Investigation on cobalt‐oxide nanoparticles cyto‐genotoxicity and inflammatory response in two types of respiratory cells

The increasing use of cobalt oxide (Co₃O₄) nanoparticles (NPs) in several applications and the suggested genotoxic potential of Co‐oxide highlight the importance of evaluating Co₃O₄ NPs toxicity. Cyto‐genotoxic and inflammatory effects induced by Co₃O₄ NPs were investigated in human alveolar (A549), and bronchial (BEAS‐2B) cells exposed to 1–40 µg ml^–1. The physicochemical properties of tested NPs were analysed by transmission electron microscopy (TEM) and dynamic light scattering (DLS). Cytotoxicity was studied to analyze cell viability (WST1 test) and membrane damage (LDH assay), direct/oxidative DNA damage was assessed by the Formamido‐pyrimidine glycosylase (Fpg)‐modified comet assay and inflammation by interleukin (IL)‐6, IL‐8 and tumor necrosis factor‐alpha (TNF‐α) release (ELISA). In A549 cells, no cytotoxicity was found, whereas BEAS‐2B cells showed a viability reduction at 40 µg ml^–1 and early membrane damage at 1, 5 and 40 µg ml–1. In A549 cells, direct and oxidative DNA damage at 20 and 40 µg ml^–1 were detected without any effects on cytokine release. In BEAS‐2B cells, significant direct DNA damage at 40 µg ml^–1 and significant oxidative DNA damage with a peak at 5 µg ml^–1, that was associated with increased TNF‐α release at 1 µg ml^–1 after 2 h and increased IL‐8 release at 20 µg ml^–1 after 24 h, were detected. The findings show in the transformed alveolar cells no cytotoxicity and genotoxic/oxidative effects at 20 and 40 µg ml^–1. In normal bronchial cells, moderate cytotoxicity, direct DNA damage only at the highest concentration and significant oxidative‐inflammatory effects at lower concentrations were detected. The findings confirm the genotoxic‐oxidative potential of Co₃O₄ NPs and show greater sensitivity of BEAS‐2B cells to cytotoxic and oxidative‐inflammatory effects suggesting the use of different cell lines and multiple end‐points to elucidate Co₃O₄ NPs toxicity. Copyright © 2015 John Wiley & Sons, Ltd.     

Direct‐oxidative DNA damage and apoptosis induction in different human respiratory cells exposed to low concentrations of sodium chromate

The mechanism of Cr(VI) genotoxicity has still not been elucidated. We used Fpg‐modified comet assay to assess direct‐oxidative DNA damage on human lung (A549) and bronchial (BEAS‐2B) cells exposed to 0.1, 0.5, 1.0 and 10 μm sodium chromate for 0.5, 1 and 4 h. Moreover we evaluated apoptosis by morphological analysis and caspase‐3 activity, also after 24 h. On A549 cells a time‐dependent DNA damage, expressed as tail DNA%, beginning from 0.5 μm was found. For oxidative DNA damage an induction after 30 min to 0.5 μm decreasing with time, and a time‐dependent increase at 10 μm was found, indicating for low Cr(VI) concentration the oxidative stress as the first event followed by direct DNA damage and for the highest concentration a time‐dependent increase in oxidative DNA damage. On BEAS‐2B cells DNA damage was induced within 1 h at 0.5–10 μm , without changes with time, showing that BEAS‐2B cells are able to resist to Cr(VI) genotoxicity. Early oxidative DNA damage at 0.1 μm decreasing with time was also found. Significant apoptosis was observed by morphological analysis in A549 cells and to a lower extent in BEAS‐2B at 10 μm . The exposure to 10 μm induced caspase‐3 activity after 4 h in BEAS‐2B and after 24 h in A549 cells. The findings show a higher responsiveness of A549 cells to genotoxic effect of Cr(VI) and early transient oxidative DNA damage in BEAS‐2B. The results emphasize the suitability of this experimental model to evaluate the early genotoxic response of different cells to non‐cytotoxic concentrations of Cr(VI) on target organ. Copyright © 2009 John Wiley & Sons, Ltd.     

Multi‐walled carbon nanotubes induce cytotoxicity and genotoxicity in human lung epithelial cells

The increasing use of nanomaterials in consumer products highlights the importance of understanding their potential toxic effects. We evaluated cytotoxic and genotoxic/oxidative effects induced by commercial multi‐walled carbon nanotubes (MWCNTs) on human lung epithelial (A549) cells treated with 5, 10, 40 and 100 µg ml^?1 for different exposure times. Scanning electron microscopy (SEM) analysis, MTT [3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl tetrazolium bromide] and lactate dehydrogenase (LDH) assays were performed to evaluate cytotoxicity. Fpg‐modified comet assay was used to evaluate direct‐oxidative DNA damage. LDH leakage was detected after 2, 4 and 24 h of exposure and viability reduction was revealed after 24 h. SEM analysis, performed after 4 and 24 h exposure, showed cell surface changes such as lower microvilli density, microvilli structure modifications and the presence of holes in plasma membrane. We found an induction of direct DNA damage after each exposure time and at all concentrations, statistically significant at 10 and 40 µg ml^?1 after 2 h, at 5, 10, 100 µg ml^?1 after 4 h and at 10 µg ml^?1 after 24 h exposure. However, oxidative DNA damage was not found. The results showed an induction of early cytotoxic effects such as loss of membrane integrity, surface morphological changes and MWCNT agglomerate entrance at all concentrations. We also demonstrated the ability of MWCNTs to induce early genotoxicity. This study emphasizes the suitability of our approach to evaluating simultaneously the early response of the cell membrane and DNA to different MWCNT concentrations and exposure times in cells of target organ. The findings contribute to elucidation of the mechanism by which MWCNTs cause toxic effects in an in vitro experimental model. Copyright © 2012 John Wiley & Sons, Ltd.     

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.     

Cyto-genotoxic and inflammatory effects of commercial Linde Type A (LTA) nanozeolites on human alveolar epithelial cells

Nanozeolites (NZs) are increasingly used in several sectors, including catalysts, ion exchange materials or thermal isolators, taking advantage of the major property of NZs to absorb residual water and moisture to preserve the insulation of devices and products, but very few data are available on their toxicity. We investigated the potential cyto-genotoxicity and pro-inflammatory effects of manufactured Linde Type A (LTA)-NZs on human alveolar cells (A549) exposed to 10, 25, 50 and 100 μg/mL. LTA NZs were characterized by dynamic light scattering (DLS). Cell viability, mortality and apoptosis were evaluated by cytofluorimetric assay after 24h exposure. Membrane damage was evaluated by lactate dehydrogenase release and direct and oxidative DNA damage induction by formamide-pyrimidine glycosylase-Comet assay after 4 and 24 h. The induction of pro-inflammatory effects was evaluated in terms of interleukin 6 (IL-6) and IL-8 cytokine release after 24 h by ELISA. We found a slight increase in apoptotic cell percentage at 50 and 100 μg/mL and dead cell percentage at 100 μg/mL after 24 h; slight, but statistically significant, direct DNA damage starting from 25 μg/mL and slight oxidative DNA damage both at 4 and at 24 h; increased release of IL-6 only at the lowest concentration after 24 h. The results show lack of cytotoxicity, early moderate genotoxicity and slight inflammatory effects at the lowest used concentration. These findings represent the first data on potential genotoxic, oxidative and inflammatory effects of LTA NZs and highlight the need to perform further studies to confirm such results.     

Evaluation of cytotoxic,oxidative stress,proinflammatory and genotoxic effect of silver nanoparticles in human lung epithelial cells

Silver nanoparticles are increasingly used in various products, due to their antibacterial properties. Despite its wide spread use, only little information on possible adverse health effects exists. Therefore, the aim of this study was to assess the toxic potential of silver nanoparticles (<100 nm) in human lung epithelial (A549) cells and the underlying mechanism of its cellular toxicity. Silver nanoparticles induced dose and time‐dependent cytotoxicity in A549 cells demonstrated by MTT and LDH assays. Silver nanoparticles were also found to induce oxidative stress in dose and time‐dependent manner indicated by depletion of GSH and induction of ROS, LPO, SOD, and catalase. Further, the activities of caspases and the level of proinflammatory cytokines, namely interleukin‐1β (IL‐1β) and interleukin‐6 (IL‐6) were significantly higher in treated cells. DNA damage, as measured by single cell gel electrophoresis, was also dose and time‐dependent signicants in A549 cells. This study investigating the effects of silver nanoparticles in human lung epithelial cells has provided valuable insights into the mechanism of potential toxicity induced by silver nanoparticles and warrants more careful assessment of silver nanoparticles before their industrial applications. © 2013 Wiley Periodicals, Inc. Environ Toxicol 30: 149–160, 2015.     

In vitro genotoxic and cytotoxic effects of doxepin and escitalopram on human peripheral lymphocytes

Antidepressants are drugs used for the treatment of many psychiatric conditions including depression. There are findings suggesting that these drugs might have genotoxic, carcinogenic, and/or mutagenic effects. Therefore, the present in vitro study is intended to investigate potential genotoxic and cytotoxic effects of the antidepressants escitalopram (selective serotonin reuptake inhibitor) and doxepin (Tricyclic antidepressant) on human peripheral lymphocytes cytokinesis-block micronucleus (CBMN), sister chromatid exchange (SCE), and single cell gel electrophoresis (alkaline comet assay) were used for the purpose of the study. In the study, four different concentrations of both drugs (1, 2.5, 5, and 10?µg/mL) were administered to human peripheral lymphocytes for 24?h. The tested concentrations of both drugs were found to exhibit no cytotoxic and mitotic inhibitory effects. SCE increase caused by 5 and 10?µg/mL of escitalopram was found statistically significant, while no statistically significant increase was observed in DNA damage and micronucleus (MN) formation. Moreover, the increase caused by doxepin in MN formation was not found statistically significant. Besides, 10?µg/mL of doxepin was demonstrated to significantly increase arbitrary unit and SCE formation. These findings suggest that the investigated concentrations of escitalopram and doxepin were non-cytotoxic but potentially genotoxic at higher concentrations.     

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.     

Genotoxic and cytotoxic potential of titanium dioxide (TiO2) nanoparticles on fish cells in vitro

Intrinsic genotoxic and cytotoxic potential of titanium dioxide (TiO2) engineered nanoparticles (ENPs) were evaluated in a metabolically competent, established fish cell line derived from rainbow trout (Oncorhyncus mykiss) gonadal tissue (i.e. RTG-2 cells). Prior to evaluation of the toxic potential, mean size of the ENPs was determined using transmission electron microscopy (TEM). As a prerequisite, an extensive characterisation of the ENPs was carried out following sonication which enabled the synthesis of an efficient dosing strategy for the cells in which exposure in phosphate buffered saline (PBS) gave an optimal agglomeration effects compared to distilled water (H2O) and minimal essential media (MEM). Interaction of the ENPs with cells under scanning electron microscope (SEM) was also studied. The genotoxic and cytotoxic potential of the ENPs were determined either alone or in combination with ultraviolet radiation (i.e. UVA). Whilst genotoxic potential was determined by evaluating DNA strand breaks using single cell gel electrophoresis (SCGE) or the comet assay and induction of cytogenetic damage using cytokinesis-blocked micronucleus (MN) assay, cytotoxicity was determined by measuring the retention of supra vital stain, neutral red, by the lysosomes using the neutral red retention (NRR) assay. In addition, while performing the comet assay, lesion specific bacterial endonuclease, formamidopyrimidine DNA glycosylase (Fpg), which recognises oxidised purine bases, was used to determine oxidative DNA damage. The results suggested that the highest concentration of the ENPs (i.e. 50 microg ml(-1)) did not produce elevations in DNA damage over 4 h (comet assay), 24 h (modified comet assay) or 48 h (MN assay) exposures in the absence of UVA irradiation, although there was a significant reduction in lysosomal integrity over 24 h exposure (NRR assay). The induction of MN did not show any enhanced levels as a function of ENP concentration. A significantly increased level of strand breaks was observed in combination with UVA (3 kJ m(-2)). In general, the NRR assay suggested elevated levels of cytotoxicity when the UVA exposure was carried out with MEM compared to PBS, although both showed an increase when in combination with the highest concentration of ENPs (i.e. 50 microg ml(-1)). Overall, the study emphasises the need for adoption of an holistic approach while evaluating the potential toxic effects of ENPs in which appropriate measures should be taken to avoid agglomeration or aggregation to facilitate efficient cellular uptake to evaluate potential biological responses.     

Analysis of cytotoxic effects of silver nanoclusters on human peripheral blood mononuclear cells ‘in vitro’

The antimicrobial properties of silver nanoparticles (AgNPs) have made these particles one of the most used nanomaterials in consumer products. Therefore, an understanding of the interactions (unwanted toxicity) between nanoparticles and human cells is of significant interest. The aim of this study was to assess the in vitro cytotoxicity effects of silver nanoclusters (AgNC, < 2 nm diameter) on peripheral blood mononuclear cells (PBMC). Using flow cytometry and comet assay methods, we demonstrate that exposure of PBMC to AgNC induced intracellular reactive oxygen species (ROS) generation, DNA damage and apoptosis at 3, 6 and 12 h, with a dose‐dependent response (0.1, 1, 3, 5 and 30 µg ml^–1). Advanced electron microscopy imaging of complete and ultrathin‐sections of PBMC confirmed the cytotoxic effects and cell damage caused by AgNC. The present study showed that AgNC produced without coating agents induced significant cytotoxic effects on PBMC owing to their high aspect ratio and active surface area, even at much lower concentrations (<1 µg ml^–1) than those applied in previous studies, resembling what would occur under real exposure conditions to nanosilver‐functionalized consumer products. Copyright © 2015 John Wiley & Sons, Ltd.     

Antigenotoxic activities of the natural dietary coumarins umbelliferone,herniarin and 7-isopentenyloxy coumarin on human lymphocytes exposed to oxidative stress

Abstract

The antigenotoxic effects of umbelliferone (UMB), herniarin (HER) and 7-isopentenyloxy coumarin (7-IP), common natural dietary coumarins, were evaluated on the human lymphocyte DNA damage using single-cell gel electrophoresis. H₂O₂-induced DNA break was measured based on the percentage of DNA in tail, and the antigenotoxic effects of the tested compounds were compared with that of ascorbic acid (10, 25, 50, 100 and 200?μM). UMB, HER and 7-IP did not show any genotoxicity, as compared to phosphate-buffered saline. Treatment with UMB, HER and 7-IP led to a significant reduction in the percentage of DNA in tail induced by H₂O₂ (p?<?0.001) at all concentrations. The presence of prenyl moiety in the chemical structure of 7-IP may contribute to its better antigenotoxic property, compared to UMB. The results of this study showed that 7-IP possessed the best antigenotoxic activity among the tested compounds.     

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.     

Long-term exposures to low doses of titanium dioxide nanoparticles induce cell transformation,but not genotoxic damage in BEAS-2B cells

Abstract

There is a great interest in a better knowledge of the health effects caused by nanomaterials exposures and, in particular to those induced by titanium dioxide nanoparticles (nano-TiO₂) due to its high use and increasing presence in the environment. To add new information on its potential genotoxic/carcinogenic risk, we have carried out experiments using chronic exposures (up to 4 weeks), low doses, and the BEAS-2B cell line that, as a human bronchial epithelium cells, can be considered a good cell target. Cell uptake has been assessed by transmission electron microscopy (TEM) and flow cytometry (FC); genotoxicity was evaluated using the comet and the micronucleus (MN) assays; and cell-transforming ability was evaluated using the soft-agar assay to detect anchorage-independent cell growth. Results show an important cell uptake at all the tested doses and sampling times used (except for 1?µg/mL and 24-h exposure). Nevertheless, no genotoxic effects were observed in the comet and in the MN assays. This lack of genotoxic effect agrees with the FC results showing no induction of intracellular reactive oxygen species (ROS), the data from the comet assay with formamidopyrimidine DNA glycosylase (FPG) enzyme showing no induction of oxidized bases, and the lack of induction of expression of heme-oxygenase (HO-1) gene both at the RNA and protein level. On the contrary, significant increases in the number of clones growing in an anchorage-independent way were observed. This study would indicate a potential carcinogenic risk associated to nano-TiO₂ exposure, not mediated by a genotoxic mechanism.     

Oxidative damage and OGG1 expression induced by a combined effect of titanium dioxide nanoparticles and lead acetate in human hepatocytes

Titanium dioxide (TiO(2)) is a widely used nanomaterial that can cause biological damage through oxidative stress. At low concentrations, TiO(2) can interact with lead acetate (PbAc) to produce different toxic responses, compared with TiO(2) or PbAc alone. In this study, we utilized the following as indicators of toxic responses in human embryo hepatocytes (L02): reactive oxygen species (ROS), reduced glutathione (GSH), superoxide dismutase (SOD), and the DNA adducts 8-hydroxydeoxyguanosine (8-OHdG) and 8-oxoguanine DNA glycosylase homolog 1 (OGG1). These were used to evaluate the oxidative stress of TiO(2) (at 0.001, 0.01, 0.1, 1, and 10 μg mL(-1)) mixed with PbAc (1 μg mL(-1)) on L02 cells without photoactivation. Compared with the negative control (1‰ dimethyl sulfoxide), TiO(2) mixed with PbAc induced increased release of ROS (at 0.001, 0.01, 0.1, 1, 10 μg mL(-1) TiO(2)), intracellular SOD activity (at 0.1 and 0.01 μg mL(-1) TiO(2)), GSH levels (at 0.01-1 μg mL(-1) TiO(2)), 8-OHdG levels (at 1 and 10 μg mL(-1) TiO(2)), OGG1 expression (at 0.001-1 μg mL(-1) TiO(2)), and cytotoxicity (at 0.1, 1, and 10 μg mL(-1) TiO(2)) in L02 cells. There were no significant changes in ROS, GSH, SOD, 8-OHdG, or OGG1 levels when L02 cells were treated with TiO(2) alone or PbAc alone. These findings indicate that TiO(2) and PbAc in combination induce cytotoxicity and oxidative stress in L02 cells in the absence of photoactivation.     

Toxicity assessment of six titanium dioxide nanoparticles in human epidermal keratinocytes

Purpose: The aim of this study was to evaluate and compare the toxicity of six different types of titanium dioxide (TiO₂) nanoparticles (NP) on human epidermal keratinocytes (HEK).

Materials and methods: Six TiO₂ NP (A (10?nm), A*(32?nm), B (27.5?nm), C (200?nm), C*(30–40?nm), and D*(200–400?nm)) were suspended in water or culture medium and characterized by transmission electron microscopy (TEM) and dynamic light scattering (DLS). In addition, these NP were assayed with cell viability, cytokine release and cellular uptake in HEK.

Results: TiO₂NP did not change in shape in the culture medium when visualized by TEM. There was an increase in agglomeration with all TiO₂NP in the medium when measured by DLS. Since TiO₂NP interfered with the CellTiter 96®AQueous One and MTT assays but had a minimal effect on alamar Blue (aB). The aB viability assay was selected to assess all six types of TiO₂NP and sample B had a statistically significant decrease in viability at 0.4?mg/ml. A slight increase in TNF-α was noted in sample A*, C, and D* at as low as 0.05?mg/ml. Sample A* and B at certain concentrations showed an increase in Interleukin (IL)-6. IL-10 and IL-1β release for all TiO₂NP were noted around the detection limit with no significant changes compared to control. A statistically significant decrease in IL-8 was noted for all TiO₂NP at the highest concentrations due to the adsorption of IL-8 by TiO₂. All TiO₂NP were localized within cytoplasmic vacuoles of HEK and the element Ti was detected by energy-dispersive x-ray spectroscopy analysis.

Conclusions: Based on cell viability, only sample B was slightly cytotoxic to HEK and samples B and A* have the potential to cause inflammation indicated by an increase in IL-6.     

In vitro evaluation of the cytotoxic and genotoxic effects of artemether,an antimalarial drug,in a gastric cancer cell line (PG100)

Artemisinin is a sesquiterpene lactone endoperoxide, obtained from Artemisia annua, and extensively used as an antimalarial drug. Many studies have reported the genotoxic and cytotoxic effects of artemisinins; however, there are no studies that compare such effects between cancer cell lines and normal human cells after treatment with artemether, an artemisinin derivative. Gastric cancer is the fourth most frequent type of cancer and the second highest cause of cancer mortality worldwide. Thus, the aim of this study was to evaluate the in vitro genotoxic and cytotoxic effects induced by artemether in gastric cancer cell line (PG100) and compare them with the results obtained in human lymphocytes exposed to the same conditions. We used MTT (3‐(4,5‐methylthiazol‐2‐yl)‐2, 5‐diphenyl‐tetrazolium bromide) assay, comet assay and ethidium bromide/acridine orange viability staining to evaluate the cytotoxic and genotoxic effects of artemether in PG100. MTT assay showed a decrease in the survival percentages for both cell types treated with different concentrations of artemether (P < 0.05). PG100 also showed a significant dose‐dependent increase in DNA damage index at concentrations of 119.4 and 238.8 µg ml^?1 (P < 0.05). Our results showed that artemether induced necrosis in PG100 at concentrations of 238.8 and 477.6 µg ml^?1, for all the tested harvest times (P < 0.05). In lymphocytes, artemether induced both apoptosis and necrosis at concentrations of 238.8 and 477.6 µg ml^?1, for all the tested harvest times (P < 0.05). In conclusion, human lymphocytes were more sensitive to the cytotoxic effects of the antimalarial drug than the gastric cancer cell line PG100. Copyright © 2011 John Wiley & Sons, Ltd.     

Cytotoxic,genotoxic and the hemolytic effect of titanium dioxide (TiO2) nanoparticles on human erythrocyte and lymphocyte cells in vitro

With the increasing clinical use of titanium dioxide (TiO₂) nanoparticles, a better understanding of their safety in the blood stream is required. The present study evaluates the toxic effect of commercially available TiO₂ nanoparticles (~100 nm) using a battery of cytotoxic, genotoxic, hemolytic and morphological parameters. The cytotoxic effects of TiO₂ nanoparticles in human lymphocyte cells were studied with respect to membrane damage, mitochondrial function, metabolic activity and lysosomal membrane stability. Genotoxicity in lymphocyte cells was quantitated using a comet assay. The mode of cell death (apoptosis/necrosis) was evaluated using PI/Annexin V staining. TiO₂ nanoparticles were also evaluated for their hemolytic properties, osmotic fragility and interaction with hemoglobin. Human erythrocyte cells were studied for morphological alterations using atomic force microscopy (AFM). Results suggest that the particles could induce a significant reduction in mitochondrial dehydrogenase activity in human lymphocyte cells. Membrane integrity remained unaffected by nanoparticle treatment. DNA damage and apoptosis were induced by TiO₂ nanoparticles in a dose‐dependent manner. A study on human erythrocyte cells revealed a hemolytic property of TiO₂ nanoparticles characterized by spherocytosis and echinocytosis. Spectral analysis revealed a hemoglobin TiO₂ nanoparticle interaction. Our in vitro study results suggest that commercially available blood contacting nanoparticles (TiO₂ nanoparticle) should be carefully evaluated for their toxic potential. Copyright © 2013 John Wiley & Sons, Ltd.     

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.     

Uranium induces apoptosis and is genotoxic to normal rat kidney (NRK-52E) proximal cells.

Uranium (U) is a heavy metal used in the nuclear industry and for military applications. U compounds are toxic. Their toxicity is mediated either by their radioactivity or their chemical properties. Mammalian kidneys and bones are the main organs affected by U toxicity. Although the most characteristic response to U exposure is renal dysfunction, little information is available on the mechanisms of its toxicity at the molecular level. This report studied the genotoxicity of U. Apoptosis induction in normal rat kidney (NRK-52(E)) proximal cells was investigated as a function of exposure time or concentrations (0-800microM). In parallel, DNA damage was evaluated by several methods. In order to distinguish between the intrinsic and the extrinsic pathways of apoptosis, caspases-8, -9, -10 assays were conducted and the mitochondrial membrane potential was measured. Three methods were selected for their complementarities in the detection of genetic lesions. The comet assay was used for the detection of primary lesions of DNA. gamma-H2AX immunostaining was achieved to detect DNA double-strand breaks. The micronucleus assay was used to detect chromosomic breaks or losses. DNA damage and apoptosis were observed in a concentration-dependent manner. This study demonstrated that U is genotoxic from 300microM and induces caspase-dependent apoptosis cell death from 200microM mainly through the intrinsic pathway in NRK-52(E) cells. These results suggest that the DNA damage caused by U is reversible at low concentration (200-400microM) but becomes irreversible and leads to cell death for higher concentrations (500-800microM).