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The possibility of eye exposure for workers participating in manufacturing of nanoparticles or consumers using products containing nanoparticles has been reported, but toxicity studies on the eye are scarce. In this study, cytotoxicity of five nanoparticles including silver, ceria, silica, titanium and zinc were tested using Statens Seruminstitut Rabbit Cornea (SIRC) cells. When cells were treated with nanoparticles with concentrations of 1-100 μg/mL for 24 hr, zinc oxide nanoparticles showed higher toxicity to cornea cells. LC50 of zinc oxide nanoparticles was less than 25 μg/mL but those of other nanoparticles could not be calculated in this test, which means more than 100 μg/mL. Generation of reactive oxygen species was observed, and expression of apoptosis related biomarkers including Bax and Bcl-2 were changed after treatment of zinc oxide nanoparticles, while no other significant toxicity- related changes were observed in cornea cells treated with Ag, CeO2, SiO2 and TiO2 nanoparticles.

Methylisothiazolinone (MIT) has been used in combination with methylchloroisothiazolinone (CMIT) for cosmetic products such as shampoo, body lotion, and skin care products. The mixture of CMIT/MIT has been found to cause allergic contact dermatitis and is thus no longer permitted for use as a preservative in leave-on cosmetics. However, MIT itself was approved as a stand-alone preservative at a maximum concentration of 100 ppm as the toxicity was derived from CMIT rather than MIT. However, in many countries, allergic skin irritation caused by MIT remains a social concern. In this study, skin irritation was assessed for the presence of MIT, propylene glycol, and their mixture using a 3D human skin model EpiDerm™. Although non-diluted MIT causes serious skin toxicity, skin irritation was not observed at a concentration of 100 ppm, the maximum permissible level for cosmetics and personal care products according to European regulations. Propylene glycol, the most widely used vehicle for MIT, did not cause skin irritation in the 3D skin model. The results are expected to provide information for regulatory policies and guidelines on the use of biocides in consumer products.

Toxicity and target organ distribution of cerium oxide nanoparticles (CeNPs) were investigated via single intravenous injection and single oral administration, respectively. Rats were sacrificed at 24 h after treatment with doses of 30 and 300 mg/kg, and cerium concentrations were measured in liver, kidney, spleen, lung, blood, urine and feces. Results revealed cerium levels in blood and tissues were considerably low in oral treated groups and most cerium was detected in feces, meaning CeNPs would not be absorbed in the gastro-intestinal system. Conversely, high concentrations of cerium were detected in all tissues of rats after intravenous injection. Liver and spleen were main target organs. Cerium levels in liver were 594.9 ± 95.3 μg/g tissue in 30 mg/kg treat group and 3741.7 ± 932.7 μg/g tissue in 300 mg/kg treat group. Cerium levels in spleen reached almost levels of liver. Cerium was also detected, that is relatively low compared to oral administration, in feces of rats treated via intravenous injection, that supports biliary excretion of CeNPs. Urine excretion of CeNPs was not detected in oral treatment and intravenous injection. In accordance with level of cerium distribution, toxicities based on hematology, serum biochemistry and histopathology were observed in rats treated by intravenous injection while no significance was revealed in orally treated groups.

Some chemicals commonly used in personal care products, household items, food vessels, cosmetics, and other consumer products are potentially harmful, and several reviews of epidemiological studies have suggested the associations between the chemical exposure from consumer products, and respiratory diseases, skin sensitization, and reproductive problems. Therefore, risk assessment is essential for management of consumer products safety. Necessarily, the estimation of human exposure is an essential step in risk assessment, and the absorption rate of those chemicals via the gastrointestinal tract, respiratory tract, and skin are very critical in determining the internal dose of the exposed chemicals. In this study, parallel artificial membrane permeability assays (PAMPA) for the gastrointestinal tract and skin were performed to evaluate the permeability of parabens (4-hydroxybenzoic acid, methyl-, propyl-, and butyl paraben), bisphenols (bisphenol A, bisphenol F, and bisphenol S), isothiazolinones (methyl-, chloromethyl-, benz-, octyl-, and dichlorooctyl isothiazolinone), and phthalates [diethyl-, dibutyl-, Di-isononyl-, and bis(2-ethylhexyl) phthalate]. Lipid solubility of test chemicals indicated by log P values was shown as the most critical factor and showed a positive association with the permeability of parabens, bisphenols, and isothiazolinones in PAMPA assay. However, phthalate showed a reverse-association between lipophilicity and permeability. The permeability of all the tested chemicals was higher in the gastrointestinal tract membrane than in the skin membrane. The pH in donor solution did not show significant effects on the permeability in all the chemicals, except the chemicals with a free hydrophilic moiety in their chemical structures.

Background Cerium oxide nanoparticles (CeO 2 NPs) are widely used in various commercial applications because of their characteristic properties. CeO 2 NPs can be easily exposed to humans in real life, but the safety assessment of CeO 2 NPs has not been fully investigated. Therefore, in this study, we conducted a combined repeated-dose and reproductive/developmental toxicity screening study to investigate the potential effects on general health hazards, including reproductive/developmental functions, after repeated CeO 2 NPs oral exposure. In addition, tissues from parental animals and their pups were collected to analyze the internal accumulation of cerium. CeO 2 NPs were orally administered to Sprague-Dawley rats at doses of 0, 100, 300 and 1000 mg/kg during their pre-mating, mating, gestation and early lactation periods. Results In the general systemic and reproductive/developmental examinations, no marked toxicities were observed in any of in-life and terminal observation parameters in this study. In the biodistribution analysis, cerium was not detected in either parental or pup tissues (blood, liver, lungs and kidneys). Conclusion Repeated oral exposure of CeO 2 NPs did not induce marked toxicities affecting general systemic and reproductive/developmental functions up to the dose level of 1000 mg/kg and the CeO 2 NPs were not systemically absorbed in parental animals or their pups. This result could be used to risk assessment for human, and additional toxicity studies with CeO 2 NPs will be necessary considering various physicochemical properties and exposure probabilities of this nanoparticles.

Yeo Jin Kim,1,2 Md Mujibur Rahman,1 Sang Min Lee,2 Jung Min Kim,3 Kwangsik Park,4 Joo-Hyon Kang,5 Young Rok Seo1,2 1Institute of Environmental Medicine for Green Chemistry, Dongguk University Biomedi Campus, Ilsandong-gu, Goyang-si, Republic of Korea; 2Department of Life Science, Dongguk University Biomedi Campus, Ilsandong-gu, Goyang-si, Republic of Korea; 3Genoplan Korea, Inc., Seocho-gu, Seoul, Republic of Korea; 4College of Pharmacy, Dongduk Women’s University, Seongbuk-gu, Seoul, Republic of Korea; 5Department of Civil & Environmental Engineering, Dongguk University, Jung-gu, Seoul, Republic of Korea Background: Silver nanoparticles (AgNPs) are widely used in industrial and household applications, arousing concern regarding their safety in humans. The risks posed by stabilizer-coated AgNPs continue to be unclear, and assessing their toxicity is for an understanding of the safety issues involved in their use in various applications. Purpose: We aimed to investigated the long-term toxicity of citrate-coated silver nanoparticles (cAgNPs) in liver tissue using several toxicity tests and transcriptomic analysis at 7 and 28 days after a single intravenous injection into rabbit ear veins (n=4). Materials and methods: The cAgNPs used in this study were in the form of a 20% (w/v) aqueous solution, and their size was 7.9±0.95 nm, measured using transmission electron microscopy. The animal experiments were performed based on the principles of good laboratory practice. Results: Our results showed that the structure and function of liver tissue were disrupted due to a single exposure to cAgNPs. In addition, in vivo comet assay showed unrepaired genotoxicity in liver tissue until 4 weeks after a single injection, suggesting a potential carcinogenic effect of cAgNPs. In our transcriptomic analysis, a total of 244 genes were found to have differential expression at 28 days after a single cAgNP injection. Carefully curated pathway analysis of these genes using Pathway Studio and Ingenuity Pathway Analysis tools revealed major molecular networks responding to cAgNP exposure and indicated a high correlation of the genes with inflammation, hepatotoxicity, and cancer. Molecular validation suggested potential biomarkers for assessing the toxicity of accumulated cAgNPs. Conclusion: Our investigation highlights the risk associated with a single cAgNP exposure with unrepaired damage persisting for at least a month. Keywords: nanotoxicity, liver toxicity, prolonged tissue damage, differentially expressed genes, molecular pathway analysis

Previous research studies on the toxicity of polyhexamethylene guanidine (PHMG) as a humidifier disinfectant majorly focused on lung fibrosis. Considering that disinfectants in humidifiers are released in aerosol form, the eyes are directly exposed and highly vulnerable to the detrimental effects of the PHMG. Therefore, in the present study we investigated the adverse effects of PHMG on the eyes; considering fibrosis as a manifestation of PHMG toxicity in the eye, we evaluated fibrosis-related biomarkers in cultured Statens Seruminstitut Rabbit Cornea (SIRC) cells. Cell viability was measured using 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay, fibrosis-related biomarkers were evaluated through polymerase chain reaction (PCR) and immunoblotting, and oxidative stress was evaluated using 2’,7’-dichlorodihydrofluorescein diacetate (H2DCFDA). Polyhexamethylene guanidine showed cytotoxicity in a time and concentration-dependent manner. Fibrosis related biomarkers including transforming growth factor-β (TGF-β), α-smooth muscle actin (α-SMA), matrix metalloproteinase (MMP), tissue inhibitor of metalloproteinase (TIMP) and hemeoxygenase-1 (HO-1) increased in both gene and protein levels. Oxidative stress also increased in the PHMG-treated cultured cells. The findings of the present study suggest that PHMG could cause toxicity in the eye as manifested by fibrosis.

Disinfectants including polyhexamethylene guanidine phosphate (PHMG) and mixtures of chloromethylisothiazolinone/ methylisothiazolinone (CMIT/MIT) have been widely used in Korea to prevent microbial growth in the humidifier water, which triggered an outbreak of serious respiratory diseases. In addition to the respiratory syndrome, disease-related symptoms including liver toxicity, asthma, and skin allergies were also found after extensive survey of people exposed to the humidifier disinfectants (HDs). In this study, eye irritation tests were performed based on the Organization for economic co-operation and development (OECD) test guidelines 492 using EpiOcularTM which is a tissue model of reconstructed human cornea-like epithelium. As results, the raw materials of PHMG (26% as active ingredient) and CMIT/MIT (1.5% as active ingredient) were classified under UN globally harmonized system of classification and labeling of chemical (GHS) category 1 or category 2. However, aqueous dilutions of raw materials such as market products of HDs that contain 0.13% of PHMG and 0.03% of CMIT/MIT or further dilutions of the market products for humidifier that contain 0.0013% of PHMG and 0.0003% of CMIT/MIT were classified under any category, which suggested absence of eye irritation at the test concentration.

The stability of citrate-capped silver nanoparticles (AgNPs) and the embryonic developmental toxicity were evaluated in the fish test water. Serious aggregation of AgNPs was observed in undiluted fish water (DM-100) in which high concentration of ionic salts exist. However, AgNPs were found to be stable for 7 days in DM-10, prepared by diluting the original fish water (DM-100) with deionized water to 10 %. The normal physiology of zebrafish embryos were evaluated in DM-10 to see if DM-10 can be used as a control vehicle for the embryonic fish toxicity test. As results, DM-10 without AgNPs did not induce any significant adverse effects on embryonic development of zebrafish determined by mortality, hatching, malformations and heart rate. When embryonic toxicity of AgNPs was tested in both DM-10 and in DM-100, AgNPs showed higher toxicity in DM-10 than in DM-100. This means that the big-sized aggregates of AgNPs were low toxic compared to the nano-sized AgNPs. AgNPs induced delayed hatching, decreased heart rate, pericardial edema, and embryo death. Accumulation of AgNPs in the embryo bodies was also observed. Based on this study, citrate-capped AgNPs are not aggregated in DM-10 and it can be used as a control vehicle in the toxicity test of fish embryonic development.

Effects of nanoparticles (NPs) on skin corrosion and irritation using three-dimensional human skin models were investigated based on the test guidelines of Organization for Economic Co-operation and Development (OECD TG431 and TG439). EpiDermTM skin was incubated with NPs including those harboring iron (FeNPs), aluminum oxide (AlNPs), titanium oxide (TNPs), and silver (AgNPs) for a defined time according to the test guidelines. Cell viabilities of EpiDermTM skins were measured by the 3-(4, 5-dimethylthi-azol-2-yl)-2.5-diphenyltetrazolium bromide based method. FeNPs, AlNPs, TNPs, and AgNPs were non-corrosive because the viability was more than 50% after 3 min exposure and more than 15% after 60 min exposure, which are the non-corrosive criteria. All NPs were also non-irritants, based on viability exceeding 50% after 60 min exposure and 42 hr post-incubation. Release of interleukin 1-alpha and histopathological analysis supported the cell viability results. These findings suggest that FeNPs, AlNPs, TNPs, and AgNPs are ‘non-corrosive’ and ‘non-irritant’ to human skin by a globally harmonized classification system.

The biological activity of particles is largely dependent on their size in biological systems. Dispersion in the aqueous phase has been both a critical impediment to and a prerequisite for particle studies. Carbon black has been used as a surrogate to investigate the biological effects of carbonaceous particles. Here, biocompatible methods were established to disperse carbon black into ultrafine and fine particles which are generally distinguished by the small size of 100 nm. Carbon black with a distinct particle size, N330 and N990 were suspended in blood plasma, cell culture media, Krebs-Ringer's solution (KR), or physiological salt solution (PSS). Large clumps were observed in all dispersion preparations; however, sonication improved dispersion - averaged particle sizes for N330 and N990 were 85.0 ± 42.9 and 112.4 ± 67.9 nm, respectively, in plasma; the corresponding sizes in culture media were 84.8 ± 38.4 and 164.1 ± 77.8 nm. However, sonication was not enough to disperse N330 less than 100 nm in either KR or PSS. Application of Tween 80 along with sonication reduced the size of N330 to less than 100 nm, and dispersed N990 larger than 100 nm (73.6 ± 28.8 and 80.1 ± 30.0 nm for N330 and 349.5 ± 161.8 and 399.8 ± 181.1 nm for N990 in KR and PSS, respectively). In contrast, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) exhibited little effect. Electron microscopy confirmed the typical aciniform structure of the carbon arrays; however, zeta potential measurement failed to explain the dispersibility of carbon black. The methods established in this study could disperse carbon black into ultrafine and fine particles, and may serve as a useful model for the study of particle toxicity, particularly size-related effects.

The adverse effects of silver nanoparticles (AgNPs) to human and environment have not been known well, although AgNPs are now widely applicated to consumer products. In this study, we investigated the inflammatory responses including cytokine production and gene expression in mice after a single intratracheal instillation of AgNPs. As results, pro-inflammatory cytokines (IL-1, TNF-a, and IL-6) and Th0 cytokine (IL-2) were progressively increased by the day 28 after a single instillation. The secretion of Th2 type cytokine was more dominant than that of Th1 type cytokine, and the increase of B cell distribution was also observed. But, histopathological changes in lung were observed only at day 1 after instillation. We identified the changes of gene expression induced by AgNPs in lung tissue using microarray. The 261 genes related to inflammation and tissue damages including Saa3, Krt 13, Lor, Krtdap, and Lcn 2 were up-regulated by over 2 fold, while 103 genes including H2-Ea, Chka, BC030476, Heg1 and Hbb-b1 were down-regulated. Based on the data, it is suggested that AgNPs may induce Th2 type dominant inflammatory responses and tissue damage in the lung of mice.

Bioavailability, tissue distribution, blood concentration, and excretion of citrate-coated silver nanoparticles (AgNPs; size, 7.9 ± 0.95 nm by TEM diameter) were investigated. Male SD rats were treated by a single oral or intravenous administration of either 1 or 10 mg/kg AgNPs. Silver concentration of blood was determined at 10 min, and at 1, 2, 4, 8, 24, 48, and 96 h after treatment. Silver in the liver, lungs, and kidneys was also measured at 24 and 96 h after treatment. Excretion of silver nanoparticles via feces and urine was determined at 24 h after treatment. After oral administration, most AgNPs were found in feces, and their blood concentration was very low. This suggests that absorption through the gastrointestinal tract was not good. However, a high level of silver in the blood was detected after tail vein injection. When rats were injected with 1 mg/kg AgNPs, the silver concentration of blood was significantly elevated at 10 min after injection; the level subsequently decreased. In the rats treated with 10 mg/kg AgNPs, the elevated level did not decrease, but was maintained during the experimental period. On the basis of the values of AUC(oral)/AUC(iv), the bioavailability of orally administered AgNPs was 1.2% in the group treated with 1 mg/kg AgNPs and 4.2% in the group treated with 10 mg/kg AgNPs. AgNPs accumulated in the liver, lungs, and kidneys; the accumulated AgNPs were released into the blood stream. AgNP levels in the urine were extremely low compared to the levels in the feces. When rats were injected with AgNPs, these particles were also detected in feces at 24 h after treatment, which suggests bile secretion of AgNPs.

Concentrations of ambient air particles such as total suspended particulates (TSP), particulate matter (PM)₁?, and PM 2.5 , were monitored simultaneously from May to June, 2007, at four locations at Alashan and Beijing in China, and Seoul and Gosan in South Korea. These monitoring sites are in the pathway of Asian dust (AD) storms traveling from China to the Korean peninsula. An AD event was observed in the Korean peninsula on May 25-26. During the sampling period, the average concentrations of TSP and PM10 were each higher than 150 ㎍/㎥ in Alashan and Beijing, but not in Seoul and Gosan. The concentration of TSP in Alashan (where the dust storm originated) on the day of the AD event reached 1824.7 ㎍/㎥, this being the highest concentration of air particles observed during the sampling period. The level was decreased to 15% of this (274.7 ㎍/㎥) in Seoul and to 8.7% (159. 0 ㎍/㎥) in Gosan when the AD was observed two days later in Korea. Gosan, a representative background site in East Asia, displayed the lowest concentration of particles, with this maximal TSP concentration of 159 ㎍/㎥. Inorganic element contents in the air particles were also measured, and the concentrations in the four different regions compared. Concentrations of anthropogenically-enriched heavy metals in PM10 and PM2.5, such as lead, arsenic, cadmium, and mercury, were higher in the metropolitan cities of Beijing and Seoul compared to non-industrialized regions of Alashan and Gosan. However, the concentrations of inorganic soil elements including iron, magnesium, aluminum sodium, and manganese, were higher in Alashan compared to those in other sampling sites.

Citrate-stabilized silver nanoplates (AgNPs) were prepared via a seed-mediated method without surfactants, such as cetyltrimethylammonium bromide (CTAB), in a short amount of time (15 min). Silver seeds with 3–4 nm in diameter were added to a growth solution containing AgNO3, trisodium citrate (TSC) and L-ascorbic acid (AA). The size of the AgNPs depended on the concentration of the silver seed and TSC. The physical properties of the AgNPs were analyzed by transmission electron microscopy (TEM) and by an ultraviolet-visible (UV-vis) spectrophotometer. In addition, we tested the nanotoxicity of AgNPs prepared in TSC solution to the spleen of a rat, and found that AgNPs induced inflammation and white spots on the surface of the spleen.

Magnetite iron nanoparticles have been widely used as contrast agents and in thermal therapy for cancer. However, their adverse effects on human health have not been fully investigated. In this study, iron oxide nanoparticles were prepared using inorganic iron chloride (size: 5.3+/-3.6 nm in phosphate buffered saline, surface charge: 23.14 mV), and their inflammatory responses were investigated. When mice were treated with iron oxide nanoparticles (250 microg/kg, 500 microg/kg, and 1mg/kg) by a single intratracheal instillation, the level of intracellular reduced glutathione (GSH) was decreased in the cells of bronchoalveolar lavage (BAL) fluid. The arrest of cell cycles in G1 phase was observed, but S-phase was significantly decreased. The concentrations of pro-inflammatory cytokines (IL-1, TNF-alpha, and IL-6) were dose-dependently increased at day 1 after instillation in the BAL fluid and in the blood. During the experimental period of 28 days, pro-inflammatory cytokines (IL-1, TNF-alpha, and IL-6), Th0 cytokine (IL-2), Th1 type cytokine (IL-12), Th2 type cytokines (IL-4 and IL-5), TGF-beta, and IgE were also elevated. Expressions of many genes related with inflammation or tissue damage such as heat shock protein, matrix metalloproteinase, tissue inhibitors of metalloproteinases, and serum amyloid A were significantly induced. Formation of microgranuloma, which is one of the indicators for chronic inflammatory response, was observed in the alveolar space. In addition, distribution of B cell and CD8+ T cell in blood lymphocytes was increased at day 28. Based on the result, iron oxide nanoparticles may subchronic induce inflammatory responses via oxidative stress in mice by a single intratracheal instillation.

The appearance of Asian Dust (AD) originating from China and Mongolia during spring each year is a meteorological phenomenon periodically observed in extensive regions of East Asia. According to a previous epidemiological study, AD has adverse effects on both human beings and ecosystems. In this study, we collected total suspension particles (TSP) in the AD period and Non-AD (NAD) period. We extracted organic components from TSP using dichloromethane (DCM), and the polyaromatic hydrocarbons (PAHs) were analyzed. The DCM extracts contained PAHs such as benzo(b)fluoranthene, benzo[g,h,i]perylene, benzo(k)fluoranthene, benzo(a)pyrene, and pyrene. No significant difference was observed in cytotoxicity of the DCM extracts from AD versus NAD when tested on the human bronchial epithelial cells, BEAS-2B. We also examined the toxic mechanisms of AD extracts in cultured BEAS-2B cells and RAW264.7 cells, and in BEAS-2B cells observed increased levels of reactive oxygen species (ROS), decreased glutathione (GSH), and induced caspase-3 activity. Increased expression of oxidative stress-related and inflammation- related genes were also observed in BEAS-2B cells, while nitric oxide (NO) levels were increased in RAW264.7 cells. Taken together, the results suggest that in these cultured cells, AD may induce cytotoxicity through oxidative stress and pro-inflammatory signals.

Platinum nanoparticles (PNPs) are potentially useful for sensing, catalysis, and other applications in the biological and medical sciences. However, toxicity data on the PNPs are very limited. In this study, we prepared PNPs using K(2)PtCl(6), (21 nm in phosphate buffered saline) and tested inflammatory responses in mice after a single intratracheal instillation. The concentrations of pro-inflammatory cytokines (IL-1, TNF-alpha, and IL-6), TH0 cytokine (IL-2), TH1-type cytokine (IL-12), and TH2-type cytokines (IL-4 and IL-5) were increased in broncho-alveolar lavage fluid by PNPs. It was found that the induciton of TH2-type cytokines were higher than TH1-type cytokine on day 28 after instillation. TGF-beta was also significantly increased in broncho-alveolar lavage fluid during the experimental period. IgE level in serum was increased with the increase of B cells distribution. Intracellular level of glutathione (GSH) was diminished by treatment of PNPs. When the distribution of T cell subtype (CD4(+)/CD8(+)) was analyzed, the ratio was decreased. Gene expression of matrix metalloproteinases was found to be significantly increased on day 1. By histopathological examination, cell infiltration of macrophages and neutrophils was observed in lung tissue during the experimental period. Based on these findings, it is suggested that the exposure to PNPs may induce inflammatory responses in mice.

Manganese, an essential trace nutrient in human beings, has been widely used in the steel industry to improve hardness, stiffness, and strength. With the increased applications of manganese compounds, discharge into the environment has rapidly increased and may exert adverse effects on human health. In this study, manganese toxicity was investigated using cultured T98G cells, which are derived from human glioblasts with the ability to differentiate into several different types of neuroglia. Cytotoxicity was shown in manganese-treated groups (100, 200, 400, and 800microM of MnCl(2)), and cell viability was decreased to 58.8% of the control group at 2days after treatment with 800microM of MnCl(2). When cells were treated with manganese for 24h, ROS dose-dependently increased while antioxidant intracellular GSH decreased. With the generation of ROS, the increased activity of caspase-3 was shown, and was followed by chromatin condensation and breakage, which is an indication of the cellular apoptotic process. ROS also triggered pro-inflammatory responses in cultured T98G cells, which were demonstrated by the increased gene expression and protein levels of IL-6 and IL-8.

As the use of carbon fullerene increases in the chemical industry, the concern over its biological and toxicological effects is also increasing. In this study, the suspension of carbon fullerene (C60) in phosphate buffered saline was prepared and toxicity was investigated using cultured RAW 264.7 and in intraperitoneally injected mice, respectively. The average size of carbon fullerene in the suspension was 53.7 ± 26.5 nm when determined by particle size analyzer. Cell viability was significantly decreased by the exposure of carbon fullerene (0.25~2.00 μg/ml) for 96 hrs in the cultured RAW 264.7 cells. Intracellular reduced glutathione (GSH) level was also decreased compared to the level of the non-treated control group during the exposure period, while the level of nitric oxide was increased. When mice were intraperitoneally injected with carbon fullerene, serum cytokine levels of IL-1 and IL-6 were increased with the increased expression of inflammatory genes in peritoneal macrophage and T cell distribution in blood lymphocytes.The results suggested inflammatory responses were induced by carbon fullerene.

Cerium oxide nanoparticles have a high thermodynamic affinity for oxygen and sulfur, which makes them useful in applications such as catalysts, solar cells, and gas sensors. In this study, we investigated the effects of intratracheal instillation of cerium oxide nanoparticles on the inflammatory responses in mice. The number of neutrophils in bronchoaveolar lavage (BAL) fluids was significantly elevated on day 1 after instillation. Inflammatory cytokines, such as interleukin (IL)-1, tumor necrosis factor (TNF)-α, and IL-6, were also increased in BAL fluid and the cytokine increase initiated the differentiation of naive T cells, followed by the induction of Th1-type cytokines [IL-12 and interferon (IFN)-γ] and Th2-type cytokines (IL-4, IL-5, and IL-10). The secretion of Th1-type cytokines was more dominant than that of Th2-type cytokines. The inflammatory responses were maintained for 28 days by a positive feedback stimulation of IFN-γ and IL-10. In the lung, the expression of inflammatory genes was increased in a time-dependent manner, and granuloma formation appeared on day 14 after instillation. This suggests that intratracheal instillation of cerium oxide nanoparticles causes a delayed-type hypersensitivity reaction and lung fibrosis in mice.

Recent publications showed that metal nanoparticles which are made from TiO2, CeO2, Al2O3, CuCl2, AgNO3 and ZnO2 induced oxidative stress and pro-inflammatory effects in cultured cells and the responses seemed to be common toxic pathway of metal nanoparticles to the ultimate toxicity in animals as well as cellular level. In this study, we compared the gene expression induced by two different types of metal oxide nanoparticles, titanium dioxide nanoparticles (TNP) and cerium dioxide nanoparticles (CNP) using microarray analysis. About 50 genes including interleukin 6, interleukin 1, platelet-derived growth factor β, and leukemia inhibitory factor were induced in cultured BEAS-2B cells treated with TNP 40 ppm. When we compared the induction levels of genes in TNP-treated cells to those in CNP-treated cells, the induction levels were very correlated in various gene categories (r = 0.645). This may suggest a possible common toxic mechanism of metal oxide nanoparticles.

Cytotoxicity induced by silver nanoparticles (AgNPs) and the role that oxidative stress plays in this process were demonstrated in human hepatoma cells. Toxicity induced by silver (Ag(+)) ions was studied in parallel using AgNO(3) as the Ag(+) ion source. Using cation exchange treatment, we confirmed that the AgNP solution contained a negligible amount of free Ag(+) ions. Metal-responsive metallothionein 1b (MT1b) mRNA expression was not induced in AgNP-treated cells, while it was induced in AgNO(3)-treated cells. These results indicate that AgNP-treated cells have limited exposure to Ag(+) ions, despite the potential release of Ag(+) ions from AgNPs in cell culture. AgNPs agglomerated in the cytoplasm and nuclei of treated cells, and induced intracellular oxidative stress. AgNPs exhibited cytotoxicity with a potency comparable to that of Ag(+) ions in in vitro cytotoxicity assays. However, the toxicity of AgNPs was prevented by use of the antioxidant N-acetylcysteine, and AgNP-induced DNA damage was also prevented by N-acetylcysteine. AgNO(3) treatment induced oxidative stress-related glutathione peroxidase 1 (GPx1) and catalase expression to a greater extent than AgNP exposure, but treatment with AgNO(3) and AgNPs induced comparable superoxide dismutase 1 (SOD1) expression levels. Our findings suggest that AgNP cytotoxicity is primarily the result of oxidative stress and is independent of the toxicity of Ag(+) ions.

Cerium oxide nanoparticles (size: 30 nm) were prepared by the supercritical synthesis method, Acute oral toxicity and tissue distribution of the nanoparticles were evaluated by a single administration in rats. Oral administration of the nanoparticles to the rats did not lead to death when the animals were treated by a dose of 5 g/kg (high dose) as well as 100 mg/kg (low dose). Abnormal clinical signs, changes in serum biochemistry and hematology were not observed in high-dose treated group compared to the vehicle control group. Lesions in liver, lung and kidney were not observed in high-dose treated group by histopathological examination. Tissue distribution analysis in liver, kidney, spleen, lung, testis and brain was performed on day 1, day 7 and day 14 after treatment. The average values of the accumulated cerium oxide nanoparticles were elevated in all tissues but statistical significance was only shown in lung. Low levels of tissue distributions after a single oral administration seem to be the low bioavailability of the nanoparticles.

Titanium dioxide nanoparticles (TNP) are nanomaterials which have various applications including photocatalysts, cosmetics, and pharmaceuticals because of their high stability, anticorrosiveness, and photocatalytic properties. Induction of cytokines and potential chronic inflammation were investigated in mice treated with TNP (5 mg/kg, 20 mg/kg, and 50 mg/kg) by a single intratracheal instillation. Pro-inflammatory cytokines such as IL-1, TNF-a, and IL-6 were significantly induced in a dose-dependent manner at day 1 after instillation. The levels of Th1-type cytokines (IL-12 and IFN-gamma) and Th2-type cytokines (IL4, IL-5 and IL-10) were also elevated dose-dependently at day 1 and the inflammatory responses were sustained until the remainder of experimental period for 14 days. By the induction of Th2-type cytokines, the increased B cell distributions both in spleen and in blood, and increased IgE production in BAL fluid and serum were observed. In lung tissue, increase of inflammatory proteins (MIP and MCP) and granuloma formation were observed. Furthermore, the expressions of genes related with antigen presentation (H2-T23, H2-T17, H2-K1, and H2-Eb1) and genes related with the induction of chemotaxis of immune cells (Ccl7, Ccl3, Cxcl1, Ccl4, Ccl2) were markedly increased using microarray analysis. From these data, it could be suggested that TNP possibly cause chronic inflammatory diseases through Th2-mediated pathway in mice.

The increased application of engineered carbon nanotubes (CNTs) has also raised the level of public concern regarding possible toxicities caused by exposure to these nanostructures. In this study, pulmonary and systemic immune responses induced by intratracheal instillation of multi-walled carbon nanotubes (MWCNTs) were investigated in mice. Total numbers of immune cells in bronchoalveolar lavage (BAL) fluid were significantly increased in treated groups (5, 20, and 50mg/kg doses of MWCNTs) and the distribution of neutrophils was elevated at day 1 after instillation. Pro-inflammatory cytokines (IL-1, TNF-alpha, IL-6, IL-4, IL-5, IL-10, IL-12, and IFN-gamma) were also increased in a dose-dependent manner, both in BAL fluid and in blood. Most of the cytokines showed the highest levels at day 1 after instillation and then decreased. Th2-type cytokines (IL-4, IL-5, and IL-10) were elevated in the treated group to levels higher than those of the Th1-type cytokines (IL-12 and IFN-gamma). Furthermore, distributions of B cells in spleen and blood were significantly increased at day 1 after instillation, indicating that Th2-type cytokines had activated B cells, causing them to proliferate. Along with the additional numbers of B cells, granuloma formation in the lung tissue and IgE production were also observed, with an intensity dependent on the dose of MWCNTs instilled. Based on these observations, it is suggested that MWCNTs may induce allergic responses in mice through B cell activation and production of IgE.

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 40 microg/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-activated 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.

An analytical method has been developed for measuring chlorostyrenes in fish tissue sample. Extraction of chlorostyrenes from fish tissue was carried out by ultrasonication using acetone/n-hexane (5:2, v/v) mixture. Most of the lipids in the extract were eliminated by freezing-lipid filtration, prior to solid-phase extraction (SPE) cleanup. During freezing-lipid filtration, about 90% of the lipids extracted from the fish samples were easily removed without any significant losses of chlorostyrenes. For purification, SPE using Florisil was used for the rapid and effective cleanup. Quantification was performed using gas chromatography-mass spectrometry in the selected ion monitoring mode. Spiking experiments were carried out to determine the recovery, precision, and limits of detection (LODs) of the method. The overall recovery was above 80% in the spiked fish tissue sample at 10 and 100 ng/g levels, respectively. The detection limits for chlorostyrenes were ranged from 0.05 to 0.1 ng/g. This developed method is demonstrated to give efficient recoveries and LODs for detecting chlorostyrenes spiked into fish tissue with high lipid content.

Mercury is a widespread environmental and industrial pollutant that induces serious adverse effects in both humans and the environment. However, the toxicities and its mechanisms have not been fully elucidated. Among the proposed mechanisms of biological toxicities, the intracellular level of thiol group (-SH) and oxidative stress have been widely studied. In this study, production of reactive oxygen species (ROS) by mercuric chloride (2, 4, 6, and 8 ppm as of mercury) was investigated in cultured human bronchial epithelial cells (BEAS-2B cell line). Exposure of cultured cells to mercuric chloride led to cell death, ROS increase, and cytosolic caspase-3 activation. The ROS increase was related to the decreased level of GSH. Chromatin condensation evaluated by 4',6-diamidino-2-phenylindole (DAPI) staining were also shown in mercury-treated cells and this suggest the apoptotic process of cells by mercuric chloride.

Toxicokinetics of zinc oxide nanoparticles (ZnONP) was studied in rats via a single intravenous (iv) injection and a single oral administration (3 mg/kg or 30 mg/kg), respectively. Blood concentrations of zinc (Zn) were monitored for 7 d and tissue distribution were determined in liver, kidneys, lung, spleen, thymus, brain, and testes. To ascertain the excretion of ZnONP, Zn levels in urine and feces were measured for 7 d. ZnONP were not readily absorbed from the gastrointestinal tract (GIT) after oral administration and were excreted mostly in feces. When the nanoparticles were injected iv to rats at a dose of 30 mg/kg, peak concentration appeared at 5 min but returned to normal range by d 2 (48 h after injection). ZnONP were distributed mainly to liver, kidneys, lung, and spleen, but not to thymus, brain, and testes. The distribution level was significantly decreased to normal by d 7. Feces excretion levels after iv injection supported biliary excretion of ZnONP. In rats injected iv with 30 mg/kg, mitotic figures in hepatocytes were significantly increased and multifocal acute injuries with dark brown pigment were noted in lungs, while no significant damage was observed in rats treated orally with the same dosage.

In this study, the total toxic effects of river water samples were assessed using a series of cell culture bioassays which encompassed different mechanisms, based on specific modes of action. River water samples were collected from three tributaries of the Youngsan River in the western portion of Korea. We confirmed that Youngsan River water was polluted with a complex mixture of estrogenic and dioxin-like compounds. The total toxic effects of the downstream water samples were found to be higher than that of the upstream water samples. In the upstream water samples, total estrogenic activity was measured to be between 0.005 and 0.049 ng-EEQ/l (17beta-estradiol-equivalent concentration) and no CYP1A activity was detected. In the downstream water samples, however, total estrogenic activity was measured to be between 0.021 ng-EEQ/l and 1.918 ng-EEQ/l, and total CYP1A activity was between 0.63 and 3.55 microg-MEQ/l (3-methylcholanthrene-equivalent concentration). When assessed according to a concentration-response curve, downstream water sample extracts exerted dual actions on estrogen receptors, depending on the concentration volume of the samples. The concentration volume range proximal to the original water sample exhibited estrogenic activity, whereas antiestrogenic activity was observed at high concentration volumes (more than 5 times concentration) in the extracts. This study involved a combination of in vitro bioassays, designed to encompass different mechanisms. The bioassays used included the estrogen receptor binding affinity test, E-screen assay, aromatase assay, and EROD assay. These tests provided a great deal of useful information regarding the potency and action modes of estrogenicity and antiestrogenicity inherent in the sampled river water. Although further study is necessary to determine the relationship between toxic responses in in vitro bioassay systems and chronic toxicity in aquatic organisms, our approach is expected to be fairly accurate with regard to the detection of endocrine-disrupting effects in an aquatic environment.

Epidemiologic studies have suggested a close correlation between arsenic exposure and cardiovascular disease. In mechanism studies of heart disease and arsenic exposure, blood vessels, vascular smooth muscle cells, and endothelial cells of the artery have long been thought to be the primary targets in arsenic exposure but there are only a few studies on cardiomyocytes. In this study, to predict more diverse responses of cardiomyocytes to arsenic exposure in the development of heart failure, gene expression profiles of cultured rat cardiomyocytes (H9C2 cell line) were evaluated using the rat whole-genome microarray when a subcytotoxic level of arsenic trioxide was treated. After 24 hr of arsenic 0.5 ppm exposure (As2O3 was used), 405 genes were up-regulated including heme oxygenase-1, and 499 genes were down-regulated including fibroblast growth factor. With the subcytotoxic dose of As2O3, oxidative stress was generated without cell death, and the transcription of stress-related genes such as heme oxygenase-1, glutathione S-transferase, metallothionein, and catalase were significantly increased. Direct measurement of reactive oxygen species using fluorescent dye showed that arsenic caused oxidative stress at the subcytotoxic level. Although no direct comparison was made among different types of cells in this study, it appears that arsenic can cause physiologic adverse reactions at a relatively low level in cardiomyocytes and that cardiomyocytes are also one of the vulnerable targets of heart failure by arsenic compounds.

Gene expression levels of choriogenin, vitellogenin, and estrogen receptor were determined using a reverse transcription-polymerase chain reaction (RT-PCR) technique after exposure to estrogenic chemicals to compare the sensitivities of the biomarkers of endocrine disruption in medaka, Oryzias latipes. Mature male medaka were treated with a single dose of 100 μ/l of 17α-ethinylestradiol, nonylphenol, and bisphenol A for 6 days, then RNA was extracted from the livers of treated fish for RT-PCR. Primers of RT-PCR for choriogenin H and L, and estrogen receptor were synthesized based on previously known cDNA sequences, and primers for vitellogenin I and II were synthesized based on the partial cDNA which was sequenced in this study. When the five biomarker genes were amplified by RT-PCR under the same condition, the mRNA induction level of each gene was elevated with different sensitivities. Conclusively, choriogenin L, which is a precursor of zona radiata protein (ZI-3) with molecular weight of 49 kD, showed the most sensitive gene expression in all the treated groups.

Objectives : Effects of nanoparticles including zinc oxide nanoparticles, titanium oxide nanoparticles, and their mixtures on skin corrosion and irritation were investigated by using in vitro 3D human skin models (KeraSkinTM) and the results were compared to those of an in vivo animal test. Methods : Skin models were incubated with nanoparticles for a definite time period and cell viability was measured by the 3-(4, 5-dimethylthiazol-2-yl)-2.5-diphenyltetrazolium bromide method. Skin corrosion and irritation were identified by the decreased viability based on the pre-determined threshold. Results : Cell viability after exposure to nanomaterial was not decreased to the pre-determined threshold level, which was 15% after 60 minutes exposure in corrosion test and 50% after 45 minutes exposure in the irritation test. IL-1α release and histopathological findings support the results of cell viability test. In vivo test using rabbits also showed non-corrosive and non-irritant results. Conclusions : The findings provide the evidence that zinc oxide nanoparticles, titanium oxide nanoparticles and their mixture are ‘non corrosive’ and ‘non-irritant’ to the human skin by a globally harmonized classification system. In vivo test using animals can be replaced by an alternative in vitro test.

Citrate-capped silver nanoparticles (AgNP) are widely used in industry, consumer products, and medical appliances. However, information on the environmental toxicity and human health is not comprehensive. Further, the physicochemical properties of AgNP make it difficult to test toxicity, as nanosized particles, due to their size, may increase by aggregation or agglomeration in some administration vehicles. In this study, stability of AgNP was investigated in different types of isotonic solutions, which is important for in vitro testing or toxicokinetic studies using intravenous (iv) injection. Size, morphology, zeta potential, and ion formation were investigated in isotonic solutions for the physicochemical characterization of AgNP. Aggregation and precipitation of AgNP were observed in phosphate-buffered saline or 0.9% NaCl, while AgNP were stable without aggregation or precipitation in 5% glucose in isotonic solution. The average size of AgNP in 5% glucose was approximately 10 nm at different temperatures of 10, 25, or 36°C and at varying concentrations from 10 to 1000 ppm. It is noteworthy that this is almost the same size distribution as that in the water-based suspension of AgNP supplied by the manufacturer. Zeta potential ranged from -40 to -60 mV, suggesting that the repulsion forces of AgNP are not disturbed to a sufficient degree to aggregate while osmolarity is in the isotonic range of 290 ± 10 mOsm/kg in 5% glucose solution. Data suggest that AgNP in a 5% glucose solution may be used in the toxicity test via iv injection without adverse consequences in blood.

Blood levels, tissue distributions, and excretion of silver (Ag) were measured in male Sprague-Dawley rats (n = 5) up to 24 h after a single oral administration of silver nanoparticles (AgNP) and silver ions (Ag(+)), respectively. The AUC24hr of Ag(+) was 3.81 ± 0.57 μg/d/ml when rats were treated with a single dose of 20 mg/kg, whereas that of AgNP was 1.58 ± 0.25 μg/d/ml. Tissue distribution of Ag in liver, kidneys, and lungs was higher when Ag(+) was administered compared to AgNP. Orally administered AgNP were predominantly excreted through feces, suggesting low bioavailability. Death or body weight changes were not observed in the Ag(+)- or AgNP-treated groups. However, decreased red blood cell counts, hematocrit, and hemoglobin were found in the Ag(+)-treated groups, while increased platelet counts and mean platelet volume were noted in the AgNP-treated rats. A serum biochemical analysis showed that aspartate aminotransferase (AST) and alanine aminotransferase (ALT) rose significantly following Ag(+) treatment (20 mg/kg). AgNP treatment (2 or 20 mg/kg) also elevated AST, whereas infiltration of mononuclear cells with liver necrosis was found only in the 20 mg/kg Ag(+)-treated animals.

A growing number of studies report that conventional cytotoxicity assays are incompatible with certain nanoparticles (NPs) due to artifacts caused by the distinctive characteristics of NPs. Lactate dehydrogenase (LDH) leakage assays have inadequately detected cytotoxicity of silver nanoparticles (AgNPs), leading to research into the underlying mechanism. When ECV304 endothelial-like umbilical cells were treated with citrate-capped AgNPs (cAgNPs) or bare AgNPs (bAgNPs), the plasma membrane was disrupted, but the LDH leakage assay failed to detect cytotoxicity, indicating interference with the assay by AgNPs. Both cAgNPs and bAgNPs inactivated LDH directly when treated to cell lysate as expected. AgNPs adsorbed LDH and thus LDH, together with AgNPs, was removed from assay reactants during sample preparation, with a resultant underestimation of LDH leakage from cells. cAgNPs, but not bAgNPs, generated reactive oxygen species (ROS), which were successfully scavenged by N-acetylcysteine or ascorbic acid. LDH inhibition by cAgNPs could be restored partially by simultaneous treatment with those antioxidants, suggesting the contribution of ROS to LDH inactivation. Additionally, the composition of the protein corona surrounding AgNPs was identified employing liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. In sum, the LDH leakage assay, a conventional cell viability test method, should be employed with caution when assessing cytotoxicity of AgNPs.

Carbon black (CB) is an industrial chemical with high potential for human exposure. Although the relationship between exposure to particulate matter (PM) and cardiovascular disease is well documented, the risk of adverse cardiovascular effects attributed to CB particles has not been clearly characterized. This study was performed to (1) investigate the effects of CB on cardiovascular system and (2) identify the target tissue or potential biomarkers. Carbon black with a distinct particle size, N330 (ultrafine particle) and N990 (fine particle), was intratracheally instilled into rats at a doses of 1, 3, or 10 mg/kg. Measurements of thrombotic activity and determination of plasma homocysteine levels, cardiac functionality, and inflammatory responses were conducted at 24-h and 1-wk time points. Exposure to N330 accelerated platelet-dependent blood clotting at 10 mg/kg, the highest exposure tested. Unexpectedly, both N330 and N990 led to prolongation of activated partial thromboplastin time (aPTT), whereas these CB particles failed to affect prothrombin time (PT). N990 produced a significant elevation in the level of plasma homocysteine, a well-established etiological factor in cardiovascular diseases. Both N330 and N990 induced apparent inflammation in the lungs; however, both particles failed to initiate systemic inflammation. Neither CB particle produced observable cardiac symptoms as detected by electrocardiography. Taken together, data show CB exposure enhanced the cardiovascular risk by inducing hyperhomocysteinemia and platelet hyperactivity, although these effects may be variable depending on particle size and exposure duration. Homocysteine may be a potential biomarker for cardiovascular toxicity following CB exposure.

Serum kinetics, tissue distribution, and excretion of citrate-coated silver nanoparticles (AgNPs) were investigated in rabbits (n = 4) up to 28 days after a single intravenous injection. Following a single injection of AgNPs, the AUC(last) was reported to be 3.65 ± 0.68 μg·day/ml in 5 mg/kg-treated group and 0.90 ± 0.16 μg·day/ml in 0.5 mg/kg-treated group, respectively. The accumulation of silver was observed in all the tested organs including liver, kidney, spleen, lung, brain, testis, and thymus at 1 day, 7 day, and 28 day of measurement. The liver and spleen seemed to be the major targets because of high accumulation of silver. Excretion via feces and urine was also monitored during the entire experimental period. Unexpectedly, much more excretion of silver occurred via feces than through urine after an intravenous injection, which suggests biliary excretion of AgNPs. General toxicity was analyzed and histopathological changes were also evaluated.

Silver particles are used in various consumer products due to their positive effects, which include sterilization and antibacterial properties. However, it has been reported that silver nanoparticles (AgNPs) have strongly acute toxic effects on various cells. Therefore, the cytotoxicity of AgNPs was investigated, using Escherichia coli as a model organism, from the standpoint of three key metrics (ionic ratio, size, and agglomeration) that are the most relevant physicochemical properties. The findings indicated that cytotoxicity is depressed by the agglomeration of AgNPs. The order of toxic sensitivity was as follows: total Ag concentration > ionic ratio > size, the order of which was inversely related to the extent of agglomeration. Environ. Toxicol. Chem. 2010;29:2154–2160. © 2010 SETAC

Toxicity of nanoparticles depends on many factors including size, shape, chemical composition, surface area, surface charge, and others. In this study, we compared the toxicity of different sized-silver nanoparticles (AgNPs) which are being widely used in consumer products due to its unique antimicrobial activity. When mice were treated with AgNPs 1mg/kg for 14 days by oral administration, small-sized AgNPs (22nm, 42nm, and 71nm) were distributed to the organs including brain, lung, liver, kidney, and testis while large-sized AgNPs (323nm) were not detected in those tissues. The levels of TGF-β in serum were also significantly increased in the treated group of small-sized AgNPs but not in large-sized AgNPs. In addition, B cell distribution was increased in small-sized AgNPs but not in large-sized-AgNPs by the phenotype analysis. However, body weight or in the ratio of organ/body weight were not different between the control group and all the AgNPs-treated groups. The repeated-dose toxicity of AgNPs (42nm) was also investigated in mice by oral administration for 28 days. By the administration of AgNPs (0.25mg/kg, 0.50mg/kg, 1.00mg/kg), adverse impacts on liver and kidney were observed in a high dose-treated group (1.00mg/kg), when determined by blood chemistry and histipathological analysis. Cytokines including IL-1, IL-6, IL-4, IL-10, IL-12, and TGF-β were also increased in a dose-dependent manner by repeated oral administration. In addition, B cell distribution in lymphocyte and IgE production were increased. Based on these results, it is suggested that repeated oral administration of nano-sized AgNPs may cause organ toxicity and inflammatory responses in mice.

Brazilin, the main constituent of Caesalpinia sappan, is an antioxidative substance that has catechol moiety in its chemical structure. Considering the antioxidant-activity of brazilin, it was expected to have protective effects on the toxicities of radical generating chemicals. The incubation of rat hepatocytes with BrCCl3 resulted in significant increase in lipid peroxidation, leakage of cytoplasmic enzymes and cytoplasmic glutathione depletion. The BrCCl3-induced toxicities on hepatocytes were reduced by the treatment of brazilin. Brazilin has been also proved to have a protective effect on the BrCCl3-induced depression of microsomal calcium sequestration activity. These results indicate that brazilin plays a protective role in BrCCl3-induced hepatocyte injury of the rat.

Silver nanoparticles (AgNPs) may induce deleterious effects in aquatic life on environmental release. The hepatotoxicity of AgNPs was assessed in the liver of adult zebrafish, with the aim of studying the roles of oxidative damage and apoptosis. Zebrafish were exposed to an AgNP solution in which free Ag+ ions were absent at the time of treatment. However, the metal-sensitive metallothionein 2 (MT2) mRNA was induced in the liver tissues of AgNP-treated zebrafish, suggesting that Ag+ ions were released from AgNPs after treatment. It is also possible that MT2 mRNA was induced in the liver tissues by AgNP-generated free radicals. A number of cellular alterations including disruption of hepatic cell cords and apoptotic changes were observed in histological analysis of the liver tissues. The levels of malondialdehyde, a byproduct of cellular lipid peroxidation, and total glutathione were increased in the tissues after treatment with AgNPs. The mRNA levels of the oxyradical-scavenging enzymes catalase and glutathione peroxidase 1a were reduced in the tissues. AgNP treatment induced DNA damage, as demonstrated by analysis with the double-strand break marker γ-H2AX and the expression of p53 protein in liver tissues. In addition, the p53-related pro-apoptotic genes Bax, Noxa, and p21 were upregulated after treatment with AgNPs. These data suggest that oxidative stress and apoptosis are associated with AgNP toxicity in the liver of adult zebrafish.

Silver nanoparticles (AgNPs) are widely applied in many household products and medical uses. However, studies on the effects of AgNPs on human health and environmental implications are in the beginning stage. Furthermore, most data on the toxicity of AgNPs have been generated using nanoparticles modified with detergents to prevent agglomeration, which may alter their toxicities. In this study, we studied toxicity using AgNPs prepared by dispersing them in fetal bovine serum (FBS), biocompatible materials. AgNPs (average size; 68.9 nm, concentrations; 0.2, 0.4, 0.8, and 1.6 ppm, exposure time; 24, 48, 72, and 96 h) showed cytotoxicity to cultured RAW264.7 cells by increasing sub G1 fraction, which indicates cellular apoptosis. AgNPs decreased intracellular glutathione level, increased NO secretion, increased TNF-α in protein and gene levels, and increased gene expression of matrix metalloproteinases (MMP-3, MMP-11, and MMP-19). When cells were treated with AgNPs, they were observed in the cytosol of the activated cells, but were not observed in the dead cells. It seemed that AgNPs were ionized in the cells to cause cytotoxicity by a Trojan-horse type mechanism suggested by previously reported studies.

In this study, the potential harmful effect of cerium dioxide (CeO(2)), and titanium dioxide (TiO(2)) nanoparticles on the environment was investigated using Caenorhabditis elegans ecotoxicity tests. Multiple toxic endpoints, such as stress-response gene expression, growth, fertility, and survival, were analyzed in C. elegans, in response to the CeO(2) and TiO(2) exposure. To investigate relationship between sizes of nanoparticles and toxicity, C. elegans were exposed to nanoparticles to the different sizes of nanoparticles (15, 45nm for CeO(2) and 7, 20nm for TiO(2)). An increase in the expression of the cyp35a2 gene, decrease in fertility and survival parameters were observed in the 15 and 45nm of CeO(2) and in the 7nm of TiO(2) nanoparticles exposed to C. elegans. Gene knock-down experiment using RNA interference (RNAi) suggested that physiological level disturbances may be related with the cyp35a2 gene expression. Smaller sized nanoparticles (7nm of TiO(2) and 15nm of CeO(2)) seemed to be more toxic than larger sized ones (20nm of TiO(2) and 45nm of CeO(2)) on the observed toxicity. The size-dependent effect in CeO(2) and TiO(2) nanoparticles-induced toxicity needs to be investigated under more detailed experimental settings with the various sizes of nanoparticles. Further studies on the mechanism by which CeO(2) and TiO(2) nanoparticles affect cyp35a2 gene expression, fertility, and survival are warranted to better understand the CeO(2) and TiO(2) nanoparticles-induced ecotoxicity in C. elegans, as are studies with the causal relationships between these parameters. Overall results suggest that CeO(2) and TiO(2) nanoparticles have a potential for provoking ecotoxicity on C. elegans and the data obtained from this study can comprise a contribution to knowledge of the ecotoxicology of nanoparticles in C. elegans, about which little data are available.