on trout erythrocyte

TiO₂ has been widely used to promote organic compounds degradation on waste aqueous solution, however, data on TiO₂ nanotoxicity to aquatic life are still limited. In this in vitro study, we compare the toxicity of two different families of TiO₂ nanoparticles on erythrocytes from Oncorhynchus mykiss trout. The crystal structure of the two TiO₂ nanoparticles was analyzed by XRD and the results indicated that one sample is composed of TiO₂ in the anatase crystal phase, while the other sample contains a mixture of both the anatase and the rutile forms of TiO₂ in a 2:8 ratio. Further characterization of the two families of TiO₂ nanoparticles was determined by SEM high resolution images and BET technique. The toxicity results indicate that both TiO₂ nanoparticles increase the hemolysis rate in a dose dependent way (1.6, 3.2, 4.8 μg mL^?1) but they do not influence superoxide anion production due to NADH addition measured by chemiluminescence. Moreover, TiO₂ nanoparticles (4.8 μg mL^?1) induce DNA damage and the entity of the damage is independent from the type of TiO₂ nanoparticles used. Modified comet assay (Endo III and Fpg) shows that TiO₂ oxidizes not only purine but also pyrimidine bases. In our experimental conditions, the exposure to TiO₂ nanoparticles does not affect the DNA repair system functionality. The data obtained contribute to better characterize the aqueous environmental risks linked to TiO₂ nanoparticles exposure. © 2011 Wiley Periodicals, Inc. Environ Toxicol 29: 117–127, 2014.     

Combined effect of titanium dioxide nanoparticles and glucose on the blood glucose homeostasis in young rats after oral administration

Titanium dioxide nanoparticles (TiO₂ NPs) were usually consumed with a high content of sugar, and children were identified as having the highest exposures due to sweet food preferences. Research on the combined effect of ingested TiO₂ NPs and glucose has great significance, particularly in young people. We examined young Sprague-Dawley rats administered TiO₂ NPs (0, 2, 10 and 50 mg/kg) orally with and without glucose (1.8 g/kg) for 90 days. Blood glucose homeostasis was assessed by monitoring blood glucose and detecting glycoproteins. Glucose tolerance was also evaluated by the oral glucose tolerance test. The levels of blood glucose-related hormones such as insulin, C-peptide and glucagon were measured. We found that subchronic co-exposure of TiO₂ NPs and glucose caused slight imbalance of blood glucose homeostasis in vivo. Mild and temporary hypoglycemia, impaired glucose tolerance and changes of glucose-regulating hormones were shown in the exposure groups. The combined effect of TiO₂ NPs and glucose was more apparent than that of TiO₂ NPs alone, which may be due to the effects of excess glucose and the interactions between TiO₂ NPs and glucose. The antagonistic effect of TiO₂ NPs with glucose did exist in the level of glycosylated hemoglobin in female rats. Gender differences were apparent in these effects induced by TiO₂ NPs and glucose. Female rats seemed to be more susceptible for blood glucose disorders. Co-exposure of TiO₂ NPs and excessive glucose could induce gender-dependent imbalance of blood glucose homeostasis in rats. It may be the reason that these consumers face greater health risks glycosylated hemoglobin.     

Oxidative stress induced by atrazine in rat erythrocytes: Mitigating effect of vitamin E

The present study investigates the propensity of atrazine to induce oxidative stress and its possible attenuation by vitamin E in rat erythrocytes, which is a convenient model to understand the oxidative damage induced by various xenobiotics. Experimental animals were administered atrazine (300?mg/kg body weight, daily) and/or vitamin E (100?mg/kg body weight, daily) orally for a period of 7, 14, and 21 days. Results indicated that the reduced glutathione (GSH) content of the erythrocytes of atrazine treated rats was significantly decreased as compared to the control group. Co-administration of vitamin E along with atrazine restored the GSH content of erythrocytes nearly to control levels. The activities of antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidase, and glutathione-s-transferase were found to be increased significantly in the erythrocytes accompanied by a decrease in the activity of the glucose-6-phosophate dehydrogenase, following atrazine exposure. On the other hand, when vitamin E was co-administered along with atrazine, activities of these enzymes were found to be restored significantly. In conclusion, results of the study demonstrated that atrazine induced oxidative stress in rat erythrocytes, in terms of increased activities of the various antioxidant enzymes, and decreased content of reduced glutathione. However, vitamin E administration ameliorated the effects of atrazine, suggesting that vitamin E is a potential antioxidant against atrazine-induced oxidative stress.